Metagenomics and the protein universe

PubMed Central

Godzik, Adam

2011-01-01

Metagenomics sequencing projects have dramatically increased our knowledge of the protein universe and provided over one-half of currently known protein sequences; they have also introduced a much broader phylogenetic diversity into the protein databases. The full analysis of metagenomic datasets is only beginning, but it has already led to the discovery of thousands of new protein families, likely representing novel functions specific to given environments. At the same time, a deeper analysis of such novel families, including experimental structure determination of some representatives, suggests that most of them represent distant homologs of already characterized protein families, and thus most of the protein diversity present in the new environments are due to functional divergence of the known protein families rather than the emergence of new ones. PMID:21497084

RanBPM: a potential therapeutic target for modulating diverse physiological disorders.

PubMed

Das, Soumyadip; Suresh, Bharathi; Kim, Hyongbum Henry; Ramakrishna, Suresh

2017-12-01

The Ran-binding protein microtubule-organizing center (RanBPM) is a highly conserved nucleocytoplasmic protein involved in a variety of intracellular signaling pathways that control diverse cellular functions. RanBPM interacts with proteins that are linked to various diseases, including Alzheimer's disease (AD), schizophrenia (SCZ), and cancer. In this article, we define the characteristics of the scaffolding protein RanBPM and focus on its interaction partners in diverse physiological disorders, such as neurological diseases, fertility disorders, and cancer. Copyright © 2017 Elsevier Ltd. All rights reserved.

Reprogramming cellular functions with engineered membrane proteins.

PubMed

Arber, Caroline; Young, Melvin; Barth, Patrick

2017-10-01

Taking inspiration from Nature, synthetic biology utilizes and modifies biological components to expand the range of biological functions for engineering new practical devices and therapeutics. While early breakthroughs mainly concerned the design of gene circuits, recent efforts have focused on engineering signaling pathways to reprogram cellular functions. Since signal transduction across cell membranes initiates and controls intracellular signaling, membrane receptors have been targeted by diverse protein engineering approaches despite limited mechanistic understanding of their function. The modular architecture of several receptor families has enabled the empirical construction of chimeric receptors combining domains from distinct native receptors which have found successful immunotherapeutic applications. Meanwhile, progress in membrane protein structure determination, computational modeling and rational design promise to foster the engineering of a broader range of membrane receptor functions. Marrying empirical and rational membrane protein engineering approaches should enable the reprogramming of cells with widely diverse fine-tuned functions. Copyright © 2017 Elsevier Ltd. All rights reserved.

UBXD Proteins: A Family of Proteins with Diverse Functions in Cancer.

PubMed

Rezvani, Khosrow

2016-10-14

The UBXD family is a diverse group of UBX (ubiquitin-regulatory X) domain-containing proteins in mammalian cells. Members of this family contain a UBX domain typically located at the carboxyl-terminal of the protein. In contrast to the UBX domain shared by all members of UBXD family, the amino-terminal domains are diverse and appear to carry out different roles in a subcellular localization-dependent manner. UBXD proteins are principally associated with the endoplasmic reticulum (ER), where they positively or negatively regulate the ER-associated degradation machinery (ERAD). The distinct protein interaction networks of UBXD proteins allow them to have specific functions independent of the ERAD pathway in a cell type- and tissue context-dependent manner. Recent reports have illustrated that a number of mammalian members of the UBXD family play critical roles in several proliferation and apoptosis pathways dysregulated in selected types of cancer. This review covers recent advances that elucidate the therapeutic potential of selected members of the UBXD family that can contribute to tumor growth.

Centrins in unicellular organisms: functional diversity and specialization.

PubMed

Zhang, Yu; He, Cynthia Y

2012-07-01

Centrins (also known as caltractins) are conserved, EF hand-containing proteins ubiquitously found in eukaryotes. Similar to calmodulins, the calcium-binding EF hands in centrins fold into two structurally similar domains separated by an alpha-helical linker region, shaping like a dumbbell. The small size (15-22 kDa) and domain organization of centrins and their functional diversity/specialization make them an ideal system to study protein structure-function relationship. Here, we review the work on centrins with a focus on their structures and functions characterized in unicellular organisms.

A Dominant Conformational Role for Amino Acid Diversity in Minimalist Protein-Protein Interfaces

PubMed Central

Gilbreth, Ryan N.; Esaki, Kaori; Koide, Akiko; Sidhu, Sachdev S.; Koide, Shohei

2008-01-01

Recent studies have shown that highly simplified interaction surfaces consisting of combinations of just two amino acids, Tyr and Ser, exhibit high affinity and specificity. The high functional levels of such minimalist interfaces might thus indicate small contributions of greater amino acid diversity seen in natural interfaces. Toward addressing this issue, we have produced a pair of binding proteins built on the fibronectin type III scaffold, termed “monobodies”. One monobody contains the Tyr/Ser binary-code interface (termed YS) and the other contains an expanded amino acid diversity interface (YSX), but both bind to an identical target, maltose binding protein (MBP). The YSX monobody bound with higher affinity, a slower off rate and a more favorable enthalpic contribution than the YS monobody. High-resolution x-ray crystal structures revealed that both proteins bound to an essentially identical epitope, providing a unique opportunity to directly investigate the role of amino acid diversity in a protein interaction interface. Surprisingly, Tyr still dominates the YSX paratope and the additional amino acid types are primarily used to conformationally optimize contacts made by tyrosines. Scanning mutagenesis showed that while all contacting Tyr side-chains are essential in the YS monobody, the YSX interface was more tolerant to mutations. These results suggest that the conformational, not chemical, diversity of additional types of amino acids provided higher functionality and evolutionary robustness, supporting the dominant role of Tyr and the importance of conformational diversity in forming protein interaction interfaces. PMID:18602117

The poly(C)-binding proteins: a multiplicity of functions and a search for mechanisms.

PubMed Central

Makeyev, Aleksandr V; Liebhaber, Stephen A

2002-01-01

The poly(C) binding proteins (PCBPs) are encoded at five dispersed loci in the mouse and human genomes. These proteins, which can be divided into two groups, hnRNPs K/J and the alphaCPs (alphaCP1-4), are linked by a common evolutionary history, a shared triple KH domain configuration, and by their poly(C) binding specificity. Given these conserved characteristics it is remarkable to find a substantial diversity in PCBP functions. The roles of these proteins in mRNA stabilization, translational activation, and translational silencing suggest a complex and diverse set of post-transcriptional control pathways. Their additional putative functions in transcriptional control and as structural components of important DNA-protein complexes further support their remarkable structural and functional versatility. Clearly the identification of additional binding targets and delineation of corresponding control mechanisms and effector pathways will establish highly informative models for further exploration. PMID:12003487

The poly(C)-binding proteins: a multiplicity of functions and a search for mechanisms.

PubMed

Makeyev, Aleksandr V; Liebhaber, Stephen A

2002-03-01

The poly(C) binding proteins (PCBPs) are encoded at five dispersed loci in the mouse and human genomes. These proteins, which can be divided into two groups, hnRNPs K/J and the alphaCPs (alphaCP1-4), are linked by a common evolutionary history, a shared triple KH domain configuration, and by their poly(C) binding specificity. Given these conserved characteristics it is remarkable to find a substantial diversity in PCBP functions. The roles of these proteins in mRNA stabilization, translational activation, and translational silencing suggest a complex and diverse set of post-transcriptional control pathways. Their additional putative functions in transcriptional control and as structural components of important DNA-protein complexes further support their remarkable structural and functional versatility. Clearly the identification of additional binding targets and delineation of corresponding control mechanisms and effector pathways will establish highly informative models for further exploration.

IA channels: diverse regulatory mechanisms.

PubMed

Carrasquillo, Yarimar; Nerbonne, Jeanne M

2014-04-01

In many peripheral and central neurons, A-type K(+) currents, IA, have been identified and shown to be key determinants in shaping action potential waveforms and repetitive firing properties, as well as in the regulation of synaptic transmission and synaptic plasticity. The functional properties and physiological roles of native neuronal IA, however, have been shown to be quite diverse in different types of neurons. Accumulating evidence suggests that this functional diversity is generated by multiple mechanisms, including the expression and subcellular distributions of IA channels encoded by different voltage-gated K(+) (Kv) channel pore-forming (α) subunits, interactions of Kv α subunits with cytosolic and/or transmembrane accessory subunits and regulatory proteins and post-translational modifications of channel subunits. Several recent reports further suggest that local protein translation in the dendrites of neurons and interactions between IA channels with other types of voltage-gated ion channels further expands the functional diversity of native neuronal IA channels. Here, we review the diverse molecular mechanisms that have been shown or proposed to underlie the functional diversity of native neuronal IA channels.

Function and diversity of P0 proteins among cotton leafroll dwarf virus isolates.

PubMed

Cascardo, Renan S; Arantes, Ighor L G; Silva, Tatiane F; Sachetto-Martins, Gilberto; Vaslin, Maité F S; Corrêa, Régis L

2015-08-12

The RNA silencing pathway is an important anti-viral defense mechanism in plants. As a counter defense, some members of the viral family Luteoviridae are able to evade host immunity by encoding the P0 RNA silencing suppressor protein. Here we explored the functional diversity of P0 proteins among eight cotton leafroll dwarf virus (CLRDV) isolates, a virus associated with a worldwide cotton disease known as cotton blue disease (CBD). CLRDV-infected cotton plants of different varieties were collected from five growing fields in Brazil and their P0 sequences compared to three previously obtained isolates. P0's silencing suppression activities were scored based on transient expression experiments in Nicotiana benthamiana leaves. High sequence diversity was observed among CLRDV P0 proteins, indicating that some isolates found in cotton varieties formerly resistant to CLRDV should be regarded as new genotypes within the species. All tested proteins were able to suppress local and systemic silencing, but with significantly variable degrees. All P0 proteins were able to mediate the decay of ARGONAUTE proteins, a key component of the RNA silencing machinery. The sequence diversity observed in CLRDV P0s is also reflected in their silencing suppression capabilities. However, the strength of local and systemic silencing suppression was not correlated for some proteins.

Site‐Selective Disulfide Modification of Proteins: Expanding Diversity beyond the Proteome

PubMed Central

Kuan, Seah Ling; Wang, Tao

2016-01-01

Abstract The synthetic transformation of polypeptides with molecular accuracy holds great promise for providing functional and structural diversity beyond the proteome. Consequently, the last decade has seen an exponential growth of site‐directed chemistry to install additional features into peptides and proteins even inside living cells. The disulfide rebridging strategy has emerged as a powerful tool for site‐selective modifications since most proteins contain disulfide bonds. In this Review, we present the chemical design, advantages and limitations of the disulfide rebridging reagents, while summarizing their relevance for synthetic customization of functional protein bioconjugates, as well as the resultant impact and advancement for biomedical applications. PMID:27778400

A Prospective Metagenomic and Metabolomic Analysis of the Impact of Exercise and/or Whey Protein Supplementation on the Gut Microbiome of Sedentary Adults.

PubMed

Cronin, Owen; Barton, Wiley; Skuse, Peter; Penney, Nicholas C; Garcia-Perez, Isabel; Murphy, Eileen F; Woods, Trevor; Nugent, Helena; Fanning, Aine; Melgar, Silvia; Falvey, Eanna C; Holmes, Elaine; Cotter, Paul D; O'Sullivan, Orla; Molloy, Michael G; Shanahan, Fergus

2018-01-01

Many components of modern living exert influence on the resident intestinal microbiota of humans with resultant impact on host health. For example, exercise-associated changes in the diversity, composition, and functional profiles of microbial populations in the gut have been described in cross-sectional studies of habitual athletes. However, this relationship is also affected by changes in diet, such as changes in dietary and supplementary protein consumption, that coincide with exercise. To determine whether increasing physical activity and/or increased protein intake modulates gut microbial composition and function, we prospectively challenged healthy but sedentary adults with a short-term exercise regime, with and without concurrent daily whey protein consumption. Metagenomics- and metabolomics-based assessments demonstrated modest changes in gut microbial composition and function following increases in physical activity. Significant changes in the diversity of the gut virome were evident in participants receiving daily whey protein supplementation. Results indicate that improved body composition with exercise is not dependent on major changes in the diversity of microbial populations in the gut. The diverse microbial characteristics previously observed in long-term habitual athletes may be a later response to exercise and fitness improvement. IMPORTANCE The gut microbiota of humans is a critical component of functional development and subsequent health. It is important to understand the lifestyle and dietary factors that affect the gut microbiome and what impact these factors may have. Animal studies suggest that exercise can directly affect the gut microbiota, and elite athletes demonstrate unique beneficial and diverse gut microbiome characteristics. These characteristics are associated with levels of protein consumption and levels of physical activity. The results of this study show that increasing the fitness levels of physically inactive humans leads to modest but detectable changes in gut microbiota characteristics. For the first time, we show that regular whey protein intake leads to significant alterations to the composition of the gut virome.

A dominant conformational role for amino acid diversity in minimalist protein–protein interfaces

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gilbreth, Ryan N.; Esaki, Kaori; Koide, Akiko

Recent studies have shown that highly simplified interaction surfaces consisting of combinations of just two amino acids, Tyr and Ser, exhibit high affinity and specificity. The high functional levels of such minimalist interfaces might thus indicate small contributions of greater amino acid diversity seen in natural interfaces. Toward addressing this issue, we have produced a pair of binding proteins built on the fibronectin type III scaffold, termed “monobodies.” One monobody contains the Tyr/Ser binary-code interface (termed YS) and the other contains an expanded amino acid diversity interface (YSX), but both bind to an identical target, maltose-binding protein. The YSX monobodymore » bound with higher affinity, a slower off rate and a more favorable enthalpic contribution than the YS monobody. High-resolution X-ray crystal structures revealed that both proteins bound to an essentially identical epitope, providing a unique opportunity to directly investigate the role of amino acid diversity in a protein interaction interface. Surprisingly, Tyr still dominates the YSX paratope and the additional amino acid types are primarily used to conformationally optimize contacts made by tyrosines. Scanning mutagenesis showed that while all contacting Tyr side chains are essential in the YS monobody, the YSX interface was more tolerant to mutations. These results suggest that the conformational, not chemical, diversity of additional types of amino acids provided higher functionality and evolutionary robustness, supporting the dominant role of Tyr and the importance of conformational diversity in forming protein interaction interfaces.« less

Evolution of Enzyme Superfamilies: Comprehensive Exploration of Sequence-Function Relationships.

PubMed

Baier, F; Copp, J N; Tokuriki, N

2016-11-22

The sequence and functional diversity of enzyme superfamilies have expanded through billions of years of evolution from a common ancestor. Understanding how protein sequence and functional "space" have expanded, at both the evolutionary and molecular level, is central to biochemistry, molecular biology, and evolutionary biology. Integrative approaches that examine protein sequence, structure, and function have begun to provide comprehensive views of the functional diversity and evolutionary relationships within enzyme superfamilies. In this review, we outline the recent advances in our understanding of enzyme evolution and superfamily functional diversity. We describe the tools that have been used to comprehensively analyze sequence relationships and to characterize sequence and function relationships. We also highlight recent large-scale experimental approaches that systematically determine the activity profiles across enzyme superfamilies. We identify several intriguing insights from this recent body of work. First, promiscuous activities are prevalent among extant enzymes. Second, many divergent proteins retain "function connectivity" via enzyme promiscuity, which can be used to probe the evolutionary potential and history of enzyme superfamilies. Finally, we discuss open questions regarding the intricacies of enzyme divergence, as well as potential research directions that will deepen our understanding of enzyme superfamily evolution.

Quantification of non-coding RNA target localization diversity and its application in cancers.

PubMed

Cheng, Lixin; Leung, Kwong-Sak

2018-04-01

Subcellular localization is pivotal for RNAs and proteins to implement biological functions. The localization diversity of protein interactions has been studied as a crucial feature of proteins, considering that the protein-protein interactions take place in various subcellular locations. Nevertheless, the localization diversity of non-coding RNA (ncRNA) target proteins has not been systematically studied, especially its characteristics in cancers. In this study, we provide a new algorithm, non-coding RNA target localization coefficient (ncTALENT), to quantify the target localization diversity of ncRNAs based on the ncRNA-protein interaction and protein subcellular localization data. ncTALENT can be used to calculate the target localization coefficient of ncRNAs and measure how diversely their targets are distributed among the subcellular locations in various scenarios. We focus our study on long non-coding RNAs (lncRNAs), and our observations reveal that the target localization diversity is a primary characteristic of lncRNAs in different biotypes. Moreover, we found that lncRNAs in multiple cancers, differentially expressed cancer lncRNAs, and lncRNAs with multiple cancer target proteins are prone to have high target localization diversity. Furthermore, the analysis of gastric cancer helps us to obtain a better understanding that the target localization diversity of lncRNAs is an important feature closely related to clinical prognosis. Overall, we systematically studied the target localization diversity of the lncRNAs and uncovered its association with cancer.

Genetic Incorporation of Twelve meta-Substituted Phenylalanine Derivatives Using A Single Pyrrolysyl-tRNA Synthetase

PubMed Central

Wang, Yane-Shih; Fang, Xinqiang; Chen, Hsueh-Ying; Wu, Bo; Wang, Zhiyong U.; Hilty, Christian; Liu, Wenshe R.

2012-01-01

When coexpressed with its cognate amber suppressing tRNACUAPyl, a pyrrolysyl-tRNA synthetase mutant N346A/C348A is able to genetically incorporate twelve meta-substituted phenylalanine derivatives into proteins site-specifically at amber mutation sites in Escherichia coli. These genetically encoded noncanonical amino acids resemble phenylalanine in size and contain diverse bioorthogonal functional groups such as halide, trifluoromethyl, nitrile, nitro, ketone, alkyne, and azide moieties. The genetic installation of these functional groups in proteins provides multiple ways to site-selectively label proteins with biophysical and biochemical probes for their functional investigations. We demonstrate that a genetically incorporated trifluoromethyl group can be used as a sensitive 19F NMR probe to study protein folding/unfolding, and that genetically incorporated reactive functional groups such as ketone, alkyne, and azide moieties can be applied to site-specifically label proteins with florescent probes. This critical discovery allows the synthesis of proteins with diverse bioorthogonal functional groups for a variety of basic studies and biotechnology development using a single recombinant expression system. PMID:23138887

The diverse functions of the hepatitis B core/capsid protein (HBc) in the viral life cycle: Implications for the development of HBc-targeting antivirals.

PubMed

Diab, Ahmed; Foca, Adrien; Zoulim, Fabien; Durantel, David; Andrisani, Ourania

2018-01-01

Virally encoded proteins have evolved to perform multiple functions, and the core protein (HBc) of the hepatitis B virus (HBV) is a perfect example. While HBc is the structural component of the viral nucleocapsid, additional novel functions for the nucleus-localized HBc have recently been described. These results extend for HBc, beyond its structural role, a regulatory function in the viral life cycle and potentially a role in pathogenesis. In this article, we review the diverse roles of HBc in HBV replication and pathogenesis, emphasizing how the unique structure of this protein is key to its various functions. We focus in particular on recent advances in understanding the significance of HBc phosphorylations, its interaction with host proteins and the role of HBc in regulating the transcription of host genes. We also briefly allude to the emerging niche for new direct-acting antivirals targeting HBc, known as Core (protein) Allosteric Modulators (CAMs). Copyright © 2017 Elsevier B.V. All rights reserved.

The many blades of the β-propeller proteins: conserved but versatile.

PubMed

Chen, Cammy K-M; Chan, Nei-Li; Wang, Andrew H-J

2011-10-01

The β-propeller is a highly symmetrical structure with 4-10 repeats of a four-stranded antiparallel β-sheet motif. Although β-propeller proteins with different blade numbers all adopt disc-like shapes, they are involved in a diverse set of functions, and defects in this family of proteins have been associated with human diseases. However, it has remained ambiguous how variations in blade number could alter the function of β-propellers. In addition to the regularly arranged β-propeller topology, a recently discovered β-pinwheel propeller has been found. Here, we review the structural and functional diversity of β-propeller proteins, including β-pinwheels, as well as recent advances in the typical and atypical propeller structures. Copyright © 2011 Elsevier Ltd. All rights reserved.

Diverse Supramolecular Nanofiber Networks Assembled by Functional Low-Complexity Domains.

PubMed

An, Bolin; Wang, Xinyu; Cui, Mengkui; Gui, Xinrui; Mao, Xiuhai; Liu, Yan; Li, Ke; Chu, Cenfeng; Pu, Jiahua; Ren, Susu; Wang, Yanyi; Zhong, Guisheng; Lu, Timothy K; Liu, Cong; Zhong, Chao

2017-07-25

Self-assembling supramolecular nanofibers, common in the natural world, are of fundamental interest and technical importance to both nanotechnology and materials science. Despite important advances, synthetic nanofibers still lack the structural and functional diversity of biological molecules, and the controlled assembly of one type of molecule into a variety of fibrous structures with wide-ranging functional attributes remains challenging. Here, we harness the low-complexity (LC) sequence domain of fused in sarcoma (FUS) protein, an essential cellular nuclear protein with slow kinetics of amyloid fiber assembly, to construct random copolymer-like, multiblock, and self-sorted supramolecular fibrous networks with distinct structural features and fluorescent functionalities. We demonstrate the utilities of these networks in the templated, spatially controlled assembly of ligand-decorated gold nanoparticles, quantum dots, nanorods, DNA origami, and hybrid structures. Owing to the distinguishable nanoarchitectures of these nanofibers, this assembly is structure-dependent. By coupling a modular genetic strategy with kinetically controlled complex supramolecular self-assembly, we demonstrate that a single type of protein molecule can be used to engineer diverse one-dimensional supramolecular nanostructures with distinct functionalities.

Structural and functional diversity of CLAVATA3/ESR (CLE)-like genes from the potato cyst nematode Globodera rostochiensis.

PubMed

Lu, Shun-Wen; Chen, Shiyan; Wang, Jianying; Yu, Hang; Chronis, Demosthenis; Mitchum, Melissa G; Wang, Xiaohong

2009-09-01

Plant CLAVATA3/ESR-related (CLE) peptides have diverse roles in plant growth and development. Here, we report the isolation and functional characterization of five new CLE genes from the potato cyst nematode Globodera rostochiensis. Unlike typical plant CLE peptides that contain a single CLE motif, four of the five Gr-CLE genes encode CLE proteins with multiple CLE motifs. These Gr-CLE genes were found to be specifically expressed within the dorsal esophageal gland cell of nematode parasitic stages, suggesting a role for their encoded proteins in plant parasitism. Overexpression phenotypes of Gr-CLE genes in Arabidopsis mimicked those of plant CLE genes, and Gr-CLE proteins could rescue the Arabidopsis clv3-2 mutant phenotype when expressed within meristems. A short root phenotype was observed when synthetic GrCLE peptides were exogenously applied to roots of Arabidopsis or potato similar to the overexpression of Gr-CLE genes in Arabidopsis and potato hairy roots. These results reveal that G. rostochiensis CLE proteins with either single or multiple CLE motifs function similarly to plant CLE proteins and that CLE signaling components are conserved in both Arabidopsis and potato roots. Furthermore, our results provide evidence to suggest that the evolution of multiple CLE motifs may be an important mechanism for generating functional diversity in nematode CLE proteins to facilitate parasitism.

Molecular Characteristics and Biological Functions of Surface-Active and Surfactant Proteins.

PubMed

Sunde, Margaret; Pham, Chi L L; Kwan, Ann H

2017-06-20

Many critical biological processes take place at hydrophobic:hydrophilic interfaces, and a wide range of organisms produce surface-active proteins and peptides that reduce surface and interfacial tension and mediate growth and development at these boundaries. Microorganisms produce both small lipid-associated peptides and amphipathic proteins that allow growth across water:air boundaries, attachment to surfaces, predation, and improved bioavailability of hydrophobic substrates. Higher-order organisms produce surface-active proteins with a wide variety of functions, including the provision of protective foam environments for vulnerable reproductive stages, evaporative cooling, and gas exchange across airway membranes. In general, the biological functions supported by these diverse polypeptides require them to have an amphipathic nature, and this is achieved by a diverse range of molecular structures, with some proteins undergoing significant conformational change or intermolecular association to generate the structures that are surface active.

TRP channel proteins and signal transduction.

PubMed

Minke, Baruch; Cook, Boaz

2002-04-01

TRP channel proteins constitute a large and diverse family of proteins that are expressed in many tissues and cell types. This family was designated TRP because of a spontaneously occurring Drosophila mutant lacking TRP that responded to a continuous light with a transient receptor potential (hence TRP). In addition to responses to light, TRPs mediate responses to nerve growth factor, pheromones, olfaction, mechanical, chemical, temperature, pH, osmolarity, vasorelaxation of blood vessels, and metabolic stress. Furthermore, mutations in several members of TRP-related channel proteins are responsible for several diseases, such as several tumors and neurodegenerative disorders. TRP-related channel proteins are found in a variety of organisms, tissues, and cell types, including nonexcitable, smooth muscle, and neuronal cells. The large functional diversity of TRPs is also reflected in their diverse permeability to ions, although, in general, they are classified as nonselective cationic channels. The molecular domains that are conserved in all members of the TRP family constitute parts of the transmembrane domains and in most members also the ankyrin-like repeats at the NH2 terminal of the protein and a "TRP domain" at the COOH terminal, which is a highly conserved 25-amino acid stretch with still unknown function. All of the above features suggest that members of the TRP family are "special assignment" channels, which are recruited to diverse signaling pathways. The channels' roles and characteristics such as gating mechanism, regulation, and permeability are determined by evolution according to the specific functional requirements.

Structure-Based Phylogenetic Analysis of the Lipocalin Superfamily.

PubMed

Lakshmi, Balasubramanian; Mishra, Madhulika; Srinivasan, Narayanaswamy; Archunan, Govindaraju

2015-01-01

Lipocalins constitute a superfamily of extracellular proteins that are found in all three kingdoms of life. Although very divergent in their sequences and functions, they show remarkable similarity in 3-D structures. Lipocalins bind and transport small hydrophobic molecules. Earlier sequence-based phylogenetic studies of lipocalins highlighted that they have a long evolutionary history. However the molecular and structural basis of their functional diversity is not completely understood. The main objective of the present study is to understand functional diversity of the lipocalins using a structure-based phylogenetic approach. The present study with 39 protein domains from the lipocalin superfamily suggests that the clusters of lipocalins obtained by structure-based phylogeny correspond well with the functional diversity. The detailed analysis on each of the clusters and sub-clusters reveals that the 39 lipocalin domains cluster based on their mode of ligand binding though the clustering was performed on the basis of gross domain structure. The outliers in the phylogenetic tree are often from single member families. Also structure-based phylogenetic approach has provided pointers to assign putative function for the domains of unknown function in lipocalin family. The approach employed in the present study can be used in the future for the functional identification of new lipocalin proteins and may be extended to other protein families where members show poor sequence similarity but high structural similarity.

Functional anthology of intrinsic disorder. 3. Ligands, post-translational modifications, and diseases associated with intrinsically disordered proteins.

PubMed

Xie, Hongbo; Vucetic, Slobodan; Iakoucheva, Lilia M; Oldfield, Christopher J; Dunker, A Keith; Obradovic, Zoran; Uversky, Vladimir N

2007-05-01

Currently, the understanding of the relationships between function, amino acid sequence, and protein structure continues to represent one of the major challenges of the modern protein science. As many as 50% of eukaryotic proteins are likely to contain functionally important long disordered regions. Many proteins are wholly disordered but still possess numerous biologically important functions. However, the number of experimentally confirmed disordered proteins with known biological functions is substantially smaller than their actual number in nature. Therefore, there is a crucial need for novel bionformatics approaches that allow projection of the current knowledge from a few experimentally verified examples to much larger groups of known and potential proteins. The elaboration of a bioinformatics tool for the analysis of functional diversity of intrinsically disordered proteins and application of this data mining tool to >200 000 proteins from the Swiss-Prot database, each annotated with at least one of the 875 functional keywords, was described in the first paper of this series (Xie, H.; Vucetic, S.; Iakoucheva, L. M.; Oldfield, C. J.; Dunker, A. K.; Obradovic, Z.; Uversky, V.N. Functional anthology of intrinsic disorder. 1. Biological processes and functions of proteins with long disordered regions. J. Proteome Res. 2007, 5, 1882-1898). Using this tool, we have found that out of the 710 Swiss-Prot functional keywords associated with at least 20 proteins, 262 were strongly positively correlated with long intrinsically disordered regions, and 302 were strongly negatively correlated. Illustrative examples of functional disorder or order were found for the vast majority of keywords showing strongest positive or negative correlation with intrinsic disorder, respectively. Some 80 Swiss-Prot keywords associated with disorder- and order-driven biological processes and protein functions were described in the first paper (see above). The second paper of the series was devoted to the presentation of 87 Swiss-Prot keywords attributed to the cellular components, domains, technical terms, developmental processes, and coding sequence diversities possessing strong positive and negative correlation with long disordered regions (Vucetic, S.; Xie, H.; Iakoucheva, L. M.; Oldfield, C. J.; Dunker, A. K.; Obradovic, Z.; Uversky, V. N. Functional anthology of intrinsic disorder. 2. Cellular components, domains, technical terms, developmental processes, and coding sequence diversities correlated with long disordered regions. J. Proteome Res. 2007, 5, 1899-1916). Protein structure and functionality can be modulated by various post-translational modifications or/and as a result of binding of specific ligands. Numerous human diseases are associated with protein misfolding/misassembly/misfunctioning. This work concludes the series of papers dedicated to the functional anthology of intrinsic disorder and describes approximately 80 Swiss-Prot functional keywords that are related to ligands, post-translational modifications, and diseases possessing strong positive or negative correlation with the predicted long disordered regions in proteins.

Rational protein design: developing next-generation biological therapeutics and nanobiotechnological tools.

PubMed

Wilson, Corey J

2015-01-01

Proteins are the most functionally diverse macromolecules observed in nature, participating in a broad array of catalytic, biosensing, transport, scaffolding, and regulatory functions. Fittingly, proteins have become one of the most promising nanobiotechnological tools to date, and through the use of recombinant DNA and other laboratory methods we have produced a vast number of biological therapeutics derived from human genes. Our emerging ability to rationally design proteins (e.g., via computational methods) holds the promise of significantly expanding the number and diversity of protein therapies and has opened the gateway to realizing true and uncompromised personalized medicine. In the last decade computational protein design has been transformed from a set of fundamental strategies to stringently test our understanding of the protein structure-function relationship, to practical tools for developing useful biological processes, nano-devices, and novel therapeutics. As protein design strategies improve (i.e., in terms of accuracy and efficiency) clinicians will be able to leverage individual genetic data and biological metrics to develop and deliver personalized protein therapeutics with minimal delay. © 2014 Wiley Periodicals, Inc.

Post-translational processing targets functionally diverse proteins in Mycoplasma hyopneumoniae

PubMed Central

Tacchi, Jessica L.; Raymond, Benjamin B. A.; Haynes, Paul A.; Berry, Iain J.; Widjaja, Michael; Bogema, Daniel R.; Woolley, Lauren K.; Jenkins, Cheryl; Minion, F. Chris; Padula, Matthew P.; Djordjevic, Steven P.

2016-01-01

Mycoplasma hyopneumoniae is a genome-reduced, cell wall-less, bacterial pathogen with a predicted coding capacity of less than 700 proteins and is one of the smallest self-replicating pathogens. The cell surface of M. hyopneumoniae is extensively modified by processing events that target the P97 and P102 adhesin families. Here, we present analyses of the proteome of M. hyopneumoniae-type strain J using protein-centric approaches (one- and two-dimensional GeLC–MS/MS) that enabled us to focus on global processing events in this species. While these approaches only identified 52% of the predicted proteome (347 proteins), our analyses identified 35 surface-associated proteins with widely divergent functions that were targets of unusual endoproteolytic processing events, including cell adhesins, lipoproteins and proteins with canonical functions in the cytosol that moonlight on the cell surface. Affinity chromatography assays that separately used heparin, fibronectin, actin and host epithelial cell surface proteins as bait recovered cleavage products derived from these processed proteins, suggesting these fragments interact directly with the bait proteins and display previously unrecognized adhesive functions. We hypothesize that protein processing is underestimated as a post-translational modification in genome-reduced bacteria and prokaryotes more broadly, and represents an important mechanism for creating cell surface protein diversity. PMID:26865024

Metaproteome of the viral concentrates from the deep chlorophyll maximum of the South China Sea

NASA Astrophysics Data System (ADS)

Xie, Zhang-Xian; Chen, Feng; Zhang, Shu-Feng; Wang, Ming-Hua; Zhang, Hao; Kong, Ling-Fen; Dai, Min-Han; Hong, Hua-Sheng; Lin, Lin; Wang, Da-Zhi

2016-04-01

Viral concentrates (VCs) have been commonly used for studying viral diversity, viral metagenomics and virus-host interactions in the natural ecosystem. However, the protein characteristics of VCs have not been explored. Here, we applied shotgun proteomics to characterize the proteins of VCs collected from the oligotrophic deep chlorophyll maximum of the South China Sea. We found that 34% of the identified proteins were assigned to the viruses, mainly being those of SAR11 related bacteria, cyanobacteria and picophytoeukaryotes. The remaining 66% were non-viral proteins mostly originating from diverse bacteria, such as SAR324, SAR11 and the Alteromonadales, and were functionally dominated by transport, translation, sulfur metabolism and one-carbon metabolism. Among the non-viral proteins, 28% were extracellular proteins and 10% were identified exclusively in the VCs, suggesting that non-viral entities might exist in the VCs. This study demonstrated that metaproteomics provides a valuable avenue to explore not only the diversity and structure of a viral community but also the novel ecological functions affiliated with microbes in the natural environment.

Regulation and physiological functions of mammalian phospholipase C.

PubMed

Nakamura, Yoshikazu; Fukami, Kiyoko

2017-04-01

Phospholipase C (PLC) is a key enzyme in phosphoinositide metabolism. PLC hydrolyses phosphatidylinositol 4,5-bis-phosphate to generate two second messengers, inositol 1,4,5-trisphosphate and diacylglycerol, that generate diverse cellular responses. PLC is activated by various signalling molecules, including Ca2+, heterometric G proteins, small G proteins, and receptor/non-receptor tyrosine kinases. In addition to their enzymatic activity, some PLC subtypes also function as a guanine nucleotide exchange factor, GTPase-activating protein, and adaptor protein, independent of their lipase activity. There are 13 PLC isozymes in mammals, and they are categorized into six classes based on structure. Generation and analysis of genetically modified mice has revealed the unexpectedly diverse physiological functions of PLC isozymes. Although all PLC isozymes catalyze the same reaction, each PLC isozyme has unique physiological functions. This review focuses on the regulation and physiological functions of PLCs. © The Authors 2017. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.

Structural and Phylogenetic Analysis of a Conserved Actinobacteria-Specific Protein (ASP1; SCO1997) from Streptomyces Coelicolor

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gao, B.; Sugiman-Marangos, S; Junop, M

2009-01-01

The Actinobacteria phylum represents one of the largest and most diverse groups of bacteria, encompassing many important and well-characterized organisms including Streptomyces, Bifidobacterium, Corynebacterium and Mycobacterium. Members of this phylum are remarkably diverse in terms of life cycle, morphology, physiology and ecology. Recent comparative genomic analysis of 19 actinobacterial species determined that only 5 genes of unknown function uniquely define this large phylum [1]. The cellular functions of these actinobacteria-specific proteins (ASP) are not known.

Functional Anthology of Intrinsic Disorder. III. Ligands, Postranslational Modifications and Diseases Associated with Intrinsically Disordered Proteins

PubMed Central

Xie, Hongbo; Vucetic, Slobodan; Iakoucheva, Lilia M.; Oldfield, Christopher J.; Dunker, A. Keith; Obradovic, Zoran; Uversky, Vladimir N.

2008-01-01

Currently, the understanding of the relationships between function, amino acid sequence and protein structure continues to represent one of the major challenges of the modern protein science. As much as 50% of eukaryotic proteins are likely to contain functionally important long disordered regions. Many proteins are wholly disordered but still possess numerous biologically important functions. However, the number of experimentally confirmed disordered proteins with known biological functions is substantially smaller than their actual number in nature. Therefore, there is a crucial need for novel bioinformatics approaches that allow projection of the current knowledge from a few experimentally verified examples to much larger groups of known and potential proteins. The elaboration of a bioinformatics tool for the analysis of functional diversity of intrinsically disordered proteins and application of this data mining tool to >200,000 proteins from Swiss-Prot database, each annotated with at least one of the 875 functional keywords was described in the first paper of this series (Xie H., Vucetic S., Iakoucheva L.M., Oldfield C.J., Dunker A.K., Obradovic Z., Uversky V.N. (2006) Functional anthology of intrinsic disorder. I. Biological processes and functions of proteins with long disordered regions. J. Proteome Res.). Using this tool, we have found that out of the 711 Swiss-Prot functional keywords associated with at least 20 proteins, 262 were strongly positively correlated with long intrinsically disordered regions, and 302 were strongly negatively correlated. Illustrative examples of functional disorder or order were found for the vast majority of keywords showing strongest positive or negative correlation with intrinsic disorder, respectively. Some 80 Swiss-Prot keywords associated with disorder- and order-driven biological processes and protein functions were described in the first paper (Xie H., Vucetic S., Iakoucheva L.M., Oldfield C.J., Dunker A.K., Obradovic Z., Uversky V.N. (2006) Functional anthology of intrinsic disorder. I. Biological processes and functions of proteins with long disordered regions. J. Proteome Res.). The second paper of the series was devoted to the presentation of 87 Swiss-Prot keywords attributed to the cellular components, domains, technical terms, developmental processes and coding sequence diversities possessing strong positive and negative correlation with long disordered regions (Vucetic S., Xie H., Iakoucheva L.M., Oldfield C.J., Dunker A.K., Obradovic Z., Uversky V.N. (2006) Functional anthology of intrinsic disorder. II. Cellular components, domains, technical terms, developmental processes and coding sequence diversities correlated with long disordered regions. J. Proteome Res.). Protein structure and functionality can be modulated by various posttranslational modifications or/and as a result of binding of specific ligands. Numerous human diseases are associated with protein misfolding/misassembly/ misfunctioning. This work concludes the series of papers dedicated to the functional anthology of intrinsic disorder and describes ~80 Swiss-Prot functional keywords that are related to ligands, posttranslational modifications and diseases possessing strong positive or negative correlation with the predicted long disordered regions in proteins. PMID:17391016

New Insight Into the Diversity of SemiSWEET Sugar Transporters and the Homologs in Prokaryotes

PubMed Central

Jia, Baolei; Hao, Lujiang; Xuan, Yuan Hu; Jeon, Che Ok

2018-01-01

Sugars will eventually be exported transporters (SWEETs) and SemiSWEETs represent a family of sugar transporters in eukaryotes and prokaryotes, respectively. SWEETs contain seven transmembrane helices (TMHs), while SemiSWEETs contain three. The functions of SemiSWEETs are less studied. In this perspective article, we analyzed the diversity and conservation of SemiSWEETs and further proposed the possible functions. 1,922 SemiSWEET homologs were retrieved from the UniProt database, which is not proportional to the sequenced prokaryotic genomes. However, these proteins are very diverse in sequences and can be classified into 19 clusters when >50% sequence identity is required. Moreover, a gene context analysis indicated that several SemiSWEETs are located in the operons that are related to diverse carbohydrate metabolism. Several proteins with seven TMHs can be found in bacteria, and sequence alignment suggested that these proteins in bacteria may be formed by the duplication and fusion. Multiple sequence alignments showed that the amino acids for sugar translocation are still conserved and coevolved, although the sequences show diversity. Among them, the functions of a few amino acids are still not clear. These findings highlight the challenges that exist in SemiSWEETs and provide future researchers the foundation to explore these uncharted areas. PMID:29872447

New Insight Into the Diversity of SemiSWEET Sugar Transporters and the Homologs in Prokaryotes.

PubMed

Jia, Baolei; Hao, Lujiang; Xuan, Yuan Hu; Jeon, Che Ok

2018-01-01

Sugars will eventually be exported transporters (SWEETs) and SemiSWEETs represent a family of sugar transporters in eukaryotes and prokaryotes, respectively. SWEETs contain seven transmembrane helices (TMHs), while SemiSWEETs contain three. The functions of SemiSWEETs are less studied. In this perspective article, we analyzed the diversity and conservation of SemiSWEETs and further proposed the possible functions. 1,922 SemiSWEET homologs were retrieved from the UniProt database, which is not proportional to the sequenced prokaryotic genomes. However, these proteins are very diverse in sequences and can be classified into 19 clusters when >50% sequence identity is required. Moreover, a gene context analysis indicated that several SemiSWEETs are located in the operons that are related to diverse carbohydrate metabolism. Several proteins with seven TMHs can be found in bacteria, and sequence alignment suggested that these proteins in bacteria may be formed by the duplication and fusion. Multiple sequence alignments showed that the amino acids for sugar translocation are still conserved and coevolved, although the sequences show diversity. Among them, the functions of a few amino acids are still not clear. These findings highlight the challenges that exist in SemiSWEETs and provide future researchers the foundation to explore these uncharted areas.

A searchable, whole genome resource designed for protein variant analysis in diverse lineages of U.S. beef cattle

USDA-ARS?s Scientific Manuscript database

A key feature of a gene's function is the variety of protein isoforms it encodes in a population. However, the genetic diversity in bovine whole genome databases tends to be underrepresented because these databases contain an abundance of sequence from the most influential sires. Our first aim was ...

Protein relative abundance patterns associated with sucrose-induced dysbiosis are conserved across taxonomically diverse oral microcosm biofilm models of dental caries.

PubMed

Rudney, Joel D; Jagtap, Pratik D; Reilly, Cavan S; Chen, Ruoqiong; Markowski, Todd W; Higgins, LeeAnn; Johnson, James E; Griffin, Timothy J

2015-12-19

The etiology of dental caries is multifactorial, but frequent consumption of free sugars, notably sucrose, appears to be a major factor driving the supragingival microbiota in the direction of dysbiosis. Recent 16S rRNA-based studies indicated that caries-associated communities were less diverse than healthy supragingival plaque but still displayed considerable taxonomic diversity between individuals. Metagenomic studies likewise have found that healthy oral sites from different people were broadly similar with respect to gene function, even though there was an extensive individual variation in their taxonomic profiles. That pattern may also extend to dysbiotic communities. In that case, shifts in community-wide protein relative abundance might provide better biomarkers of dysbiosis that can be achieved through taxonomy alone. In this study, we used a paired oral microcosm biofilm model of dental caries to investigate differences in community composition and protein relative abundance in the presence and absence of sucrose. This approach provided large quantities of protein, which facilitated deep metaproteomic analysis. Community composition was evaluated using 16S rRNA sequencing and metaproteomic approaches. Although taxonomic diversity was reduced by sucrose pulsing, considerable inter-subject variation in community composition remained. By contrast, functional analysis using the SEED ontology found that sucrose induced changes in protein relative abundance patterns for pathways involving glycolysis, lactate production, aciduricity, and ammonia/glutamate metabolism that were conserved across taxonomically diverse dysbiotic oral microcosm biofilm communities. Our findings support the concept of using function-based changes in protein relative abundance as indicators of dysbiosis. Our microcosm model cannot replicate all aspects of the oral environment, but the deep level of metaproteomic analysis it allows makes it suitable for discovering which proteins are most consistently abundant during dysbiosis. It then may be possible to define biomarkers that could be used to detect at-risk tooth surfaces before the development of overt carious lesions.

Phytochemicals perturb membranes and promiscuously alter protein function.

PubMed

Ingólfsson, Helgi I; Thakur, Pratima; Herold, Karl F; Hobart, E Ashley; Ramsey, Nicole B; Periole, Xavier; de Jong, Djurre H; Zwama, Martijn; Yilmaz, Duygu; Hall, Katherine; Maretzky, Thorsten; Hemmings, Hugh C; Blobel, Carl; Marrink, Siewert J; Koçer, Armağan; Sack, Jon T; Andersen, Olaf S

2014-08-15

A wide variety of phytochemicals are consumed for their perceived health benefits. Many of these phytochemicals have been found to alter numerous cell functions, but the mechanisms underlying their biological activity tend to be poorly understood. Phenolic phytochemicals are particularly promiscuous modifiers of membrane protein function, suggesting that some of their actions may be due to a common, membrane bilayer-mediated mechanism. To test whether bilayer perturbation may underlie this diversity of actions, we examined five bioactive phenols reported to have medicinal value: capsaicin from chili peppers, curcumin from turmeric, EGCG from green tea, genistein from soybeans, and resveratrol from grapes. We find that each of these widely consumed phytochemicals alters lipid bilayer properties and the function of diverse membrane proteins. Molecular dynamics simulations show that these phytochemicals modify bilayer properties by localizing to the bilayer/solution interface. Bilayer-modifying propensity was verified using a gramicidin-based assay, and indiscriminate modulation of membrane protein function was demonstrated using four proteins: membrane-anchored metalloproteases, mechanosensitive ion channels, and voltage-dependent potassium and sodium channels. Each protein exhibited similar responses to multiple phytochemicals, consistent with a common, bilayer-mediated mechanism. Our results suggest that many effects of amphiphilic phytochemicals are due to cell membrane perturbations, rather than specific protein binding.

Phytochemicals Perturb Membranes and Promiscuously Alter Protein Function

PubMed Central

2015-01-01

A wide variety of phytochemicals are consumed for their perceived health benefits. Many of these phytochemicals have been found to alter numerous cell functions, but the mechanisms underlying their biological activity tend to be poorly understood. Phenolic phytochemicals are particularly promiscuous modifiers of membrane protein function, suggesting that some of their actions may be due to a common, membrane bilayer-mediated mechanism. To test whether bilayer perturbation may underlie this diversity of actions, we examined five bioactive phenols reported to have medicinal value: capsaicin from chili peppers, curcumin from turmeric, EGCG from green tea, genistein from soybeans, and resveratrol from grapes. We find that each of these widely consumed phytochemicals alters lipid bilayer properties and the function of diverse membrane proteins. Molecular dynamics simulations show that these phytochemicals modify bilayer properties by localizing to the bilayer/solution interface. Bilayer-modifying propensity was verified using a gramicidin-based assay, and indiscriminate modulation of membrane protein function was demonstrated using four proteins: membrane-anchored metalloproteases, mechanosensitive ion channels, and voltage-dependent potassium and sodium channels. Each protein exhibited similar responses to multiple phytochemicals, consistent with a common, bilayer-mediated mechanism. Our results suggest that many effects of amphiphilic phytochemicals are due to cell membrane perturbations, rather than specific protein binding. PMID:24901212

Prokaryotic cytoskeletons: protein filaments organizing small cells.

PubMed

Wagstaff, James; Löwe, Jan

2018-04-01

Most, if not all, bacterial and archaeal cells contain at least one protein filament system. Although these filament systems in some cases form structures that are very similar to eukaryotic cytoskeletons, the term 'prokaryotic cytoskeletons' is used to refer to many different kinds of protein filaments. Cytoskeletons achieve their functions through polymerization of protein monomers and the resulting ability to access length scales larger than the size of the monomer. Prokaryotic cytoskeletons are involved in many fundamental aspects of prokaryotic cell biology and have important roles in cell shape determination, cell division and nonchromosomal DNA segregation. Some of the filament-forming proteins have been classified into a small number of conserved protein families, for example, the almost ubiquitous tubulin and actin superfamilies. To understand what makes filaments special and how the cytoskeletons they form enable cells to perform essential functions, the structure and function of cytoskeletal molecules and their filaments have been investigated in diverse bacteria and archaea. In this Review, we bring these data together to highlight the diverse ways that linear protein polymers can be used to organize other molecules and structures in bacteria and archaea.

The Protein Interactome of Mycobacteriophage Giles Predicts Functions for Unknown Proteins.

PubMed

Mehla, Jitender; Dedrick, Rebekah M; Caufield, J Harry; Siefring, Rachel; Mair, Megan; Johnson, Allison; Hatfull, Graham F; Uetz, Peter

2015-08-01

Mycobacteriophages are viruses that infect mycobacterial hosts and are prevalent in the environment. Nearly 700 mycobacteriophage genomes have been completely sequenced, revealing considerable diversity and genetic novelty. Here, we have determined the protein complement of mycobacteriophage Giles by mass spectrometry and mapped its genome-wide protein interactome to help elucidate the roles of its 77 predicted proteins, 50% of which have no known function. About 22,000 individual yeast two-hybrid (Y2H) tests with four different Y2H vectors, followed by filtering and retest screens, resulted in 324 reproducible protein-protein interactions, including 171 (136 nonredundant) high-confidence interactions. The complete set of high-confidence interactions among Giles proteins reveals new mechanistic details and predicts functions for unknown proteins. The Giles interactome is the first for any mycobacteriophage and one of just five known phage interactomes so far. Our results will help in understanding mycobacteriophage biology and aid in development of new genetic and therapeutic tools to understand Mycobacterium tuberculosis. Mycobacterium tuberculosis causes over 9 million new cases of tuberculosis each year. Mycobacteriophages, viruses of mycobacterial hosts, hold considerable potential to understand phage diversity, evolution, and mycobacterial biology, aiding in the development of therapeutic tools to control mycobacterial infections. The mycobacteriophage Giles protein-protein interaction network allows us to predict functions for unknown proteins and shed light on major biological processes in phage biology. For example, Giles gp76, a protein of unknown function, is found to associate with phage packaging and maturation. The functions of mycobacteriophage-derived proteins may suggest novel therapeutic approaches for tuberculosis. Our ORFeome clone set of Giles proteins and the interactome data will be useful resources for phage interactomics. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

HnRNP-like proteins as post-transcriptional regulators.

PubMed

Yeap, Wan-Chin; Namasivayam, Parameswari; Ho, Chai-Ling

2014-10-01

Plant cells contain a diverse repertoire of RNA-binding proteins (RBPs) that coordinate a network of post-transcriptional regulation. RBPs govern diverse developmental processes by modulating the gene expression of specific transcripts. Recent gene annotation and RNA sequencing clearly showed that heterogeneous nuclear ribonucleoprotein (hnRNP)-like proteins which form a family of RBPs, are also expressed in higher plants and serve specific plant functions. In addition to their involvement in post-transcriptional regulation from mRNA capping to translation, they are also involved in telomere regulation, gene silencing and regulation in chloroplast. Here, we review the involvement of plant hnRNP-like proteins in post-transcription regulation of RNA processes and their functional roles in control of plant developmental processes especially plant-specific functions including flowering, chloroplastic-specific mRNA regulation, long-distance phloem transportation and plant responses to environmental stresses. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Functional expansion of human tRNA synthetases achieved by structural inventions

PubMed Central

Guo, Min; Schimmel, Paul; Yang, Xiang-Lei

2010-01-01

Known as an essential component of the translational apparatus, the aminoacyl-tRNA synthetase family catalyzes the first step reaction in protein synthesis, that is, to specifically attach each amino acid to its cognate tRNA. While preserving this essential role, tRNA synthetases developed other roles during evolution. Human tRNA synthetases, in particular, have diverse functions in different pathways involving angiogenesis, inflammation and apoptosis. The functional diversity is further illustrated in the association with various diseases through genetic mutations that do not affect aminoacylation or protein synthesis. Here we review the accumulated knowledge on how human tRNA synthetases used structural inventions to achieve functional expansions. PMID:19932696

Archaeal Viruses: Diversity, Replication, and Structure.

PubMed

Dellas, Nikki; Snyder, Jamie C; Bolduc, Benjamin; Young, Mark J

2014-11-01

The Archaea-and their viruses-remain the most enigmatic of life's three domains. Once thought to inhabit only extreme environments, archaea are now known to inhabit diverse environments. Even though the first archaeal virus was described over 40 years ago, only 117 archaeal viruses have been discovered to date. Despite this small number, these viruses have painted a portrait of enormous morphological and genetic diversity. For example, research centered around the various steps of the archaeal virus life cycle has led to the discovery of unique mechanisms employed by archaeal viruses during replication, maturation, and virion release. In many instances, archaeal virus proteins display very low levels of sequence homology to other proteins listed in the public database, and therefore, structural characterization of these proteins has played an integral role in functional assignment. These structural studies have not only provided insights into structure-function relationships but have also identified links between viruses across all three domains of life.

Functional characterization of the late embryogenesis abundant (LEA) protein gene family from Pinus tabuliformis (Pinaceae) in Escherichia coli.

PubMed

Gao, Jie; Lan, Ting

2016-01-19

Late embryogenesis abundant (LEA) proteins are a large and highly diverse gene family present in a wide range of plant species. LEAs are proposed to play a role in various stress tolerance responses. Our study represents the first-ever survey of LEA proteins and their encoding genes in a widely distributed pine (Pinus tabuliformis) in China. Twenty-three LEA genes were identified from the P. tabuliformis belonging to seven groups. Proteins with repeated motifs are an important feature specific to LEA groups. Ten of 23 pine LEA genes were selectively expressed in specific tissues, and showed expression divergence within each group. In addition, we selected 13 genes representing each group and introduced theses genes into Escherichia coli to assess the protective function of PtaLEA under heat and salt stresses. Compared with control cells, the E. coli cells expressing PtaLEA fusion protein exhibited enhanced salt and heat resistance and viability, indicating the protein may play a protective role in cells under stress conditions. Furthermore, among these enhanced tolerance genes, a certain extent of function divergence appeared within a gene group as well as between gene groups, suggesting potential functional diversity of this gene family in conifers.

Coiled Coils - A Model System for the 21st Century.

PubMed

Lupas, Andrei N; Bassler, Jens

2017-02-01

α-Helical coiled coils were described more than 60 years ago as simple, repetitive structures mediating oligomerization and mechanical stability. Over the past 20 years, however, they have emerged as one of the most diverse protein folds in nature, enabling many biological functions beyond mechanical rigidity, such as membrane fusion, signal transduction, and solute transport. Despite this great diversity, their structures can be described by parametric equations, making them uniquely suited for rational protein design. Far from having been exhausted as a source of structural insight and a basis for functional engineering, coiled coils are poised to become even more important for protein science in the coming decades. Copyright © 2016 Elsevier Ltd. All rights reserved.

Ice-binding proteins: a remarkable diversity of structures for stopping and starting ice growth.

PubMed

Davies, Peter L

2014-11-01

Antifreeze proteins (AFPs) were discovered in marine fishes that need protection from freezing. These ice-binding proteins (IBPs) are widespread across biological kingdoms, and their functions include freeze tolerance and ice adhesion. Consistent with recent independent evolution, AFPs have remarkably diverse folds that rely heavily on hydrogen- and disulfide-bonding. AFP ice-binding sites are typically flat, extensive, relatively hydrophobic, and are thought to organize water into an ice-like arrangement that merges and freezes with the quasi-liquid layer next to the ice lattice. In this article, the roles, properties, and structure-function interactions of IBPs are reviewed, and their relationship to ice nucleation proteins, which promote freezing at high subzero temperatures, is explored. Copyright © 2014 Elsevier Ltd. All rights reserved.

DNA-mediated engineering of multicomponent enzyme crystals

PubMed Central

Brodin, Jeffrey D.; Auyeung, Evelyn; Mirkin, Chad A.

2015-01-01

The ability to predictably control the coassembly of multiple nanoscale building blocks, especially those with disparate chemical and physical properties such as biomolecules and inorganic nanoparticles, has far-reaching implications in catalysis, sensing, and photonics, but a generalizable strategy for engineering specific contacts between these particles is an outstanding challenge. This is especially true in the case of proteins, where the types of possible interparticle interactions are numerous, diverse, and complex. Herein, we explore the concept of trading protein–protein interactions for DNA–DNA interactions to direct the assembly of two nucleic-acid–functionalized proteins with distinct surface chemistries into six unique lattices composed of catalytically active proteins, or of a combination of proteins and DNA-modified gold nanoparticles. The programmable nature of DNA–DNA interactions used in this strategy allows us to control the lattice symmetries and unit cell constants, as well as the compositions and habit, of the resulting crystals. This study provides a potentially generalizable strategy for constructing a unique class of materials that take advantage of the diverse morphologies, surface chemistries, and functionalities of proteins for assembling functional crystalline materials. PMID:25831510

DNA-mediated engineering of multicomponent enzyme crystals

DOE PAGES

Brodin, Jeffrey D.; Auyeung, Evelyn; Mirkin, Chad A.

2015-03-23

The ability to predictably control the coassembly of multiple nanoscale building blocks, especially those with disparate chemical and physical properties such as biomolecules and inorganic nanoparticles, has far-reaching implications in catalysis, sensing, and photonics, but a generalizable strategy for engineering specific contacts between these particles is an outstanding challenge. This is especially true in the case of proteins, where the types of possible interparticle interactions are numerous, diverse, and complex. In this paper, we explore the concept of trading protein–protein interactions for DNA–DNA interactions to direct the assembly of two nucleic-acid–functionalized proteins with distinct surface chemistries into six unique latticesmore » composed of catalytically active proteins, or of a combination of proteins and DNA-modified gold nanoparticles. The programmable nature of DNA–DNA interactions used in this strategy allows us to control the lattice symmetries and unit cell constants, as well as the compositions and habit, of the resulting crystals. Finally, this study provides a potentially generalizable strategy for constructing a unique class of materials that take advantage of the diverse morphologies, surface chemistries, and functionalities of proteins for assembling functional crystalline materials.« less

Protein change in plant evolution: tracing one thread connecting molecular and phenotypic diversity

PubMed Central

Bartlett, Madelaine E.; Whipple, Clinton J.

2013-01-01

Proteins change over the course of evolutionary time. New protein-coding genes and gene families emerge and diversify, ultimately affecting an organism’s phenotype and interactions with its environment. Here we survey the range of structural protein change observed in plants and review the role these changes have had in the evolution of plant form and function. Verified examples tying evolutionary change in protein structure to phenotypic change remain scarce. We will review the existing examples, as well as draw from investigations into domestication, and quantitative trait locus (QTL) cloning studies searching for the molecular underpinnings of natural variation. The evolutionary significance of many cloned QTL has not been assessed, but all the examples identified so far have begun to reveal the extent of protein structural diversity tolerated in natural systems. This molecular (and phenotypic) diversity could come to represent part of natural selection’s source material in the adaptive evolution of novel traits. Protein structure and function can change in many distinct ways, but the changes we identified in studies of natural diversity and protein evolution were predicted to fall primarily into one of six categories: altered active and binding sites; altered protein–protein interactions; altered domain content; altered activity as an activator or repressor; altered protein stability; and hypomorphic and hypermorphic alleles. There was also variability in the evolutionary scale at which particular changes were observed. Some changes were detected at both micro- and macroevolutionary timescales, while others were observed primarily at deep or shallow phylogenetic levels. This variation might be used to determine the trajectory of future investigations in structural molecular evolution. PMID:24124420

Functional diversity of HIV-1 envelope proteins expressed by contemporaneous plasma viruses

PubMed Central

Nora, Tamara; Bouchonnet, Francine; Labrosse, Béatrice; Charpentier, Charlotte; Mammano, Fabrizio; Clavel, François; Hance, Allan J

2008-01-01

Background Numerous studies have shown that viral quasi-species with genetically diverse envelope proteins (Env) replicate simultaneously in patients infected with the human immunodeficiency virus type 1 (HIV-1). Less information is available concerning the extent that envelope sequence diversity translates into a diversity of phenotypic properties, including infectivity and resistance to entry inhibitors. Methods To study these questions, we isolated genetically distinct contemporaneous clonal viral populations from the plasma of 5 HIV-1 infected individuals (n = 70), and evaluated the infectivity of recombinant viruses expressing Env proteins from the clonal viruses in several target cells. The sensitivity to entry inhibitors (enfuvirtide, TAK-799), soluble CD4 and monoclonal antibodies (2G12, 48d, 2F5) was also evaluated for a subset of the recombinant viruses (n = 20). Results Even when comparisons were restricted to viruses with similar tropism, the infectivity for a given target cell of viruses carrying different Env proteins from the same patient varied over an approximately 10-fold range, and differences in their relative ability to infect different target cells were also observed. Variable region haplotypes associated with high and low infectivity could be identified for one patient. In addition, clones carrying unique mutations in V3 often displayed low infectivity. No correlation was observed between viral infectivity and sensitivity to inhibition by any of the six entry inhibitors evaluated, indicating that these properties can be dissociated. Significant inter-patient differences, independent of infectivity, were observed for the sensitivity of Env proteins to several entry inhibitors and their ability to infect different target cells. Conclusion These findings demonstrate the marked functional heterogeneity of HIV-1 Env proteins expressed by contemporaneous circulating viruses, and underscore the advantage of clonal analyses in characterizing the spectrum of functional properties of the genetically diverse viral populations present in a given patient. PMID:18312646

Green fluorescent protein nanopolygons as monodisperse supramolecular assemblies of functional proteins with defined valency

PubMed Central

Kim, Young Eun; Kim, Yu-na; Kim, Jung A.; Kim, Ho Min; Jung, Yongwon

2015-01-01

Supramolecular protein assemblies offer novel nanoscale architectures with molecular precision and unparalleled functional diversity. A key challenge, however, is to create precise nano-assemblies of functional proteins with both defined structures and a controlled number of protein-building blocks. Here we report a series of supramolecular green fluorescent protein oligomers that are assembled in precise polygonal geometries and prepared in a monodisperse population. Green fluorescent protein is engineered to be self-assembled in cells into oligomeric assemblies that are natively separated in a single-protein resolution by surface charge manipulation, affording monodisperse protein (nano)polygons from dimer to decamer. Several functional proteins are multivalently displayed on the oligomers with controlled orientations. Spatial arrangements of protein oligomers and displayed functional proteins are directly visualized by a transmission electron microscope. By employing our functional protein assemblies, we provide experimental insight into multivalent protein–protein interactions and tools to manipulate receptor clustering on live cell surfaces. PMID:25972078

Diversity-oriented synthetic strategy for developing a chemical modulator of protein-protein interaction

NASA Astrophysics Data System (ADS)

Kim, Jonghoon; Jung, Jinjoo; Koo, Jaeyoung; Cho, Wansang; Lee, Won Seok; Kim, Chanwoo; Park, Wonwoo; Park, Seung Bum

2016-10-01

Diversity-oriented synthesis (DOS) can provide a collection of diverse and complex drug-like small molecules, which is critical in the development of new chemical probes for biological research of undruggable targets. However, the design and synthesis of small-molecule libraries with improved biological relevance as well as maximized molecular diversity represent a key challenge. Herein, we employ functional group-pairing strategy for the DOS of a chemical library containing privileged substructures, pyrimidodiazepine or pyrimidine moieties, as chemical navigators towards unexplored bioactive chemical space. To validate the utility of this DOS library, we identify a new small-molecule inhibitor of leucyl-tRNA synthetase-RagD protein-protein interaction, which regulates the amino acid-dependent activation of mechanistic target of rapamycin complex 1 signalling pathway. This work highlights that privileged substructure-based DOS strategy can be a powerful research tool for the construction of drug-like compounds to address challenging biological targets.

Insights into animal and plant lectins with antimicrobial activities.

PubMed

Dias, Renata de Oliveira; Machado, Leandro Dos Santos; Migliolo, Ludovico; Franco, Octavio Luiz

2015-01-05

Lectins are multivalent proteins with the ability to recognize and bind diverse carbohydrate structures. The glyco -binding and diverse molecular structures observed in these protein classes make them a large and heterogeneous group with a wide range of biological activities in microorganisms, animals and plants. Lectins from plants and animals are commonly used in direct defense against pathogens and in immune regulation. This review focuses on sources of animal and plant lectins, describing their functional classification and tridimensional structures, relating these properties with biotechnological purposes, including antimicrobial activities. In summary, this work focuses on structural-functional elucidation of diverse lectin groups, shedding some light on host-pathogen interactions; it also examines their emergence as biotechnological tools through gene manipulation and development of new drugs.

PHB in Cardiovascular and Other Diseases: Present Knowledge and Implications.

PubMed

Chowdhury, Debabrata; Kumar, Dinesh; Sarma, Pranjal; Tangutur, Anjana Devi; Bhadra, Manika Pal

2017-11-30

Prohibitin (PHB) is overtly conserved evolutionarily and ubiquitously expressed protein with pleiotropic functions in diverse cellular compartments. However, regulation and function of these proteins in different cells, tissues and in various diseases is different as evidenced by expression of these proteins which is found to be reduced in heart diseases, kidney diseases, lung disease, Crohn's disease and ulcerative colitis but this protein is highly expressed in diverse cancers. The mechanism by which this protein acts at the molecular level in different subcellular localizations or in different cells or tissues in different conditions (diseases or normal) has remained poorly understood. There are several studies reported to understand and decipher PHB's role in diseases and/or cancers of ovary, lung, stomach, thyroid, liver, blood, prostrate, gastric, esophagus, glioma, breast, bladder etc. where PHB is shown to act through mechanisms by acting as oncogene, tumor suppressor, antioxidant, antiapoptotic, in angiogenesis, autophagy etc. This review specifically gives attention to the functional role and regulatory mechanism of PHB proteins in cardiovascular health and diseases and its associated implications. Various molecular pathways involved in PHB function and its regulation are analyzed. PHB is rapidly emerging as a critical target molecule for cardiovascular signaling. Progress in delineating CVD and mechanisms of PHB in diverse molecular pathways is essential for determining when and how PHB targeted therapy might be feasible. In this regard, new therapies targeting PHB may best be applied in the future together with molecular profiling of CVD for clinical stratification of disease diagnosis and prognosis. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Diversity in arrestin function.

PubMed

Kendall, Ryan T; Luttrell, Louis M

2009-09-01

The termination of heptahelical receptor signaling is a multilevel process coordinated, in large part, by members of the arrestin family of proteins. Arrestin binding to agonist-occupied receptors promotes desensitization by interrupting receptor-G protein coupling, while simultaneously recruiting machinery for receptor endocytosis, vesicular trafficking, and receptor fate determination. By simultaneously binding other proteins, arrestins also act as ligand-regulated scaffolds that recruit protein and lipid kinase, phosphatase, phosphodiesterase, and ubiquitin ligase activity into receptor-based multiprotein 'signalsome' complexes. Arrestin-binding thus 'switches' receptors from a transient G protein-coupled state to a persistent arrestin-coupled state that continues to signal as the receptor transits intracellular compartments. While it is clear that signalsome assembly has profound effects on the duration and spatial characteristics of heptahelical receptor signals, the physiologic functions of this novel signaling mechanism are poorly understood. Growing evidence suggests that signalsomes regulate such diverse processes as endocytosis and exocytosis, cell migration, survival, and contractility.

Visualisation and graph-theoretic analysis of a large-scale protein structural interactome

PubMed Central

Bolser, Dan; Dafas, Panos; Harrington, Richard; Park, Jong; Schroeder, Michael

2003-01-01

Background Large-scale protein interaction maps provide a new, global perspective with which to analyse protein function. PSIMAP, the Protein Structural Interactome Map, is a database of all the structurally observed interactions between superfamilies of protein domains with known three-dimensional structure in the PDB. PSIMAP incorporates both functional and evolutionary information into a single network. Results We present a global analysis of PSIMAP using several distinct network measures relating to centrality, interactivity, fault-tolerance, and taxonomic diversity. We found the following results: Centrality: we show that the center and barycenter of PSIMAP do not coincide, and that the superfamilies forming the barycenter relate to very general functions, while those constituting the center relate to enzymatic activity. Interactivity: we identify the P-loop and immunoglobulin superfamilies as the most highly interactive. We successfully use connectivity and cluster index, which characterise the connectivity of a superfamily's neighbourhood, to discover superfamilies of complex I and II. This is particularly significant as the structure of complex I is not yet solved. Taxonomic diversity: we found that highly interactive superfamilies are in general taxonomically very diverse and are thus amongst the oldest. Fault-tolerance: we found that the network is very robust as for the majority of superfamilies removal from the network will not break up the network. Conclusions Overall, we can single out the P-loop containing nucleotide triphosphate hydrolases superfamily as it is the most highly connected and has the highest taxonomic diversity. In addition, this superfamily has the highest interaction rank, is the barycenter of the network (it has the shortest average path to every other superfamily in the network), and is an articulation vertex, whose removal will disconnect the network. More generally, we conclude that the graph-theoretic and taxonomic analysis of PSIMAP is an important step towards the understanding of protein function and could be an important tool for tracing the evolution of life at the molecular level. PMID:14531933

Nanochemistry of Protein-Based Delivery Agents

PubMed Central

Rajendran, Subin R. C. K.; Udenigwe, Chibuike C.; Yada, Rickey Y.

2016-01-01

The past decade has seen an increased interest in the conversion of food proteins into functional biomaterials, including their use for loading and delivery of physiologically active compounds such as nutraceuticals and pharmaceuticals. Proteins possess a competitive advantage over other platforms for the development of nanodelivery systems since they are biocompatible, amphipathic, and widely available. Proteins also have unique molecular structures and diverse functional groups that can be selectively modified to alter encapsulation and release properties. A number of physical and chemical methods have been used for preparing protein nanoformulations, each based on different underlying protein chemistry. This review focuses on the chemistry of the reorganization and/or modification of proteins into functional nanostructures for delivery, from the perspective of their preparation, functionality, stability and physiological behavior. PMID:27489854

Nanochemistry of protein-based delivery agents

NASA Astrophysics Data System (ADS)

Rajendran, Subin; Udenigwe, Chibuike; Yada, Rickey

2016-07-01

The past decade has seen an increased interest in the conversion of food proteins into functional biomaterials, including their use for loading and delivery of physiologically active compounds such as nutraceuticals and pharmaceuticals. Proteins possess a competitive advantage over other platforms for the development of nanodelivery systems since they are biocompatible, amphipathic, and widely available. Proteins also have unique molecular structures and diverse functional groups that can be selectively modified to alter encapsulation and release properties. A number of physical and chemical methods have been used for preparing protein nanoformulations, each based on different underlying protein chemistry. This review focuses on the chemistry of the reorganization and/or modification of proteins into functional nanostructures for delivery, from the perspective of their preparation, functionality, stability and physiological behavior.

Shedding new light on opsin evolution

PubMed Central

Porter, Megan L.; Blasic, Joseph R.; Bok, Michael J.; Cameron, Evan G.; Pringle, Thomas; Cronin, Thomas W.; Robinson, Phyllis R.

2012-01-01

Opsin proteins are essential molecules in mediating the ability of animals to detect and use light for diverse biological functions. Therefore, understanding the evolutionary history of opsins is key to understanding the evolution of light detection and photoreception in animals. As genomic data have appeared and rapidly expanded in quantity, it has become possible to analyse opsins that functionally and histologically are less well characterized, and thus to examine opsin evolution strictly from a genetic perspective. We have incorporated these new data into a large-scale, genome-based analysis of opsin evolution. We use an extensive phylogeny of currently known opsin sequence diversity as a foundation for examining the evolutionary distributions of key functional features within the opsin clade. This new analysis illustrates the lability of opsin protein-expression patterns, site-specific functionality (i.e. counterion position) and G-protein binding interactions. Further, it demonstrates the limitations of current model organisms, and highlights the need for further characterization of many of the opsin sequence groups with unknown function. PMID:22012981

Regulation, Signaling, and Physiological Functions of G-Proteins.

PubMed

Syrovatkina, Viktoriya; Alegre, Kamela O; Dey, Raja; Huang, Xin-Yun

2016-09-25

Heterotrimeric guanine-nucleotide-binding regulatory proteins (G-proteins) mainly relay the information from G-protein-coupled receptors (GPCRs) on the plasma membrane to the inside of cells to regulate various biochemical functions. Depending on the targeted cell types, tissues, and organs, these signals modulate diverse physiological functions. The basic schemes of heterotrimeric G-proteins have been outlined. In this review, we briefly summarize what is known about the regulation, signaling, and physiological functions of G-proteins. We then focus on a few less explored areas such as the regulation of G-proteins by non-GPCRs and the physiological functions of G-proteins that cannot be easily explained by the known G-protein signaling pathways. There are new signaling pathways and physiological functions for G-proteins to be discovered and further interrogated. With the advancements in structural and computational biological techniques, we are closer to having a better understanding of how G-proteins are regulated and of the specificity of G-protein interactions with their regulators. Copyright © 2016 Elsevier Ltd. All rights reserved.

Mapping of protein- and chromatin-interactions at the nuclear lamina.

PubMed

Kubben, Nard; Voncken, Jan Willem; Misteli, Tom

2010-01-01

The nuclear envelope and the lamina define the nuclear periphery and are implicated in many nuclear processes including chromatin organization, transcription and DNA replication. Mutations in lamin A proteins, major components of the lamina, interfere with these functions and cause a set of phenotypically diverse diseases referred to as laminopathies. The phenotypic diversity of laminopathies is thought to be the result of alterations in specific protein- and chromatin interactions due to lamin A mutations. Systematic identification of lamin A-protein and -chromatin interactions will be critical to uncover the molecular etiology of laminopathies. Here we summarize and critically discuss recent technology to analyze lamina-protein and-chromatin interactions.

Inorganic pyrophosphatases: structural diversity serving the function

NASA Astrophysics Data System (ADS)

Samygina, V. R.

2016-05-01

The review is devoted to ubiquitous enzymes, inorganic pyrophosphatases, which are essential in all living organisms. Despite the long history of investigations, these enzymes continue to attract interest. The review focuses on the three-dimensional structures of various representatives of this class of proteins. The structural diversity, the relationship between the structure and some properties of pyrophosphatases and various mechanisms of enzyme action related to the structural diversity of these enzymes are discussed. Interactions of pyrophosphatase with other proteins and possible practical applications are considered. The bibliography includes 56 references.

Trichomonas vaginalis vast BspA-like gene family: evidence for functional diversity from structural organisation and transcriptomics

PubMed Central

2010-01-01

Background Trichomonas vaginalis is the most common non-viral human sexually transmitted pathogen and importantly, contributes to facilitating the spread of HIV. Yet very little is known about its surface and secreted proteins mediating interactions with, and permitting the invasion and colonisation of, the host mucosa. Initial annotations of T. vaginalis genome identified a plethora of candidate extracellular proteins. Results Data mining of the T. vaginalis genome identified 911 BspA-like entries (TvBspA) sharing TpLRR-like leucine-rich repeats, which represent the largest gene family encoding potential extracellular proteins for the pathogen. A broad range of microorganisms encoding BspA-like proteins was identified and these are mainly known to live on mucosal surfaces, among these T. vaginalis is endowed with the largest gene family. Over 190 TvBspA proteins with inferred transmembrane domains were characterised by a considerable structural diversity between their TpLRR and other types of repetitive sequences and two subfamilies possessed distinct classic sorting signal motifs for endocytosis. One TvBspA subfamily also shared a glycine-rich protein domain with proteins from Clostridium difficile pathogenic strains and C. difficile phages. Consistent with the hypothesis that TvBspA protein structural diversity implies diverse roles, we demonstrated for several TvBspA genes differential expression at the transcript level in different growth conditions. Identified variants of repetitive segments between several TvBspA paralogues and orthologues from two clinical isolates were also consistent with TpLRR and other repetitive sequences to be functionally important. For one TvBspA protein cell surface expression and antibody responses by both female and male T. vaginalis infected patients were also demonstrated. Conclusions The biased mucosal habitat for microbial species encoding BspA-like proteins, the characterisation of a vast structural diversity for the TvBspA proteins, differential expression of a subset of TvBspA genes and the cellular localisation and immunological data for one TvBspA; all point to the importance of the TvBspA proteins to various aspects of T. vaginalis pathobiology at the host-pathogen interface. PMID:20144183

Intrinsic disorder mediates the diverse regulatory functions of the Cdk inhibitor p21

PubMed Central

Wang, Yuefeng; Fisher, John C.; Mathew, Rose; Ou, Li; Otieno, Steve; Sublett, Jack; Xiao, Limin; Chen, Jianhan; Roussel, Martine F.; Kriwacki, Richard W.

2011-01-01

Traditionally, well-defined three-dimensional structure was thought to be essential for protein function. However, myriad biological functions are performed by highly dynamic, intrinsically disordered proteins (IDPs). IDPs often fold upon binding their biological targets and frequently exhibit “binding diversity” by targeting multiple ligands. We sought to understand the physical basis of IDP binding diversity and herein report that the cyclin-dependent kinase (Cdk) inhibitor, p21Cip1, adaptively binds to and inhibits the various Cdk/cyclin complexes that regulate eukaryotic cell division. Based on results from NMR spectroscopy, and biochemical and cellular assays, we show that structural adaptability of a helical sub-domain within p21 termed LH enables two other sub-domains termed D1 and D2 to specifically bind conserved surface features of the cyclin and Cdk subunits, respectively, within otherwise structurally distinct Cdk/cyclin complexes. Adaptive folding upon binding is likely to mediate the diverse biological functions of the thousands of IDPs present in eukaryotes. PMID:21358637

Protein-protein interactions in the regulation of WRKY transcription factors.

PubMed

Chi, Yingjun; Yang, Yan; Zhou, Yuan; Zhou, Jie; Fan, Baofang; Yu, Jing-Quan; Chen, Zhixiang

2013-03-01

It has been almost 20 years since the first report of a WRKY transcription factor, SPF1, from sweet potato. Great progress has been made since then in establishing the diverse biological roles of WRKY transcription factors in plant growth, development, and responses to biotic and abiotic stress. Despite the functional diversity, almost all analyzed WRKY proteins recognize the TTGACC/T W-box sequences and, therefore, mechanisms other than mere recognition of the core W-box promoter elements are necessary to achieve the regulatory specificity of WRKY transcription factors. Research over the past several years has revealed that WRKY transcription factors physically interact with a wide range of proteins with roles in signaling, transcription, and chromatin remodeling. Studies of WRKY-interacting proteins have provided important insights into the regulation and mode of action of members of the important family of transcription factors. It has also emerged that the slightly varied WRKY domains and other protein motifs conserved within each of the seven WRKY subfamilies participate in protein-protein interactions and mediate complex functional interactions between WRKY proteins and between WRKY and other regulatory proteins in the modulation of important biological processes. In this review, we summarize studies of protein-protein interactions for WRKY transcription factors and discuss how the interacting partners contribute, at different levels, to the establishment of the complex regulatory and functional network of WRKY transcription factors.

Vectors for fluorescent protein tagging in Phytophthora: tools for functional genomics and cell biology.

PubMed

Ah-Fong, Audrey M V; Judelson, Howard S

2011-09-01

Fluorescent tagging has become the strategy of choice for examining the subcellular localisation of proteins. To develop a versatile community resource for this method in oomycetes, plasmids were constructed that allow the expression of either of four spectrally distinct proteins [cyan fluorescent protein (CFP), green fluorescent protein (GFP), yellow fluorescent protein (YFP), and mCherry], alone or fused at their N- or C-termini, to sequences of interest. Equivalent sets of plasmids were made using neomycin or hygromycin phosphotransferases (nptII, hpt) as selectable markers, to facilitate double-labelling and aid work in diverse species. The fluorescent proteins and drug-resistance markers were fused to transcriptional regulatory sequences from the oomycete Bremia lactucae, which are known to function in diverse oomycetes, although the promoter in the fluorescence cassette (ham34) can be replaced easily by a promoter of interest. The function of each plasmid was confirmed in Phytophthora infestans. Moreover, fusion proteins were generated using targeting sequences for the endoplasmic reticulum, Golgi, mitochondria, nuclei, and peroxisomes. Studies of the distribution of the fusions in mycelia and sporangia provided insight into cellular organisation at different stages of development. This toolbox of vectors should advance studies of gene function and cell biology in Phytophthora and other oomycetes. Copyright © 2011 British Mycological Society. Published by Elsevier Ltd. All rights reserved.

DEAD-box Helicases as Integrators of RNA, Nucleotide and Protein Binding

PubMed Central

Putnam, Andrea A.

2013-01-01

DEAD-box helicases perform diverse cellular functions in virtually all steps of RNA metabolism from Bacteria to Humans. Although DEAD-box helicases share a highly conserved core domain, the enzymes catalyze a wide range of biochemical reactions. In addition to the well established RNA unwinding and corresponding ATPase activities, DEAD-box helicases promote duplex formation and displace proteins from RNA. They can also function as assembly platforms for larger ribonucleoprotein complexes, and as metabolite sensors. This review aims to provide a perspective on the diverse biochemical features of DEAD-box helicases and connections to structural information. We discuss these data in the context of a model that views the enzymes as integrators of RNA, nucleotide, and protein binding. PMID:23416748

CRISPR-Cas Adaptive Immune Systems of the Sulfolobales: Unravelling Their Complexity and Diversity

PubMed Central

Garrett, Roger A.; Shah, Shiraz A.; Erdmann, Susanne; Liu, Guannan; Mousaei, Marzieh; León-Sobrino, Carlos; Peng, Wenfang; Gudbergsdottir, Soley; Deng, Ling; Vestergaard, Gisle; Peng, Xu; She, Qunxin

2015-01-01

The Sulfolobales have provided good model organisms for studying CRISPR-Cas systems of the crenarchaeal kingdom of the archaea. These organisms are infected by a wide range of exceptional archaea-specific viruses and conjugative plasmids, and their CRISPR-Cas systems generally exhibit extensive structural and functional diversity. They carry large and multiple CRISPR loci and often multiple copies of diverse Type I and Type III interference modules as well as more homogeneous adaptation modules. These acidothermophilic organisms have recently provided seminal insights into both the adaptation process, the diverse modes of interference, and their modes of regulation. The functions of the adaptation and interference modules tend to be loosely coupled and the stringency of the crRNA-DNA sequence matching during DNA interference is relatively low, in contrast to some more streamlined CRISPR-Cas systems of bacteria. Despite this, there is evidence for a complex and differential regulation of expression of the diverse functional modules in response to viral infection. Recent work also supports critical roles for non-core Cas proteins, especially during Type III-directed interference, and this is consistent with these proteins tending to coevolve with core Cas proteins. Various novel aspects of CRISPR-Cas systems of the Sulfolobales are considered including an alternative spacer acquisition mechanism, reversible spacer acquisition, the formation and significance of antisense CRISPR RNAs, and a novel mechanism for avoidance of CRISPR-Cas defense. Finally, questions regarding the basis for the complexity, diversity, and apparent redundancy, of the intracellular CRISPR-Cas systems are discussed. PMID:25764276

Multifunctional Mitochondrial AAA Proteases

PubMed Central

Glynn, Steven E.

2017-01-01

Mitochondria perform numerous functions necessary for the survival of eukaryotic cells. These activities are coordinated by a diverse complement of proteins encoded in both the nuclear and mitochondrial genomes that must be properly organized and maintained. Misregulation of mitochondrial proteostasis impairs organellar function and can result in the development of severe human diseases. ATP-driven AAA+ proteins play crucial roles in preserving mitochondrial activity by removing and remodeling protein molecules in accordance with the needs of the cell. Two mitochondrial AAA proteases, i-AAA and m-AAA, are anchored to either face of the mitochondrial inner membrane, where they engage and process an array of substrates to impact protein biogenesis, quality control, and the regulation of key metabolic pathways. The functionality of these proteases is extended through multiple substrate-dependent modes of action, including complete degradation, partial processing, or dislocation from the membrane without proteolysis. This review discusses recent advances made toward elucidating the mechanisms of substrate recognition, handling, and degradation that allow these versatile proteases to control diverse activities in this multifunctional organelle. PMID:28589125

Multifunctional Mitochondrial AAA Proteases.

PubMed

Glynn, Steven E

2017-01-01

Mitochondria perform numerous functions necessary for the survival of eukaryotic cells. These activities are coordinated by a diverse complement of proteins encoded in both the nuclear and mitochondrial genomes that must be properly organized and maintained. Misregulation of mitochondrial proteostasis impairs organellar function and can result in the development of severe human diseases. ATP-driven AAA+ proteins play crucial roles in preserving mitochondrial activity by removing and remodeling protein molecules in accordance with the needs of the cell. Two mitochondrial AAA proteases, i-AAA and m-AAA, are anchored to either face of the mitochondrial inner membrane, where they engage and process an array of substrates to impact protein biogenesis, quality control, and the regulation of key metabolic pathways. The functionality of these proteases is extended through multiple substrate-dependent modes of action, including complete degradation, partial processing, or dislocation from the membrane without proteolysis. This review discusses recent advances made toward elucidating the mechanisms of substrate recognition, handling, and degradation that allow these versatile proteases to control diverse activities in this multifunctional organelle.

Vertebrate Presynaptic Active Zone Assembly: a Role Accomplished by Diverse Molecular and Cellular Mechanisms.

PubMed

Torres, Viviana I; Inestrosa, Nibaldo C

2018-06-01

Among all the biological systems in vertebrates, the central nervous system (CNS) is the most complex, and its function depends on specialized contacts among neurons called synapses. The assembly and organization of synapses must be exquisitely regulated for a normal brain function and network activity. There has been a tremendous effort in recent decades to understand the molecular and cellular mechanisms participating in the formation of new synapses and their organization, maintenance, and regulation. At the vertebrate presynapses, proteins such as Piccolo, Bassoon, RIM, RIM-BPs, CAST/ELKS, liprin-α, and Munc13 are constant residents and participate in multiple and dynamic interactions with other regulatory proteins, which define network activity and normal brain function. Here, we review the function of these active zone (AZ) proteins and diverse factors involved in AZ assembly and maintenance, with an emphasis on axonal trafficking of precursor vesicles, protein homo- and hetero-oligomeric interactions as a mechanism of AZ trapping and stabilization, and the role of F-actin in presynaptic assembly and its modulation by Wnt signaling.

Functional understanding of the diverse exon-intron structures of human GPCR genes.

PubMed

Hammond, Dorothy A; Olman, Victor; Xu, Ying

2014-02-01

The GPCR genes have a variety of exon-intron structures even though their proteins are all structurally homologous. We have examined all human GPCR genes with at least two functional protein isoforms, totaling 199, aiming to gain an understanding of what may have contributed to the large diversity of the exon-intron structures of the GPCR genes. The 199 genes have a total of 808 known protein splicing isoforms with experimentally verified functions. Our analysis reveals that 1301 (80.6%) adjacent exon-exon pairs out of the total of 1,613 in the 199 genes have either exactly one exon skipped or the intron in-between retained in at least one of the 808 protein splicing isoforms. This observation has a statistical significance p-value of 2.051762 * e(-09), assuming that the observed splicing isoforms are independent of the exon-intron structures. Our interpretation of this observation is that the exon boundaries of the GPCR genes are not randomly determined; instead they may be selected to facilitate specific alternative splicing for functional purposes.

The Functional Quality of Soluble Recombinant Polypeptides Produced in Escherichia coli Is Defined by a Wide Conformational Spectrum▿

PubMed Central

Martínez-Alonso, Mónica; González-Montalbán, Nuria; García-Fruitós, Elena; Villaverde, Antonio

2008-01-01

We have observed that a soluble recombinant green fluorescent protein produced in Escherichia coli occurs in a wide conformational spectrum. This results in differently fluorescent protein fractions in which morphologically diverse soluble aggregates abound. Therefore, the functional quality of soluble versions of aggregation-prone recombinant proteins is defined statistically rather than by the prevalence of a canonical native structure. PMID:18836021

pocketZebra: a web-server for automated selection and classification of subfamily-specific binding sites by bioinformatic analysis of diverse protein families

PubMed Central

Suplatov, Dmitry; Kirilin, Eugeny; Arbatsky, Mikhail; Takhaveev, Vakil; Švedas, Vytas

2014-01-01

The new web-server pocketZebra implements the power of bioinformatics and geometry-based structural approaches to identify and rank subfamily-specific binding sites in proteins by functional significance, and select particular positions in the structure that determine selective accommodation of ligands. A new scoring function has been developed to annotate binding sites by the presence of the subfamily-specific positions in diverse protein families. pocketZebra web-server has multiple input modes to meet the needs of users with different experience in bioinformatics. The server provides on-site visualization of the results as well as off-line version of the output in annotated text format and as PyMol sessions ready for structural analysis. pocketZebra can be used to study structure–function relationship and regulation in large protein superfamilies, classify functionally important binding sites and annotate proteins with unknown function. The server can be used to engineer ligand-binding sites and allosteric regulation of enzymes, or implemented in a drug discovery process to search for potential molecular targets and novel selective inhibitors/effectors. The server, documentation and examples are freely available at http://biokinet.belozersky.msu.ru/pocketzebra and there are no login requirements. PMID:24852248

Profiling of proteins secreted in the bovine oviduct reveals diverse functions of this luminal microenvironment.

PubMed

Pillai, Viju Vijayan; Weber, Darren M; Phinney, Brett S; Selvaraj, Vimal

2017-01-01

The oviductal microenvironment is a site for key events that involve gamete maturation, fertilization and early embryo development. Secretions into the oviductal lumen by either the lining epithelium or by transudation of plasma constituents are known to contain elements conducive for reproductive success. Although previous studies have identified some of these factors involved in reproduction, knowledge of secreted proteins in the oviductal fluid remains rudimentary with limited definition of function even in extensively studied species like cattle. In this study, we used a shotgun proteomics approach followed by bioinformatics sequence prediction to identify secreted proteins present in the bovine oviductal fluid (ex vivo) and secretions from the bovine oviductal epithelial cells (in vitro). From a total of 2087 proteins identified, 266 proteins could be classified as secreted, 109 (41%) of which were common for both in vivo and in vitro conditions. Pathway analysis indicated different classes of proteins that included growth factors, metabolic regulators, immune modulators, enzymes, and extracellular matrix components. Functional analysis revealed mechanisms in the oviductal lumen linked to immune homeostasis, gamete maturation, fertilization and early embryo development. These results point to several novel components that work together with known elements mediating functional homeostasis, and highlight the diversity of machinery associated with oviductal physiology and early events in cattle fertility.

A-Kinase Anchoring Proteins: From protein complexes to physiology and disease

PubMed Central

Carnegie, Graeme K.; Means, Christopher K.; Scott, John D.

2009-01-01

Protein scaffold complexes are a key mechanism by which a common signaling pathway can serve many different functions. Sequestering a signaling enzyme to a specific subcellular environment not only ensures that the enzyme is near its relevant targets, but also segregates this activity to prevent indiscriminate phosphorylation of other substrates. One family of diverse, well-studied scaffolding proteins are the A-kinase anchoring proteins (AKAPs). These anchoring proteins form multi-protein complexes that integrate cAMP signaling with other pathways and signaling events. In this review we focus on recent advances in the elucidation of AKAP function. PMID:19319965

A-kinase anchoring proteins: from protein complexes to physiology and disease.

PubMed

Carnegie, Graeme K; Means, Christopher K; Scott, John D

2009-04-01

Protein scaffold complexes are a key mechanism by which a common signaling pathway can serve many different functions. Sequestering a signaling enzyme to a specific subcellular environment not only ensures that the enzyme is near its relevant targets, but also segregates this activity to prevent indiscriminate phosphorylation of other substrates. One family of diverse, well-studied scaffolding proteins are the A-kinase anchoring proteins (AKAPs). These anchoring proteins form multi-protein complexes that integrate cAMP signaling with other pathways and signaling events. In this review, we focus on recent advances in the elucidation of AKAP function.

MoonProt: a database for proteins that are known to moonlight

PubMed Central

Mani, Mathew; Chen, Chang; Amblee, Vaishak; Liu, Haipeng; Mathur, Tanu; Zwicke, Grant; Zabad, Shadi; Patel, Bansi; Thakkar, Jagravi; Jeffery, Constance J.

2015-01-01

Moonlighting proteins comprise a class of multifunctional proteins in which a single polypeptide chain performs multiple biochemical functions that are not due to gene fusions, multiple RNA splice variants or pleiotropic effects. The known moonlighting proteins perform a variety of diverse functions in many different cell types and species, and information about their structures and functions is scattered in many publications. We have constructed the manually curated, searchable, internet-based MoonProt Database (http://www.moonlightingproteins.org) with information about the over 200 proteins that have been experimentally verified to be moonlighting proteins. The availability of this organized information provides a more complete picture of what is currently known about moonlighting proteins. The database will also aid researchers in other fields, including determining the functions of genes identified in genome sequencing projects, interpreting data from proteomics projects and annotating protein sequence and structural databases. In addition, information about the structures and functions of moonlighting proteins can be helpful in understanding how novel protein functional sites evolved on an ancient protein scaffold, which can also help in the design of proteins with novel functions. PMID:25324305

Glycan array data management at Consortium for Functional Glycomics.

PubMed

Venkataraman, Maha; Sasisekharan, Ram; Raman, Rahul

2015-01-01

Glycomics or the study of structure-function relationships of complex glycans has reshaped post-genomics biology. Glycans mediate fundamental biological functions via their specific interactions with a variety of proteins. Recognizing the importance of glycomics, large-scale research initiatives such as the Consortium for Functional Glycomics (CFG) were established to address these challenges. Over the past decade, the Consortium for Functional Glycomics (CFG) has generated novel reagents and technologies for glycomics analyses, which in turn have led to generation of diverse datasets. These datasets have contributed to understanding glycan diversity and structure-function relationships at molecular (glycan-protein interactions), cellular (gene expression and glycan analysis), and whole organism (mouse phenotyping) levels. Among these analyses and datasets, screening of glycan-protein interactions on glycan array platforms has gained much prominence and has contributed to cross-disciplinary realization of the importance of glycomics in areas such as immunology, infectious diseases, cancer biomarkers, etc. This manuscript outlines methodologies for capturing data from glycan array experiments and online tools to access and visualize glycan array data implemented at the CFG.

Functional assignment to JEV proteins using SVM.

PubMed

Sahoo, Ganesh Chandra; Dikhit, Manas Ranjan; Das, Pradeep

2008-01-01

Identification of different protein functions facilitates a mechanistic understanding of Japanese encephalitis virus (JEV) infection and opens novel means for drug development. Support vector machines (SVM), useful for predicting the functional class of distantly related proteins, is employed to ascribe a possible functional class to Japanese encephalitis virus protein. Our study from SVMProt and available JE virus sequences suggests that structural and nonstructural proteins of JEV genome possibly belong to diverse protein functions, are expected to occur in the life cycle of JE virus. Protein functions common to both structural and non-structural proteins are iron-binding, metal-binding, lipid-binding, copper-binding, transmembrane, outer membrane, channels/Pores - Pore-forming toxins (proteins and peptides) group of proteins. Non-structural proteins perform functions like actin binding, zinc-binding, calcium-binding, hydrolases, Carbon-Oxygen Lyases, P-type ATPase, proteins belonging to major facilitator family (MFS), secreting main terminal branch (MTB) family, phosphotransfer-driven group translocators and ATP-binding cassette (ABC) family group of proteins. Whereas structural proteins besides belonging to same structural group of proteins (capsid, structural, envelope), they also perform functions like nuclear receptor, antibiotic resistance, RNA-binding, DNA-binding, magnesium-binding, isomerase (intra-molecular), oxidoreductase and participate in type II (general) secretory pathway (IISP).

Functional assignment to JEV proteins using SVM

PubMed Central

Sahoo, Ganesh Chandra; Dikhit, Manas Ranjan; Das, Pradeep

2008-01-01

Identification of different protein functions facilitates a mechanistic understanding of Japanese encephalitis virus (JEV) infection and opens novel means for drug development. Support vector machines (SVM), useful for predicting the functional class of distantly related proteins, is employed to ascribe a possible functional class to Japanese encephalitis virus protein. Our study from SVMProt and available JE virus sequences suggests that structural and nonstructural proteins of JEV genome possibly belong to diverse protein functions, are expected to occur in the life cycle of JE virus. Protein functions common to both structural and non-structural proteins are iron-binding, metal-binding, lipid-binding, copper-binding, transmembrane, outer membrane, channels/Pores - Pore-forming toxins (proteins and peptides) group of proteins. Non-structural proteins perform functions like actin binding, zinc-binding, calcium-binding, hydrolases, Carbon-Oxygen Lyases, P-type ATPase, proteins belonging to major facilitator family (MFS), secreting main terminal branch (MTB) family, phosphotransfer-driven group translocators and ATP-binding cassette (ABC) family group of proteins. Whereas structural proteins besides belonging to same structural group of proteins (capsid, structural, envelope), they also perform functions like nuclear receptor, antibiotic resistance, RNA-binding, DNA-binding, magnesium-binding, isomerase (intra-molecular), oxidoreductase and participate in type II (general) secretory pathway (IISP). PMID:19052658

Functional Cell Surface Display and Controlled Secretion of Diverse Agarolytic Enzymes by Escherichia coli with a Novel Ligation-Independent Cloning Vector Based on the Autotransporter YfaL

PubMed Central

Ko, Hyeok-Jin; Park, Eunhye; Song, Joseph; Yang, Taek Ho; Lee, Hee Jong; Kim, Kyoung Heon

2012-01-01

Autotransporters have been employed as the anchoring scaffold for cell surface display by replacing their passenger domains with heterologous proteins to be displayed. We adopted an autotransporter (YfaL) of Escherichia coli for the cell surface display system. The critical regions in YfaL for surface display were identified for the construction of a ligation-independent cloning (LIC)-based display system. The designed system showed no detrimental effect on either the growth of the host cell or overexpressing heterologous proteins on the cell surface. We functionally displayed monomeric red fluorescent protein (mRFP1) as a reporter protein and diverse agarolytic enzymes from Saccharophagus degradans 2-40, including Aga86C and Aga86E, which previously had failed to be functional expressed. The system could display different sizes of proteins ranging from 25.3 to 143 kDa. We also attempted controlled release of the displayed proteins by incorporating a tobacco etch virus protease cleavage site into the C termini of the displayed proteins. The maximum level of the displayed protein was 6.1 × 104 molecules per a single cell, which corresponds to 5.6% of the entire cell surface of actively growing E. coli. PMID:22344647

Protein Science by DNA Sequencing: How Advances in Molecular Biology Are Accelerating Biochemistry.

PubMed

Higgins, Sean A; Savage, David F

2018-01-09

A fundamental goal of protein biochemistry is to determine the sequence-function relationship, but the vastness of sequence space makes comprehensive evaluation of this landscape difficult. However, advances in DNA synthesis and sequencing now allow researchers to assess the functional impact of every single mutation in many proteins, but challenges remain in library construction and the development of general assays applicable to a diverse range of protein functions. This Perspective briefly outlines the technical innovations in DNA manipulation that allow massively parallel protein biochemistry and then summarizes the methods currently available for library construction and the functional assays of protein variants. Areas in need of future innovation are highlighted with a particular focus on assay development and the use of computational analysis with machine learning to effectively traverse the sequence-function landscape. Finally, applications in the fundamentals of protein biochemistry, disease prediction, and protein engineering are presented.

Diversification of HP1-like Chromo Domain Proteins in Tetrahymena thermophila.

PubMed

Wiley, Emily A; Horrell, Scott; Yoshino, Alyssa; Schornak, Cara C; Bagnani, Claire; Chalker, Douglas L

2018-01-01

Proteins that possess a chromo domain are well-known for their roles in heterochromatin assembly and maintenance. The Heterochromatin Protein 1 (HP1) family, with a chromo domain and carboxy-terminal chromo shadow domain, targets heterochromatin through interaction with histone H3 methylated on lysine 9 (H3K9me2/3). The structural and functional diversity of these proteins observed in both fission yeast and metazoans correlate with chromatin specialization. To expand these studies, we examined chromo domain proteins in the ciliate Tetrahymena thermophila, which has functionally diverse and developmentally regulated heterochromatin domains. We identified thirteen proteins similar to HP1. Together they possess only a fraction of the possible chromo domain subtypes and most lack a recognizable chromo shadow domain. Using fluorescence microscopy to track chromatin localization of tagged proteins through the life cycle, we show evidence that in T. thermophila this family has diversified with biological roles in RNAi-directed DNA elimination, germline genome structure, and somatic heterochromatin. Those proteins with H3K27me3 binding sequence characteristics localize to chromatin in mature nuclei, whereas those with H3K9me2/3 binding characteristics localize to developing nuclei undergoing DNA elimination. Findings point to an expanded and diversified family of chromo domain proteins that parallels heterochromatin diversity in ciliates. © 2017 The Authors. Journal of Eukaryotic Microbiology published by Wiley Periodicals, Inc. on behalf of International Society of Protistologists.

Changes in translation rate modulate stress-induced damage of diverse proteins

PubMed Central

Kim, Heejung

2013-01-01

Proteostasis is the maintenance of the proper function of cellular proteins. Hypertonic stress disrupts proteostasis and causes rapid and widespread protein aggregation and misfolding in the nematode Caenorhabditis elegans. Optimal survival in hypertonic environments requires degradation of damaged proteins. Inhibition of protein synthesis occurs in response to diverse environmental stressors and may function in part to minimize stress-induced protein damage. We recently tested this idea directly and demonstrated that translation inhibition by acute exposure to cycloheximide suppresses hypertonicity-induced aggregation of polyglutamine::YFP (Q35::YFP) in body wall muscle cells. In this article, we further characterized the relationship between protein synthesis and hypertonic stress-induced protein damage. We demonstrate that inhibition of translation reduces hypertonic stress-induced formation and growth of Q35::YFP, Q44::YFP, and α-synuclein aggregates; misfolding of paramyosin and ras GTPase; and aggregation of multiple endogenous proteins expressed in diverse cell types. Activation of general control nonderepressible-2 (GCN-2) kinase signaling during hypertonic stress inhibits protein synthesis via phosphorylation of eukaryotic initiation factor-2α (eIF-2α). Inhibition of GCN-2 activation prevents the reduction in translation rate and greatly exacerbates the formation and growth of Q35::YFP aggregates and the aggregation of endogenous proteins. The current studies together with our previous work provide the first direct demonstration that hypertonic stress-induced reduction in protein synthesis minimizes protein aggregation and misfolding. Reduction in translation rate also serves as a signal that activates osmoprotective gene expression. The cellular proteostasis network thus plays a critical role in minimizing hypertonic stress-induced protein damage, in degrading stress-damaged proteins, and in cellular osmosensing and signaling. PMID:24153430

Seven Transmembrane Receptors as Shapeshifting Proteins: The Impact of Allosteric Modulation and Functional Selectivity on New Drug Discovery

PubMed Central

Miller, Laurence J.

2010-01-01

It is useful to consider seven transmembrane receptors (7TMRs) as disordered proteins able to allosterically respond to a number of binding partners. Considering 7TMRs as allosteric systems, affinity and efficacy can be thought of in terms of energy flow between a modulator, conduit (the receptor protein), and a number of guests. These guests can be other molecules, receptors, membrane-bound proteins, or signaling proteins in the cytosol. These vectorial flows of energy can yield standard canonical guest allostery (allosteric modification of drug effect), effects along the plane of the cell membrane (receptor oligomerization), or effects directed into the cytosol (differential signaling as functional selectivity). This review discusses these apparently diverse pharmacological effects in terms of molecular dynamics and protein ensemble theory, which tends to unify 7TMR behavior toward cells. Special consideration will be given to functional selectivity (biased agonism and biased antagonism) in terms of mechanism of action and potential therapeutic application. The explosion of technology that has enabled observation of diverse 7TMR behavior has also shown how drugs can have multiple (pluridimensional) efficacies and how this can cause paradoxical drug classification and nomenclatures. PMID:20392808

Computational approaches for de novo design and redesign of metal-binding sites on proteins.

PubMed

Akcapinar, Gunseli Bayram; Sezerman, Osman Ugur

2017-04-28

Metal ions play pivotal roles in protein structure, function and stability. The functional and structural diversity of proteins in nature expanded with the incorporation of metal ions or clusters in proteins. Approximately one-third of these proteins in the databases contain metal ions. Many biological and chemical processes in nature involve metal ion-binding proteins, aka metalloproteins. Many cellular reactions that underpin life require metalloproteins. Most of the remarkable, complex chemical transformations are catalysed by metalloenzymes. Realization of the importance of metal-binding sites in a variety of cellular events led to the advancement of various computational methods for their prediction and characterization. Furthermore, as structural and functional knowledgebase about metalloproteins is expanding with advances in computational and experimental fields, the focus of the research is now shifting towards de novo design and redesign of metalloproteins to extend nature's own diversity beyond its limits. In this review, we will focus on the computational toolbox for prediction of metal ion-binding sites, de novo metalloprotein design and redesign. We will also give examples of tailor-made artificial metalloproteins designed with the computational toolbox. © 2017 The Author(s).

Novel Rhizosphere Soil Alleles for the Enzyme 1-Aminocyclopropane-1-Carboxylate Deaminase Queried for Function with an In Vivo Competition Assay

PubMed Central

Jin, Zhao; Di Rienzi, Sara C.; Janzon, Anders; Werner, Jeff J.; Angenent, Largus T.; Dangl, Jeffrey L.; Fowler, Douglas M.

2015-01-01

Metagenomes derived from environmental microbiota encode a vast diversity of protein homologs. How this diversity impacts protein function can be explored through selection assays aimed to optimize function. While artificially generated gene sequence pools are typically used in selection assays, their usage may be limited because of technical or ethical reasons. Here, we investigate an alternative strategy, the use of soil microbial DNA as a starting point. We demonstrate this approach by optimizing the function of a widely occurring soil bacterial enzyme, 1-aminocyclopropane-1-carboxylate (ACC) deaminase. We identified a specific ACC deaminase domain region (ACCD-DR) that, when PCR amplified from the soil, produced a variant pool that we could swap into functional plasmids carrying ACC deaminase-encoding genes. Functional clones of ACC deaminase were selected for in a competition assay based on their capacity to provide nitrogen to Escherichia coli in vitro. The most successful ACCD-DR variants were identified after multiple rounds of selection by sequence analysis. We observed that previously identified essential active-site residues were fixed in the original unselected library and that additional residues went to fixation after selection. We identified a divergent essential residue whose presence hints at the possible use of alternative substrates and a cluster of neutral residues that did not influence ACCD performance. Using an artificial ACCD-DR variant library generated by DNA oligomer synthesis, we validated the same fixation patterns. Our study demonstrates that soil metagenomes are useful starting pools of protein-coding-gene diversity that can be utilized for protein optimization and functional characterization when synthetic libraries are not appropriate. PMID:26637602

A Global Overview of the Genetic and Functional Diversity in the Helicobacter pylori cag Pathogenicity Island

PubMed Central

Moodley, Yoshan; Uhr, Markus; Stamer, Christiana; Vauterin, Marc; Suerbaum, Sebastian; Achtman, Mark

2010-01-01

The Helicobacter pylori cag pathogenicity island (cagPAI) encodes a type IV secretion system. Humans infected with cagPAI–carrying H. pylori are at increased risk for sequelae such as gastric cancer. Housekeeping genes in H. pylori show considerable genetic diversity; but the diversity of virulence factors such as the cagPAI, which transports the bacterial oncogene CagA into host cells, has not been systematically investigated. Here we compared the complete cagPAI sequences for 38 representative isolates from all known H. pylori biogeographic populations. Their gene content and gene order were highly conserved. The phylogeny of most cagPAI genes was similar to that of housekeeping genes, indicating that the cagPAI was probably acquired only once by H. pylori, and its genetic diversity reflects the isolation by distance that has shaped this bacterial species since modern humans migrated out of Africa. Most isolates induced IL-8 release in gastric epithelial cells, indicating that the function of the Cag secretion system has been conserved despite some genetic rearrangements. More than one third of cagPAI genes, in particular those encoding cell-surface exposed proteins, showed signatures of diversifying (Darwinian) selection at more than 5% of codons. Several unknown gene products predicted to be under Darwinian selection are also likely to be secreted proteins (e.g. HP0522, HP0535). One of these, HP0535, is predicted to code for either a new secreted candidate effector protein or a protein which interacts with CagA because it contains two genetic lineages, similar to cagA. Our study provides a resource that can guide future research on the biological roles and host interactions of cagPAI proteins, including several whose function is still unknown. PMID:20808891

A global overview of the genetic and functional diversity in the Helicobacter pylori cag pathogenicity island.

PubMed

Olbermann, Patrick; Josenhans, Christine; Moodley, Yoshan; Uhr, Markus; Stamer, Christiana; Vauterin, Marc; Suerbaum, Sebastian; Achtman, Mark; Linz, Bodo

2010-08-19

The Helicobacter pylori cag pathogenicity island (cagPAI) encodes a type IV secretion system. Humans infected with cagPAI-carrying H. pylori are at increased risk for sequelae such as gastric cancer. Housekeeping genes in H. pylori show considerable genetic diversity; but the diversity of virulence factors such as the cagPAI, which transports the bacterial oncogene CagA into host cells, has not been systematically investigated. Here we compared the complete cagPAI sequences for 38 representative isolates from all known H. pylori biogeographic populations. Their gene content and gene order were highly conserved. The phylogeny of most cagPAI genes was similar to that of housekeeping genes, indicating that the cagPAI was probably acquired only once by H. pylori, and its genetic diversity reflects the isolation by distance that has shaped this bacterial species since modern humans migrated out of Africa. Most isolates induced IL-8 release in gastric epithelial cells, indicating that the function of the Cag secretion system has been conserved despite some genetic rearrangements. More than one third of cagPAI genes, in particular those encoding cell-surface exposed proteins, showed signatures of diversifying (Darwinian) selection at more than 5% of codons. Several unknown gene products predicted to be under Darwinian selection are also likely to be secreted proteins (e.g. HP0522, HP0535). One of these, HP0535, is predicted to code for either a new secreted candidate effector protein or a protein which interacts with CagA because it contains two genetic lineages, similar to cagA. Our study provides a resource that can guide future research on the biological roles and host interactions of cagPAI proteins, including several whose function is still unknown.

The Ubiquitin–Proteasome System of Saccharomyces cerevisiae

PubMed Central

Finley, Daniel; Ulrich, Helle D.; Sommer, Thomas; Kaiser, Peter

2012-01-01

Protein modifications provide cells with exquisite temporal and spatial control of protein function. Ubiquitin is among the most important modifiers, serving both to target hundreds of proteins for rapid degradation by the proteasome, and as a dynamic signaling agent that regulates the function of covalently bound proteins. The diverse effects of ubiquitylation reflect the assembly of structurally distinct ubiquitin chains on target proteins. The resulting ubiquitin code is interpreted by an extensive family of ubiquitin receptors. Here we review the components of this regulatory network and its effects throughout the cell. PMID:23028185

Trafficking and function of the cystic fibrosis transmembrane conductance regulator: a complex network of posttranslational modifications

PubMed Central

McClure, Michelle L.; Barnes, Stephen; Brodsky, Jeffrey L.

2016-01-01

Posttranslational modifications add diversity to protein function. Throughout its life cycle, the cystic fibrosis transmembrane conductance regulator (CFTR) undergoes numerous covalent posttranslational modifications (PTMs), including glycosylation, ubiquitination, sumoylation, phosphorylation, and palmitoylation. These modifications regulate key steps during protein biogenesis, such as protein folding, trafficking, stability, function, and association with protein partners and therefore may serve as targets for therapeutic manipulation. More generally, an improved understanding of molecular mechanisms that underlie CFTR PTMs may suggest novel treatment strategies for CF and perhaps other protein conformational diseases. This review provides a comprehensive summary of co- and posttranslational CFTR modifications and their significance with regard to protein biogenesis. PMID:27474090

Identification of a functionally distinct truncated BDNF mRNA splice variant and protein in Trachemys scripta elegans.

PubMed

Ambigapathy, Ganesh; Zheng, Zhaoqing; Li, Wei; Keifer, Joyce

2013-01-01

Brain-derived neurotrophic factor (BDNF) has a diverse functional role and complex pattern of gene expression. Alternative splicing of mRNA transcripts leads to further diversity of mRNAs and protein isoforms. Here, we describe the regulation of BDNF mRNA transcripts in an in vitro model of eyeblink classical conditioning and a unique transcript that forms a functionally distinct truncated BDNF protein isoform. Nine different mRNA transcripts from the BDNF gene of the pond turtle Trachemys scripta elegans (tBDNF) are selectively regulated during classical conditioning: exon I mRNA transcripts show no change, exon II transcripts are downregulated, while exon III transcripts are upregulated. One unique transcript that codes from exon II, tBDNF2a, contains a 40 base pair deletion in the protein coding exon that generates a truncated tBDNF protein. The truncated transcript and protein are expressed in the naïve untrained state and are fully repressed during conditioning when full-length mature tBDNF is expressed, thereby having an alternate pattern of expression in conditioning. Truncated BDNF is not restricted to turtles as a truncated mRNA splice variant has been described for the human BDNF gene. Further studies are required to determine the ubiquity of truncated BDNF alternative splice variants across species and the mechanisms of regulation and function of this newly recognized BDNF protein.

Identification of a Functionally Distinct Truncated BDNF mRNA Splice Variant and Protein in Trachemys scripta elegans

PubMed Central

Ambigapathy, Ganesh; Zheng, Zhaoqing; Li, Wei; Keifer, Joyce

2013-01-01

Brain-derived neurotrophic factor (BDNF) has a diverse functional role and complex pattern of gene expression. Alternative splicing of mRNA transcripts leads to further diversity of mRNAs and protein isoforms. Here, we describe the regulation of BDNF mRNA transcripts in an in vitro model of eyeblink classical conditioning and a unique transcript that forms a functionally distinct truncated BDNF protein isoform. Nine different mRNA transcripts from the BDNF gene of the pond turtle Trachemys scripta elegans (tBDNF) are selectively regulated during classical conditioning: exon I mRNA transcripts show no change, exon II transcripts are downregulated, while exon III transcripts are upregulated. One unique transcript that codes from exon II, tBDNF2a, contains a 40 base pair deletion in the protein coding exon that generates a truncated tBDNF protein. The truncated transcript and protein are expressed in the naïve untrained state and are fully repressed during conditioning when full-length mature tBDNF is expressed, thereby having an alternate pattern of expression in conditioning. Truncated BDNF is not restricted to turtles as a truncated mRNA splice variant has been described for the human BDNF gene. Further studies are required to determine the ubiquity of truncated BDNF alternative splice variants across species and the mechanisms of regulation and function of this newly recognized BDNF protein. PMID:23825634

Diversity and functions of protein glycosylation in insects.

PubMed

Walski, Tomasz; De Schutter, Kristof; Van Damme, Els J M; Smagghe, Guy

2017-04-01

The majority of proteins is modified with carbohydrate structures. This modification, called glycosylation, was shown to be crucial for protein folding, stability and subcellular location, as well as protein-protein interactions, recognition and signaling. Protein glycosylation is involved in multiple physiological processes, including embryonic development, growth, circadian rhythms, cell attachment as well as maintenance of organ structure, immunity and fertility. Although the general principles of glycosylation are similar among eukaryotic organisms, insects synthesize a distinct repertoire of glycan structures compared to plants and vertebrates. Consequently, a number of unique insect glycans mediate functions specific to this class of invertebrates. For instance, the core α1,3-fucosylation of N-glycans is absent in vertebrates, while in insects this modification is crucial for the development of wings and the nervous system. At present, most of the data on insect glycobiology comes from research in Drosophila. Yet, progressively more information on the glycan structures and the importance of glycosylation in other insects like beetles, caterpillars, aphids and bees is becoming available. This review gives a summary of the current knowledge and recent progress related to glycan diversity and function(s) of protein glycosylation in insects. We focus on N- and O-glycosylation, their synthesis, physiological role(s), as well as the molecular and biochemical basis of these processes. Copyright © 2017 Elsevier Ltd. All rights reserved.

Insights into Hox protein function from a large scale combinatorial analysis of protein domains.

PubMed

Merabet, Samir; Litim-Mecheri, Isma; Karlsson, Daniel; Dixit, Richa; Saadaoui, Mehdi; Monier, Bruno; Brun, Christine; Thor, Stefan; Vijayraghavan, K; Perrin, Laurent; Pradel, Jacques; Graba, Yacine

2011-10-01

Protein function is encoded within protein sequence and protein domains. However, how protein domains cooperate within a protein to modulate overall activity and how this impacts functional diversification at the molecular and organism levels remains largely unaddressed. Focusing on three domains of the central class Drosophila Hox transcription factor AbdominalA (AbdA), we used combinatorial domain mutations and most known AbdA developmental functions as biological readouts to investigate how protein domains collectively shape protein activity. The results uncover redundancy, interactivity, and multifunctionality of protein domains as salient features underlying overall AbdA protein activity, providing means to apprehend functional diversity and accounting for the robustness of Hox-controlled developmental programs. Importantly, the results highlight context-dependency in protein domain usage and interaction, allowing major modifications in domains to be tolerated without general functional loss. The non-pleoitropic effect of domain mutation suggests that protein modification may contribute more broadly to molecular changes underlying morphological diversification during evolution, so far thought to rely largely on modification in gene cis-regulatory sequences.

Insights into Hox Protein Function from a Large Scale Combinatorial Analysis of Protein Domains

PubMed Central

Karlsson, Daniel; Dixit, Richa; Saadaoui, Mehdi; Monier, Bruno; Brun, Christine; Thor, Stefan; Vijayraghavan, K.; Perrin, Laurent; Pradel, Jacques; Graba, Yacine

2011-01-01

Protein function is encoded within protein sequence and protein domains. However, how protein domains cooperate within a protein to modulate overall activity and how this impacts functional diversification at the molecular and organism levels remains largely unaddressed. Focusing on three domains of the central class Drosophila Hox transcription factor AbdominalA (AbdA), we used combinatorial domain mutations and most known AbdA developmental functions as biological readouts to investigate how protein domains collectively shape protein activity. The results uncover redundancy, interactivity, and multifunctionality of protein domains as salient features underlying overall AbdA protein activity, providing means to apprehend functional diversity and accounting for the robustness of Hox-controlled developmental programs. Importantly, the results highlight context-dependency in protein domain usage and interaction, allowing major modifications in domains to be tolerated without general functional loss. The non-pleoitropic effect of domain mutation suggests that protein modification may contribute more broadly to molecular changes underlying morphological diversification during evolution, so far thought to rely largely on modification in gene cis-regulatory sequences. PMID:22046139

Using Signature Genes as Tools To Assess Environmental Viral Ecology and Diversity

PubMed Central

Adriaenssens, Evelien M.

2014-01-01

Viruses (including bacteriophages) are the most abundant biological entities on the planet. As such, they are thought to have a major impact on all aspects of microbial community structure and function. Despite this critical role in ecosystem processes, the study of virus/phage diversity has lagged far behind parallel studies of the bacterial and eukaryotic kingdoms, largely due to the absence of any universal phylogenetic marker. Here we review the development and use of signature genes to investigate viral diversity, as a viable strategy for data sets of specific virus groups. Genes that have been used include those encoding structural proteins, such as portal protein, major capsid protein, and tail sheath protein, auxiliary metabolism genes, such as psbA, psbB, and phoH, and several polymerase genes. These marker genes have been used in combination with PCR-based fingerprinting and/or sequencing strategies to investigate spatial, temporal, and seasonal variations and diversity in a wide range of habitats. PMID:24837394

Molecular mimicry between protein and tRNA.

PubMed

Nakamura, Y

2001-01-01

Mimicry is a sophisticated development in animals, fish, and plants that allows them to fool others by imitating a shape or color for diverse purposes, such as to prey, evade, lure, pollinate, or threaten. This is not restricted to the macro-world, but extends to the micro-world as molecular mimicry. Recent advances in structural and molecular biology uncovered a set of translation factors that resembles a tRNA shape and, in one case, even mimics a tRNA function for deciphering the genetic code. Nature must have evolved this art of molecular mimicry between protein and ribonucleic acid by using different protein structures until the translation factors sat in the cockpit of a ribosome machine, on behalf of tRNA, and achieved diverse actions. Structural, functional, and evolutionary aspects of molecular mimicry will be discussed.

JNK Signaling: Regulation and Functions Based on Complex Protein-Protein Partnerships

PubMed Central

Zeke, András; Misheva, Mariya

2016-01-01

SUMMARY The c-Jun N-terminal kinases (JNKs), as members of the mitogen-activated protein kinase (MAPK) family, mediate eukaryotic cell responses to a wide range of abiotic and biotic stress insults. JNKs also regulate important physiological processes, including neuronal functions, immunological actions, and embryonic development, via their impact on gene expression, cytoskeletal protein dynamics, and cell death/survival pathways. Although the JNK pathway has been under study for >20 years, its complexity is still perplexing, with multiple protein partners of JNKs underlying the diversity of actions. Here we review the current knowledge of JNK structure and isoforms as well as the partnerships of JNKs with a range of intracellular proteins. Many of these proteins are direct substrates of the JNKs. We analyzed almost 100 of these target proteins in detail within a framework of their classification based on their regulation by JNKs. Examples of these JNK substrates include a diverse assortment of nuclear transcription factors (Jun, ATF2, Myc, Elk1), cytoplasmic proteins involved in cytoskeleton regulation (DCX, Tau, WDR62) or vesicular transport (JIP1, JIP3), cell membrane receptors (BMPR2), and mitochondrial proteins (Mcl1, Bim). In addition, because upstream signaling components impact JNK activity, we critically assessed the involvement of signaling scaffolds and the roles of feedback mechanisms in the JNK pathway. Despite a clarification of many regulatory events in JNK-dependent signaling during the past decade, many other structural and mechanistic insights are just beginning to be revealed. These advances open new opportunities to understand the role of JNK signaling in diverse physiological and pathophysiological states. PMID:27466283

Elucidating the ensemble of functionally-relevant transitions in protein systems with a robotics-inspired method.

PubMed

Molloy, Kevin; Shehu, Amarda

2013-01-01

Many proteins tune their biological function by transitioning between different functional states, effectively acting as dynamic molecular machines. Detailed structural characterization of transition trajectories is central to understanding the relationship between protein dynamics and function. Computational approaches that build on the Molecular Dynamics framework are in principle able to model transition trajectories at great detail but also at considerable computational cost. Methods that delay consideration of dynamics and focus instead on elucidating energetically-credible conformational paths connecting two functionally-relevant structures provide a complementary approach. Effective sampling-based path planning methods originating in robotics have been recently proposed to produce conformational paths. These methods largely model short peptides or address large proteins by simplifying conformational space. We propose a robotics-inspired method that connects two given structures of a protein by sampling conformational paths. The method focuses on small- to medium-size proteins, efficiently modeling structural deformations through the use of the molecular fragment replacement technique. In particular, the method grows a tree in conformational space rooted at the start structure, steering the tree to a goal region defined around the goal structure. We investigate various bias schemes over a progress coordinate for balance between coverage of conformational space and progress towards the goal. A geometric projection layer promotes path diversity. A reactive temperature scheme allows sampling of rare paths that cross energy barriers. Experiments are conducted on small- to medium-size proteins of length up to 214 amino acids and with multiple known functionally-relevant states, some of which are more than 13Å apart of each-other. Analysis reveals that the method effectively obtains conformational paths connecting structural states that are significantly different. A detailed analysis on the depth and breadth of the tree suggests that a soft global bias over the progress coordinate enhances sampling and results in higher path diversity. The explicit geometric projection layer that biases the exploration away from over-sampled regions further increases coverage, often improving proximity to the goal by forcing the exploration to find new paths. The reactive temperature scheme is shown effective in increasing path diversity, particularly in difficult structural transitions with known high-energy barriers.

A widely employed germ cell marker is an ancient disordered protein with reproductive functions in diverse eukaryotes

PubMed Central

Carmell, Michelle A; Dokshin, Gregoriy A; Skaletsky, Helen; Hu, Yueh-Chiang; van Wolfswinkel, Josien C; Igarashi, Kyomi J; Bellott, Daniel W; Nefedov, Michael; Reddien, Peter W; Enders, George C; Uversky, Vladimir N; Mello, Craig C; Page, David C

2016-01-01

The advent of sexual reproduction and the evolution of a dedicated germline in multicellular organisms are critical landmarks in eukaryotic evolution. We report an ancient family of GCNA (germ cell nuclear antigen) proteins that arose in the earliest eukaryotes, and feature a rapidly evolving intrinsically disordered region (IDR). Phylogenetic analysis reveals that GCNA proteins emerged before the major eukaryotic lineages diverged; GCNA predates the origin of a dedicated germline by a billion years. Gcna gene expression is enriched in reproductive cells across eukarya – either just prior to or during meiosis in single-celled eukaryotes, and in stem cells and germ cells of diverse multicellular animals. Studies of Gcna-mutant C. elegans and mice indicate that GCNA has functioned in reproduction for at least 600 million years. Homology to IDR-containing proteins implicated in DNA damage repair suggests that GCNA proteins may protect the genomic integrity of cells carrying a heritable genome. DOI: http://dx.doi.org/10.7554/eLife.19993.001 PMID:27718356

Density functional study of molecular interactions in secondary structures of proteins.

PubMed

Takano, Yu; Kusaka, Ayumi; Nakamura, Haruki

2016-01-01

Proteins play diverse and vital roles in biology, which are dominated by their three-dimensional structures. The three-dimensional structure of a protein determines its functions and chemical properties. Protein secondary structures, including α-helices and β-sheets, are key components of the protein architecture. Molecular interactions, in particular hydrogen bonds, play significant roles in the formation of protein secondary structures. Precise and quantitative estimations of these interactions are required to understand the principles underlying the formation of three-dimensional protein structures. In the present study, we have investigated the molecular interactions in α-helices and β-sheets, using ab initio wave function-based methods, the Hartree-Fock method (HF) and the second-order Møller-Plesset perturbation theory (MP2), density functional theory, and molecular mechanics. The characteristic interactions essential for forming the secondary structures are discussed quantitatively.

Mini Heme-Proteins: Designability of Structure and Diversity of Functions.

PubMed

Rai, Jagdish

2017-08-30

Natural heme proteins may have heme bound to poly-peptide chain as a cofactor via noncovalent forces or heme as a prosthetic group may be covalently bound to the proteins. Nature has used porphyrins in diverse functions like electron transfer, oxidation, reduction, ligand binding, photosynthesis, signaling, etc. by modulating its properties through diverse protein matrices. Synthetic chemists have tried to utilize these molecules in equally diverse industrial and medical applications due to their versatile electro-chemical and optical properties. The heme iron has catalytic activity which can be modulated and enhanced for specific applications by protein matrix around it. Heme proteins can be designed into novel enzymes for sterio specific catalysis ranging from oxidation to reduction. These designed heme-proteins can have applications in industrial catalysis and biosensing. A peptide folds around heme easily due to hydrophobic effect of the large aromatic ring of heme. The directional property of co-ordinate bonding between peptide and metal ion in heme further specifies the structure. Therefore heme proteins can be easily designed for targeted structure and catalytic activity. The central aromatic chemical entity in heme viz. porphyrin is a very ancient molecule. Its presence in the prebiotic soup and in all forms of life suggests that it has played a vital role in the origin and progressive evolution of living organisms. Porphyrin macrocycles are highly conjugated systems composed of four modified pyrrole subunits interconnected at their α -carbon atoms via methine (=CH-) bridges. Initial minimalist models of hemoproteins focused on effect of heme-ligand co-ordinate bonding on chemical reactivity, spectroscopy, electrochemistry and magnetic properties of heme. The great sensitivity of these spectroscopic features of heme to its surrounding makes them extremely useful in structural elucidation of designed heme-peptide complexes. Therefore heme proteins are easier to work on for designing novel proteins for industrial and medical applications. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Double-stranded telomeric DNA binding proteins: Diversity matters.

PubMed

Červenák, Filip; Juríková, Katarína; Sepšiová, Regina; Neboháčová, Martina; Nosek, Jozef; Tomáška, L'ubomír

2017-01-01

Telomeric sequences constitute only a small fraction of the whole genome yet they are crucial for ensuring genomic stability. This function is in large part mediated by protein complexes recruited to telomeric sequences by specific telomere-binding proteins (TBPs). Although the principal tasks of nuclear telomeres are the same in all eukaryotes, TBPs in various taxa exhibit a surprising diversity indicating their distinct evolutionary origin. This diversity is especially pronounced in ascomycetous yeasts where they must have co-evolved with rapidly diversifying sequences of telomeric repeats. In this article we (i) provide a historical overview of the discoveries leading to the current list of TBPs binding to double-stranded (ds) regions of telomeres, (ii) describe examples of dsTBPs highlighting their diversity in even closely related species, and (iii) speculate about possible evolutionary trajectories leading to a long list of various dsTBPs fulfilling the same general role(s) in their own unique ways.

Evolutionary turnover of kinetochore proteins: a ship of Theseus?

PubMed Central

Drinnenberg, Ines A.; Henikoff, Steven; Malik, Harmit S.

2016-01-01

Summary The kinetochore is a multi-protein complex that mediates the attachment of a eukaryotic chromosome to the mitotic spindle. The protein composition of kinetochores is similar across species as divergent as yeast and human. However, recent findings have revealed an unexpected degree of compositional diversity in kinetochores. For example, kinetochore proteins that are essential in some species have been lost in others, whereas new kinetochore proteins have emerged in other lineages. Even in lineages with similar kinetochore composition, individual kinetochore proteins have functionally diverged to acquire either essential or redundant roles. Thus, despite functional conservation, the repertoire of kinetochore proteins has undergone recurrent evolutionary turnover. PMID:26877204

The logic of DNA replication in double-stranded DNA viruses: insights from global analysis of viral genomes

PubMed Central

Kazlauskas, Darius; Krupovic, Mart; Venclovas, Česlovas

2016-01-01

Abstract Genomic DNA replication is a complex process that involves multiple proteins. Cellular DNA replication systems are broadly classified into only two types, bacterial and archaeo-eukaryotic. In contrast, double-stranded (ds) DNA viruses feature a much broader diversity of DNA replication machineries. Viruses differ greatly in both completeness and composition of their sets of DNA replication proteins. In this study, we explored whether there are common patterns underlying this extreme diversity. We identified and analyzed all major functional groups of DNA replication proteins in all available proteomes of dsDNA viruses. Our results show that some proteins are common to viruses infecting all domains of life and likely represent components of the ancestral core set. These include B-family polymerases, SF3 helicases, archaeo-eukaryotic primases, clamps and clamp loaders of the archaeo-eukaryotic type, RNase H and ATP-dependent DNA ligases. We also discovered a clear correlation between genome size and self-sufficiency of viral DNA replication, the unanticipated dominance of replicative helicases and pervasive functional associations among certain groups of DNA replication proteins. Altogether, our results provide a comprehensive view on the diversity and evolution of replication systems in the DNA virome and uncover fundamental principles underlying the orchestration of viral DNA replication. PMID:27112572

Dynamic protein S-palmitoylation mediates parasite life cycle progression and diverse mechanisms of virulence.

PubMed

Brown, Robert W B; Sharma, Aabha I; Engman, David M

2017-04-01

Eukaryotic parasites possess complex life cycles and utilize an assortment of molecular mechanisms to overcome physical barriers, suppress and/or bypass the host immune response, including invading host cells where they can replicate in a protected intracellular niche. Protein S-palmitoylation is a dynamic post-translational modification in which the fatty acid palmitate is covalently linked to cysteine residues on proteins by the enzyme palmitoyl acyltransferase (PAT) and can be removed by lysosomal palmitoyl-protein thioesterase (PPT) or cytosolic acyl-protein thioesterase (APT). In addition to anchoring proteins to intracellular membranes, functions of dynamic palmitoylation include - targeting proteins to specific intracellular compartments via trafficking pathways, regulating the cycling of proteins between membranes, modulating protein function and regulating protein stability. Recent studies in the eukaryotic parasites - Plasmodium falciparum, Toxoplasma gondii, Trypanosoma brucei, Cryptococcus neoformans and Giardia lamblia - have identified large families of PATs and palmitoylated proteins. Many palmitoylated proteins are important for diverse aspects of pathogenesis, including differentiation into infective life cycle stages, biogenesis and tethering of secretory organelles, assembling the machinery powering motility and targeting virulence factors to the plasma membrane. This review aims to summarize our current knowledge of palmitoylation in eukaryotic parasites, highlighting five exemplary mechanisms of parasite virulence dependent on palmitoylation.

"Multiple partial recognitions in dynamic equilibrium" in the binding sites of proteins form the molecular basis of promiscuous recognition of structurally diverse ligands.

PubMed

Kohda, Daisuke

2018-04-01

Promiscuous recognition of ligands by proteins is as important as strict recognition in numerous biological processes. In living cells, many short, linear amino acid motifs function as targeting signals in proteins to specify the final destination of the protein transport. In general, the target signal is defined by a consensus sequence containing wild-characters, and hence represented by diverse amino acid sequences. The classical lock-and-key or induced-fit/conformational selection mechanism may not cover all aspects of the promiscuous recognition. On the basis of our crystallographic and NMR studies on the mitochondrial Tom20 protein-presequence interaction, we proposed a new hypothetical mechanism based on "a rapid equilibrium of multiple states with partial recognitions". This dynamic, multiple recognition mode enables the Tom20 receptor to recognize diverse mitochondrial presequences with nearly equal affinities. The plant Tom20 is evolutionally unrelated to the animal Tom20 in our study, but is a functional homolog of the animal/fungal Tom20. NMR studies by another research group revealed that the presequence binding by the plant Tom20 was not fully explained by simple interaction modes, suggesting the presence of a similar dynamic, multiple recognition mode. Circumstantial evidence also suggested that similar dynamic mechanisms may be applicable to other promiscuous recognitions of signal peptides by the SRP54/Ffh and SecA proteins.

pocketZebra: a web-server for automated selection and classification of subfamily-specific binding sites by bioinformatic analysis of diverse protein families.

PubMed

Suplatov, Dmitry; Kirilin, Eugeny; Arbatsky, Mikhail; Takhaveev, Vakil; Svedas, Vytas

2014-07-01

The new web-server pocketZebra implements the power of bioinformatics and geometry-based structural approaches to identify and rank subfamily-specific binding sites in proteins by functional significance, and select particular positions in the structure that determine selective accommodation of ligands. A new scoring function has been developed to annotate binding sites by the presence of the subfamily-specific positions in diverse protein families. pocketZebra web-server has multiple input modes to meet the needs of users with different experience in bioinformatics. The server provides on-site visualization of the results as well as off-line version of the output in annotated text format and as PyMol sessions ready for structural analysis. pocketZebra can be used to study structure-function relationship and regulation in large protein superfamilies, classify functionally important binding sites and annotate proteins with unknown function. The server can be used to engineer ligand-binding sites and allosteric regulation of enzymes, or implemented in a drug discovery process to search for potential molecular targets and novel selective inhibitors/effectors. The server, documentation and examples are freely available at http://biokinet.belozersky.msu.ru/pocketzebra and there are no login requirements. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

Exploration of Uncharted Regions of the Protein Universe

PubMed Central

Jaroszewski, Lukasz; Li, Zhanwen; Krishna, S. Sri; Bakolitsa, Constantina; Wooley, John; Deacon, Ashley M.; Wilson, Ian A.; Godzik, Adam

2009-01-01

The genome projects have unearthed an enormous diversity of genes of unknown function that are still awaiting biological and biochemical characterization. These genes, as most others, can be grouped into families based on sequence similarity. The PFAM database currently contains over 2,200 such families, referred to as domains of unknown function (DUF). In a coordinated effort, the four large-scale centers of the NIH Protein Structure Initiative have determined the first three-dimensional structures for more than 250 of these DUF families. Analysis of the first 248 reveals that about two thirds of the DUF families likely represent very divergent branches of already known and well-characterized families, which allows hypotheses to be formulated about their biological function. The remainder can be formally categorized as new folds, although about one third of these show significant substructure similarity to previously characterized folds. These results infer that, despite the enormous increase in the number and the diversity of new genes being uncovered, the fold space of the proteins they encode is gradually becoming saturated. The previously unexplored sectors of the protein universe appear to be primarily shaped by extreme diversification of known protein families, which then enables organisms to evolve new functions and adapt to particular niches and habitats. Notwithstanding, these DUF families still constitute the richest source for discovery of the remaining protein folds and topologies. PMID:19787035

The rendez-vous of mobile sieve-element and abundant companion-cell proteins.

PubMed

De Marco, Federica; Le Hir, Rozenn; Dinant, Sylvie

2018-06-01

Thousands of sieve tube exudate proteins (STEP) have now been identified and predicted to fulfill a diversity of functions. However, most STEPs should be considered putative, since methods to collect sieve tube exudates have many technical drawbacks, and advanced functional characterization will be required to distinguish contaminant from bonafide proteins, and determine the latter's location and activity in sieve elements (SE). One major challenge is to develop new approaches to elucidate the function of these SE proteins, which in turn, is expected to shed light on intriguing aspects of SE cell biology. Copyright © 2018 Elsevier Ltd. All rights reserved.

The secret life of kinases: insights into non-catalytic signalling functions from pseudokinases.

PubMed

Jacobsen, Annette V; Murphy, James M

2017-06-15

Over the past decade, our understanding of the mechanisms by which pseudokinases, which comprise ∼10% of the human and mouse kinomes, mediate signal transduction has advanced rapidly with increasing structural, biochemical, cellular and genetic studies. Pseudokinases are the catalytically defective counterparts of conventional, active protein kinases and have been attributed functions as protein interaction domains acting variously as allosteric modulators of conventional protein kinases and other enzymes, as regulators of protein trafficking or localisation, as hubs to nucleate assembly of signalling complexes, and as transmembrane effectors of such functions. Here, by categorising mammalian pseudokinases based on their known functions, we illustrate the mechanistic diversity among these proteins, which can be viewed as a window into understanding the non-catalytic functions that can be exerted by conventional protein kinases. © 2017 The Author(s); published by Portland Press Limited on behalf of the Biochemical Society.

Function of alternative splicing

PubMed Central

Kelemen, Olga; Convertini, Paolo; Zhang, Zhaiyi; Wen, Yuan; Shen, Manli; Falaleeva, Marina; Stamm, Stefan

2017-01-01

Almost all polymerase II transcripts undergo alternative pre-mRNA splicing. Here, we review the functions of alternative splicing events that have been experimentally determined. The overall function of alternative splicing is to increase the diversity of mRNAs expressed from the genome. Alternative splicing changes proteins encoded by mRNAs, which has profound functional effects. Experimental analysis of these protein isoforms showed that alternative splicing regulates binding between proteins, between proteins and nucleic acids as well as between proteins and membranes. Alternative splicing regulates the localization of proteins, their enzymatic properties and their interaction with ligands. In most cases, changes caused by individual splicing isoforms are small. However, cells typically coordinate numerous changes in ‘splicing programs’, which can have strong effects on cell proliferation, cell survival and properties of the nervous system. Due to its widespread usage and molecular versatility, alternative splicing emerges as a central element in gene regulation that interferes with almost every biological function analyzed. PMID:22909801

A Score of the Ability of a Three-Dimensional Protein Model to Retrieve Its Own Sequence as a Quantitative Measure of Its Quality and Appropriateness

PubMed Central

Martínez-Castilla, León P.; Rodríguez-Sotres, Rogelio

2010-01-01

Background Despite the remarkable progress of bioinformatics, how the primary structure of a protein leads to a three-dimensional fold, and in turn determines its function remains an elusive question. Alignments of sequences with known function can be used to identify proteins with the same or similar function with high success. However, identification of function-related and structure-related amino acid positions is only possible after a detailed study of every protein. Folding pattern diversity seems to be much narrower than sequence diversity, and the amino acid sequences of natural proteins have evolved under a selective pressure comprising structural and functional requirements acting in parallel. Principal Findings The approach described in this work begins by generating a large number of amino acid sequences using ROSETTA [Dantas G et al. (2003) J Mol Biol 332:449–460], a program with notable robustness in the assignment of amino acids to a known three-dimensional structure. The resulting sequence-sets showed no conservation of amino acids at active sites, or protein-protein interfaces. Hidden Markov models built from the resulting sequence sets were used to search sequence databases. Surprisingly, the models retrieved from the database sequences belonged to proteins with the same or a very similar function. Given an appropriate cutoff, the rate of false positives was zero. According to our results, this protocol, here referred to as Rd.HMM, detects fine structural details on the folding patterns, that seem to be tightly linked to the fitness of a structural framework for a specific biological function. Conclusion Because the sequence of the native protein used to create the Rd.HMM model was always amongst the top hits, the procedure is a reliable tool to score, very accurately, the quality and appropriateness of computer-modeled 3D-structures, without the need for spectroscopy data. However, Rd.HMM is very sensitive to the conformational features of the models' backbone. PMID:20830209

Phosphatidylenthanolamine Binding Protein aka Raf Kinase Inhibitor Protein: A Brief History of Its Discovery and the Remarkable Diversity of Biological Functions.

PubMed

Sedivy, John M

2011-01-01

Phosphatidylethanolamine-binding protein (PEBP) was identified almost three decades ago as an abundant protein in bovine brain. PEBP is the prototype of a highly conserved family of proteins represented in all three major phylogenetic divisions, eukaryota, bacteria, and archaea, with no significant sequence homology to other proteins. PEBP proteins have been studied in many species. The most thoroughly explored biological role of PEBP is that of a modulator of intracellular signaling pathways, which is mediated by its ability to bind and inhibit a number of protein kinases. The first such interaction that came to light was with the Raf1 kinase, and PEBP is thus widely referred to in the literature under its alternate name RKIP (Raf kinase inhibitory protein). The activity of RKIP itself is subject to regulation by phosphorylation. Intriguingly, PEBP has also been reported to possess additional, and diverse, biological functions unrelated to protein kinase networks that remain to be investigated in detail. Recent findings that RKIP may function as a suppressor of cancer metastasis are of great interest and importance. Prognostic and therapeutic applications of RKIP in human cancer were the subject of the first international workshop on RKIP that was held at the University of California, Los Angeles, in March 2010. This paper was presented at the workshop as a summary of the history of this still small but rapidly evolving field.

Robust enzyme design: bioinformatic tools for improved protein stability.

PubMed

Suplatov, Dmitry; Voevodin, Vladimir; Švedas, Vytas

2015-03-01

The ability of proteins and enzymes to maintain a functionally active conformation under adverse environmental conditions is an important feature of biocatalysts, vaccines, and biopharmaceutical proteins. From an evolutionary perspective, robust stability of proteins improves their biological fitness and allows for further optimization. Viewed from an industrial perspective, enzyme stability is crucial for the practical application of enzymes under the required reaction conditions. In this review, we analyze bioinformatic-driven strategies that are used to predict structural changes that can be applied to wild type proteins in order to produce more stable variants. The most commonly employed techniques can be classified into stochastic approaches, empirical or systematic rational design strategies, and design of chimeric proteins. We conclude that bioinformatic analysis can be efficiently used to study large protein superfamilies systematically as well as to predict particular structural changes which increase enzyme stability. Evolution has created a diversity of protein properties that are encoded in genomic sequences and structural data. Bioinformatics has the power to uncover this evolutionary code and provide a reproducible selection of hotspots - key residues to be mutated in order to produce more stable and functionally diverse proteins and enzymes. Further development of systematic bioinformatic procedures is needed to organize and analyze sequences and structures of proteins within large superfamilies and to link them to function, as well as to provide knowledge-based predictions for experimental evaluation. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The Role of Auxiliary Subunits for the Functional Diversity of Voltage-Gated Calcium Channels

PubMed Central

Campiglio, Marta; Flucher, Bernhard E

2015-01-01

Voltage-gated calcium channels (VGCCs) represent the sole mechanism to convert membrane depolarization into cellular functions like secretion, contraction, or gene regulation. VGCCs consist of a pore-forming α1 subunit and several auxiliary channel subunits. These subunits come in multiple isoforms and splice-variants giving rise to a stunning molecular diversity of possible subunit combinations. It is generally believed that specific auxiliary subunits differentially regulate the channels and thereby contribute to the great functional diversity of VGCCs. If auxiliary subunits can associate and dissociate from pre-existing channel complexes, this would allow dynamic regulation of channel properties. However, most auxiliary subunits modulate current properties very similarly, and proof that any cellular calcium channel function is indeed modulated by the physiological exchange of auxiliary subunits is still lacking. In this review we summarize available information supporting a differential modulation of calcium channel functions by exchange of auxiliary subunits, as well as experimental evidence in support of alternative functions of the auxiliary subunits. At the heart of the discussion is the concept that, in their native environment, VGCCs function in the context of macromolecular signaling complexes and that the auxiliary subunits help to orchestrate the diverse protein–protein interactions found in these calcium channel signalosomes. Thus, in addition to a putative differential modulation of current properties, differential subcellular targeting properties and differential protein–protein interactions of the auxiliary subunits may explain the need for their vast molecular diversity. J. Cell. Physiol. 999: 00–00, 2015. © 2015 The Authors. Journal of Cellular Physiology Published by Wiley Periodicals, Inc. J. Cell. Physiol. 230: 2019–2031, 2015. © 2015 Wiley Periodicals, Inc. PMID:25820299

Platyhelminth Venom Allergen-Like (VAL) proteins: revealing structural diversity, class-specific features and biological associations across the phylum

PubMed Central

CHALMERS, IAIN W.; HOFFMANN, KARL F.

2012-01-01

SUMMARY During platyhelminth infection, a cocktail of proteins is released by the parasite to aid invasion, initiate feeding, facilitate adaptation and mediate modulation of the host immune response. Included amongst these proteins is the Venom Allergen-Like (VAL) family, part of the larger sperm coating protein/Tpx-1/Ag5/PR-1/Sc7 (SCP/TAPS) superfamily. To explore the significance of this protein family during Platyhelminthes development and host interactions, we systematically summarize all published proteomic, genomic and immunological investigations of the VAL protein family to date. By conducting new genomic and transcriptomic interrogations to identify over 200 VAL proteins (228) from species in all 4 traditional taxonomic classes (Trematoda, Cestoda, Monogenea and Turbellaria), we further expand our knowledge related to platyhelminth VAL diversity across the phylum. Subsequent phylogenetic and tertiary structural analyses reveal several class-specific VAL features, which likely indicate a range of roles mediated by this protein family. Our comprehensive analysis of platyhelminth VALs represents a unifying synopsis for understanding diversity within this protein family and a firm context in which to initiate future functional characterization of these enigmatic members. PMID:22717097

Ankyrin-repeat containing proteins of microbes: a conserved structure with functional diversity

PubMed Central

Al-Khodor, Souhaila; Price, Christopher T.; Kalia, Awdhesh; Kwaik, Yousef Abu

2009-01-01

Summary The ankyrin repeat (ANK) is the most common protein-protein interaction motif in nature and predominantly found in eukaryotic proteins. The genome sequencing of various pathogenic or symbiotic bacteria and eukaryotic viruses identified numerous genes encoding ANK-containing proteins that were proposed to have been acquired from eukaryotes by horizontal gene transfer. However, the recent discovery of additional ANK-containing proteins encoded in the genomes of archaea and free-living bacteria suggests either a more ancient origin of the ANK motif or multiple convergent evolution events. Many bacterial pathogens employ various types of secretion systems to deliver ANK-containing proteins into eukaryotic cells where they mimic or manipulate various host functions. Understanding the molecular and biochemical functions of this family of proteins will enhance our understanding of important host-microbe interactions. PMID:19962898

Glutaredoxin in cancer development, progression, chemo-resistance and clinical applications

USDA-ARS?s Scientific Manuscript database

The Glutaredoxin (Grx) proteins, coupled with glutathione and glutathione reductase, constitute a major antioxidant system that counteracts the effects of oxidative stress in the cell. Grx proteins regulate diverse cellular functions and play an essential role in redox homeostasis. Abnormal regulati...

JNK Signaling: Regulation and Functions Based on Complex Protein-Protein Partnerships.

PubMed

Zeke, András; Misheva, Mariya; Reményi, Attila; Bogoyevitch, Marie A

2016-09-01

The c-Jun N-terminal kinases (JNKs), as members of the mitogen-activated protein kinase (MAPK) family, mediate eukaryotic cell responses to a wide range of abiotic and biotic stress insults. JNKs also regulate important physiological processes, including neuronal functions, immunological actions, and embryonic development, via their impact on gene expression, cytoskeletal protein dynamics, and cell death/survival pathways. Although the JNK pathway has been under study for >20 years, its complexity is still perplexing, with multiple protein partners of JNKs underlying the diversity of actions. Here we review the current knowledge of JNK structure and isoforms as well as the partnerships of JNKs with a range of intracellular proteins. Many of these proteins are direct substrates of the JNKs. We analyzed almost 100 of these target proteins in detail within a framework of their classification based on their regulation by JNKs. Examples of these JNK substrates include a diverse assortment of nuclear transcription factors (Jun, ATF2, Myc, Elk1), cytoplasmic proteins involved in cytoskeleton regulation (DCX, Tau, WDR62) or vesicular transport (JIP1, JIP3), cell membrane receptors (BMPR2), and mitochondrial proteins (Mcl1, Bim). In addition, because upstream signaling components impact JNK activity, we critically assessed the involvement of signaling scaffolds and the roles of feedback mechanisms in the JNK pathway. Despite a clarification of many regulatory events in JNK-dependent signaling during the past decade, many other structural and mechanistic insights are just beginning to be revealed. These advances open new opportunities to understand the role of JNK signaling in diverse physiological and pathophysiological states. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Trends in genome dynamics among major orders of insects revealed through variations in protein families.

PubMed

Rappoport, Nadav; Linial, Michal

2015-08-07

Insects belong to a class that accounts for the majority of animals on earth. With over one million identified species, insects display a huge diversity and occupy extreme environments. At present, there are dozens of fully sequenced insect genomes that cover a range of habitats, social behavior and morphologies. In view of such diverse collection of genomes, revealing evolutionary trends and charting functional relationships of proteins remain challenging. We analyzed the relatedness of 17 complete proteomes representative of proteomes from insects including louse, bee, beetle, ants, flies and mosquitoes, as well as an out-group from the crustaceans. The analyzed proteomes mostly represented the orders of Hymenoptera and Diptera. The 287,405 protein sequences from the 18 proteomes were automatically clustered into 20,933 families, including 799 singletons. A comprehensive analysis based on statistical considerations identified the families that were significantly expanded or reduced in any of the studied organisms. Among all the tested species, ants are characterized by an exceptionally high rate of family gain and loss. By assigning annotations to hundreds of species-specific families, the functional diversity among species and between the major clades (Diptera and Hymenoptera) is revealed. We found that many species-specific families are associated with receptor signaling, stress-related functions and proteases. The highest variability among insects associates with the function of transposition and nucleic acids processes (collectively coined TNAP). Specifically, the wasp and ants have an order of magnitude more TNAP families and proteins relative to species that belong to Diptera (mosquitoes and flies). An unsupervised clustering methodology combined with a comparative functional analysis unveiled proteomic signatures in the major clades of winged insects. We propose that the expansion of TNAP families in Hymenoptera potentially contributes to the accelerated genome dynamics that characterize the wasp and ants.

Protein tyrosine nitration in plants: Present knowledge, computational prediction and future perspectives.

PubMed

Kolbert, Zsuzsanna; Feigl, Gábor; Bordé, Ádám; Molnár, Árpád; Erdei, László

2017-04-01

Nitric oxide (NO) and related molecules (reactive nitrogen species) regulate diverse physiological processes mainly through posttranslational modifications such as protein tyrosine nitration (PTN). PTN is a covalent and specific modification of tyrosine (Tyr) residues resulting in altered protein structure and function. In the last decade, great efforts have been made to reveal candidate proteins, target Tyr residues and functional consequences of nitration in plants. This review intends to evaluate the accumulated knowledge about the biochemical mechanism, the structural and functional consequences and the selectivity of plants' protein nitration and also about the decomposition or conversion of nitrated proteins. At the same time, this review emphasizes yet unanswered or uncertain questions such as the reversibility/irreversibility of tyrosine nitration, the involvement of proteasomes in the removal of nitrated proteins or the effect of nitration on Tyr phosphorylation. The different NO producing systems of algae and higher plants raise the possibility of diversely regulated protein nitration. Therefore studying PTN from an evolutionary point of view would enrich our present understanding with novel aspects. Plant proteomic research can be promoted by the application of computational prediction tools such as GPS-YNO 2 and iNitro-Tyr software. Using the reference Arabidopsis proteome, Authors performed in silico analysis of tyrosine nitration in order to characterize plant tyrosine nitroproteome. Nevertheless, based on the common results of the present prediction and previous experiments the most likely nitrated proteins were selected thus recommending candidates for detailed future research. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

DISC1 Causes Associative Memory and Neurodevelopmental Defects in Fruit Flies

PubMed Central

Furukubo-Tokunaga, Katsuo; Kurita, Kazuki; Honjo, Ken; Pandey, Himani; Ando, Tetsuya; Takayama, Kojiro; Arai, Yuko; Mochizuki, Hiroaki; Ando, Mai; Kamiya, Atsushi; Sawa, Akira

2016-01-01

Originally found in a Scottish family with diverse mental disorders, the DISC1 protein has been characterized as an intracellular scaffold protein that associates with diverse binding partners in neural development. To explore its functions in a genetically tractable system, we expressed the human DISC1 in fruit flies (Drosophila melanogaster). As in mammalian neurons, DISC1 is localized to diverse subcellular domains of developing fly neurons including the nuclei, axons and dendrites. Overexpression of DISC1 impairs associative memory. Experiments with deletion/mutation constructs have revealed the importance of amino terminal domain (46–290) for memory suppression whereas carboxyl domain (598–854) and the amino terminal residues (1–45) including the nuclear localization signal (NLS1) are dispensable. DISC1 overexpression also causes suppression of axonal and dendritic branching of mushroom body neurons, which mediate a variety of cognitive functions in the fly brain. Analyses with deletion constructs reveal that protein domains 598–854 and 349–402 are both required for the suppression of axonal branching while amino-terminal domains including NLS1 are dispensable. In contrast, NLS1 was required for the suppression of dendritic branching, suggesting a mechanism involving gene expression. Moreover, domain 403–596 is also required for the suppression of dendritic branching. We also show that overexpression of DISC1 suppresses glutamatergic synaptogenesis in developing neuromuscular junctions. Deletion/mutation experiments have revealed the importance of protein domains 403–596 and 349–402 for synaptic suppression, while amino terminal domains including NLS1 are dispensable. Finally, we show that DISC1 functionally interacts with the fly homolog of Dysbindin (DTNBP1) via direct protein-protein interaction in developing synapses. PMID:26976042

The Individual and Population Genetics of Antibody Immunity.

PubMed

Watson, Corey T; Glanville, Jacob; Marasco, Wayne A

2017-07-01

Antibodies (Abs) produced by immunoglobulin (IG) genes are the most diverse proteins expressed in humans. While part of this diversity is generated by recombination during B-cell development and mutations during affinity maturation, the germ-line IG loci are also diverse across human populations and ethnicities. Recently, proof-of-concept studies have demonstrated genotype-phenotype correlations between specific IG germ-line variants and the quality of Ab responses during vaccination and disease. However, the functional consequences of IG genetic variation in Ab function and immunological outcomes remain underexplored. In this opinion article, we outline interconnections between IG genomic diversity and Ab-expressed repertoires and structure. We further propose a strategy for integrating IG genotyping with functional Ab profiling data as a means to better predict and optimize humoral responses in genetically diverse human populations, with immediate implications for personalized medicine. Copyright © 2017 Elsevier Ltd. All rights reserved.

Rhabdovirus accessory genes.

PubMed

Walker, Peter J; Dietzgen, Ralf G; Joubert, D Albert; Blasdell, Kim R

2011-12-01

The Rhabdoviridae is one of the most ecologically diverse families of RNA viruses with members infecting a wide range of organisms including placental mammals, marsupials, birds, reptiles, fish, insects and plants. The availability of complete nucleotide sequences for an increasing number of rhabdoviruses has revealed that their ecological diversity is reflected in the diversity and complexity of their genomes. The five canonical rhabdovirus structural protein genes (N, P, M, G and L) that are shared by all rhabdoviruses are overprinted, overlapped and interspersed with a multitude of novel and diverse accessory genes. Although not essential for replication in cell culture, several of these genes have been shown to have roles associated with pathogenesis and apoptosis in animals, and cell-to-cell movement in plants. Others appear to be secreted or have the characteristics of membrane-anchored glycoproteins or viroporins. However, most encode proteins of unknown function that are unrelated to any other known proteins. Understanding the roles of these accessory genes and the strategies by which rhabdoviruses use them to engage, divert and re-direct cellular processes will not only present opportunities to develop new anti-viral therapies but may also reveal aspects of cellar function that have broader significance in biology, agriculture and medicine. Crown Copyright © 2011. Published by Elsevier B.V. All rights reserved.

Direct Correlation between Motile Behavior and Protein Abundance in Single Cells

PubMed Central

Gillet, Sébastien; Frankel, Nicholas W.; Weibel, Douglas B.

2016-01-01

Understanding how stochastic molecular fluctuations affect cell behavior requires the quantification of both behavior and protein numbers in the same cells. Here, we combine automated microscopy with in situ hydrogel polymerization to measure single-cell protein expression after tracking swimming behavior. We characterized the distribution of non-genetic phenotypic diversity in Escherichia coli motility, which affects single-cell exploration. By expressing fluorescently tagged chemotaxis proteins (CheR and CheB) at different levels, we quantitatively mapped motile phenotype (tumble bias) to protein numbers using thousands of single-cell measurements. Our results disagreed with established models until we incorporated the role of CheB in receptor deamidation and the slow fluctuations in receptor methylation. Beyond refining models, our central finding is that changes in numbers of CheR and CheB affect the population mean tumble bias and its variance independently. Therefore, it is possible to adjust the degree of phenotypic diversity of a population by adjusting the global level of expression of CheR and CheB while keeping their ratio constant, which, as shown in previous studies, confers functional robustness to the system. Since genetic control of protein expression is heritable, our results suggest that non-genetic diversity in motile behavior is selectable, supporting earlier hypotheses that such diversity confers a selective advantage. PMID:27599206

DNA mimic proteins: functions, structures, and bioinformatic analysis.

PubMed

Wang, Hao-Ching; Ho, Chun-Han; Hsu, Kai-Cheng; Yang, Jinn-Moon; Wang, Andrew H-J

2014-05-13

DNA mimic proteins have DNA-like negative surface charge distributions, and they function by occupying the DNA binding sites of DNA binding proteins to prevent these sites from being accessed by DNA. DNA mimic proteins control the activities of a variety of DNA binding proteins and are involved in a wide range of cellular mechanisms such as chromatin assembly, DNA repair, transcription regulation, and gene recombination. However, the sequences and structures of DNA mimic proteins are diverse, making them difficult to predict by bioinformatic search. To date, only a few DNA mimic proteins have been reported. These DNA mimics were not found by searching for functional motifs in their sequences but were revealed only by structural analysis of their charge distribution. This review highlights the biological roles and structures of 16 reported DNA mimic proteins. We also discuss approaches that might be used to discover new DNA mimic proteins.

BAR domain proteins regulate Rho GTPase signaling.

PubMed

Aspenström, Pontus

2014-01-01

BAR proteins comprise a heterogeneous group of multi-domain proteins with diverse biological functions. The common denominator is the Bin-Amphiphysin-Rvs (BAR) domain that not only confers targeting to lipid bilayers, but also provides scaffolding to mold lipid membranes into concave or convex surfaces. This function of BAR proteins is an important determinant in the dynamic reconstruction of membrane vesicles, as well as of the plasma membrane. Several BAR proteins function as linkers between cytoskeletal regulation and membrane dynamics. These links are provided by direct interactions between BAR proteins and actin-nucleation-promoting factors of the Wiskott-Aldrich syndrome protein family and the Diaphanous-related formins. The Rho GTPases are key factors for orchestration of this intricate interplay. This review describes how BAR proteins regulate the activity of Rho GTPases, as well as how Rho GTPases regulate the function of BAR proteins. This mutual collaboration is a central factor in the regulation of vital cellular processes, such as cell migration, cytokinesis, intracellular transport, endocytosis, and exocytosis.

Proteopedia: 3D Visualization and Annotation of Transcription Factor-DNA Readout Modes

ERIC Educational Resources Information Center

Dantas Machado, Ana Carolina; Saleebyan, Skyler B.; Holmes, Bailey T.; Karelina, Maria; Tam, Julia; Kim, Sharon Y.; Kim, Keziah H.; Dror, Iris; Hodis, Eran; Martz, Eric; Compeau, Patricia A.; Rohs, Remo

2012-01-01

3D visualization assists in identifying diverse mechanisms of protein-DNA recognition that can be observed for transcription factors and other DNA binding proteins. We used Proteopedia to illustrate transcription factor-DNA readout modes with a focus on DNA shape, which can be a function of either nucleotide sequence (Hox proteins) or base pairing…

New insights into the targeting of a sub-set of tail-anchored proteins to the outer mitochondrial membrane

USDA-ARS?s Scientific Manuscript database

Tail-anchored (TA) proteins are a unique class of functionally diverse membrane proteins that are defined by their single C-terminal membrane-spanning domain and their ability to insert post-translationally into specific organelles with an Nout-Cin orientation. The molecular mechanisms by which TA p...

Regulator of G-protein signalling and GoLoco proteins suppress TRPC4 channel function via acting at Gαi/o.

PubMed

Jeon, Jae-Pyo; Thakur, Dhananjay P; Tian, Jin-Bin; So, Insuk; Zhu, Michael X

2016-05-15

Transient receptor potential canonical 4 (TRPC4) forms non-selective cation channels implicated in the regulation of diverse physiological functions. Previously, TRPC4 was shown to be activated by the Gi/o subgroup of heterotrimeric G-proteins involving Gαi/o, rather than Gβγ, subunits. Because the lifetime and availability of Gα-GTP are regulated by regulators of G-protein signalling (RGS) and Gαi/o-Loco (GoLoco) domain-containing proteins via their GTPase-activating protein (GAP) and guanine-nucleotide-dissociation inhibitor (GDI) functions respectively, we tested how RGS and GoLoco domain proteins affect TRPC4 currents activated via Gi/o-coupled receptors. Using whole-cell patch-clamp recordings, we show that both RGS and GoLoco proteins [RGS4, RGS6, RGS12, RGS14, LGN or activator of G-protein signalling 3 (AGS3)] suppress receptor-mediated TRPC4 activation without causing detectable basal current or altering surface expression of the channel protein. The inhibitory effects are dependent on the GAP and GoLoco domains and facilitated by enhancing membrane targeting of the GoLoco protein AGS3. In addition, RGS, but not GoLoco, proteins accelerate desensitization of receptor-activation evoked TRPC4 currents. The inhibitory effects of RGS and GoLoco domains are additive and are most prominent with RGS12 and RGS14, which contain both RGS and GoLoco domains. Our data support the notion that the Gα, but not Gβγ, arm of the Gi/o signalling is involved in TRPC4 activation and unveil new roles for RGS and GoLoco domain proteins in fine-tuning TRPC4 activities. The versatile and diverse functions of RGS and GoLoco proteins in regulating G-protein signalling may underlie the complexity of receptor-operated TRPC4 activation in various cell types under different conditions. © 2016 The Author(s). published by Portland Press Limited on behalf of the Biochemical Society.

The Recombinant Sea Urchin Immune Effector Protein, rSpTransformer-E1, Binds to Phosphatidic Acid and Deforms Membranes

PubMed Central

Lun, Cheng Man; Samuel, Robin L.; Gillmor, Susan D.; Boyd, Anthony; Smith, L. Courtney

2017-01-01

The purple sea urchin, Strongylocentrotus purpuratus, possesses a sophisticated innate immune system that functions without adaptive capabilities and responds to pathogens effectively by expressing the highly diverse SpTransformer gene family (formerly the Sp185/333 gene family). The swift gene expression response and the sequence diversity of SpTransformer cDNAs suggest that the encoded proteins have immune functions. Individual sea urchins can express up to 260 distinct SpTransformer proteins, and their diversity suggests that different versions may have different functions. Although the deduced proteins are diverse, they share an overall structure of a hydrophobic leader, a glycine-rich N-terminal region, a histidine-rich region, and a C-terminal region. Circular dichroism analysis of a recombinant SpTransformer protein, rSpTransformer-E1 (rSpTrf-E1) demonstrates that it is intrinsically disordered and transforms to α helical in the presence of buffer additives and binding targets. Although native SpTrf proteins are associated with the membranes of perinuclear vesicles in the phagocyte class of coelomocytes and are present on the surface of small phagocytes, they have no predicted transmembrane region or conserved site for glycophosphatidylinositol linkage. To determine whether native SpTrf proteins associate with phagocyte membranes through interactions with lipids, when rSpTrf-E1 is incubated with lipid-embedded nylon strips, it binds to phosphatidic acid (PA) through both the glycine-rich region and the histidine-rich region. Synthetic liposomes composed of PA and phosphatidylcholine show binding between rSpTrf-E1 and PA by fluorescence resonance energy transfer, which is associated with leakage of luminal contents suggesting changes in lipid organization and perhaps liposome lysis. Interactions with liposomes also change membrane curvature leading to liposome budding, fusion, and invagination, which is associated with PA clustering induced by rSpTrf-E1 binding. Longer incubations result in the extraction of PA from the liposomes, which form disorganized clusters. CD shows that when rSpTrf-E1 binds to PA, it changes its secondary structure from disordered to α helical. These results provide evidence for how SpTransformer proteins may associate with molecules that have exposed phosphates including PA on cell membranes and how the characteristic of protein multimerization may drive changes in the organization of membrane lipids. PMID:28553283

Enzyme Immobilization: Nanobiotechnology: Putting Molecular Machines to Work

DOE Office of Scientific and Technical Information (OSTI.GOV)

None

2009-04-01

Describes, in general terms, the concepts of high-throughput protein expression coupled with immobilizations in functionalized nanoporous materials to carry out multiple kinds of diverse reactions. The animations also illustrate that immobilized enzymes potentially can refold inactive proteins. Transcripts of videos available upon request

Genome-wide analysis of the Dof transcription factor gene family reveals soybean-specific duplicable and functional characteristics.

PubMed

Guo, Yong; Qiu, Li-Juan

2013-01-01

The Dof domain protein family is a classic plant-specific zinc-finger transcription factor family involved in a variety of biological processes. There is great diversity in the number of Dof genes in different plants. However, there are only very limited reports on the characterization of Dof transcription factors in soybean (Glycine max). In the present study, 78 putative Dof genes were identified from the whole-genome sequence of soybean. The predicted GmDof genes were non-randomly distributed within and across 19 out of 20 chromosomes and 97.4% (38 pairs) were preferentially retained duplicate paralogous genes located in duplicated regions of the genome. Soybean-specific segmental duplications contributed significantly to the expansion of the soybean Dof gene family. These Dof proteins were phylogenetically clustered into nine distinct subgroups among which the gene structure and motif compositions were considerably conserved. Comparative phylogenetic analysis of these Dof proteins revealed four major groups, similar to those reported for Arabidopsis and rice. Most of the GmDofs showed specific expression patterns based on RNA-seq data analyses. The expression patterns of some duplicate genes were partially redundant while others showed functional diversity, suggesting the occurrence of sub-functionalization during subsequent evolution. Comprehensive expression profile analysis also provided insights into the soybean-specific functional divergence among members of the Dof gene family. Cis-regulatory element analysis of these GmDof genes suggested diverse functions associated with different processes. Taken together, our results provide useful information for the functional characterization of soybean Dof genes by combining phylogenetic analysis with global gene-expression profiling.

The logic of DNA replication in double-stranded DNA viruses: insights from global analysis of viral genomes.

PubMed

Kazlauskas, Darius; Krupovic, Mart; Venclovas, Česlovas

2016-06-02

Genomic DNA replication is a complex process that involves multiple proteins. Cellular DNA replication systems are broadly classified into only two types, bacterial and archaeo-eukaryotic. In contrast, double-stranded (ds) DNA viruses feature a much broader diversity of DNA replication machineries. Viruses differ greatly in both completeness and composition of their sets of DNA replication proteins. In this study, we explored whether there are common patterns underlying this extreme diversity. We identified and analyzed all major functional groups of DNA replication proteins in all available proteomes of dsDNA viruses. Our results show that some proteins are common to viruses infecting all domains of life and likely represent components of the ancestral core set. These include B-family polymerases, SF3 helicases, archaeo-eukaryotic primases, clamps and clamp loaders of the archaeo-eukaryotic type, RNase H and ATP-dependent DNA ligases. We also discovered a clear correlation between genome size and self-sufficiency of viral DNA replication, the unanticipated dominance of replicative helicases and pervasive functional associations among certain groups of DNA replication proteins. Altogether, our results provide a comprehensive view on the diversity and evolution of replication systems in the DNA virome and uncover fundamental principles underlying the orchestration of viral DNA replication. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

To Be Specific or Not: The Critical Relationship Between Hox And TALE Proteins.

PubMed

Merabet, Samir; Mann, Richard S

2016-06-01

Hox proteins are key regulatory transcription factors that act in different tissues of the embryo to provide specific spatial and temporal coordinates to each cell. These patterning functions often depend on the presence of the TALE-homeodomain class cofactors, which form cooperative DNA-binding complexes with all Hox proteins. How this family of cofactors contributes to the highly diverse and specific functions of Hox proteins in vivo remains an important unsolved question. We review here the most recent advances in understanding the molecular mechanisms underlying Hox-TALE function. In particular, we discuss the role of DNA shape, DNA-binding affinity, and protein-protein interaction flexibility in dictating Hox-TALE specificity. We propose several models to explain how these mechanisms are integrated with each other in the context of the many distinct functions that Hox and TALE factors carry out in vivo. Copyright © 2016 Elsevier Ltd. All rights reserved.

Integrative Identification of Arabidopsis Mitochondrial Proteome and Its Function Exploitation through Protein Interaction Network

PubMed Central

Cui, Jian; Liu, Jinghua; Li, Yuhua; Shi, Tieliu

2011-01-01

Mitochondria are major players on the production of energy, and host several key reactions involved in basic metabolism and biosynthesis of essential molecules. Currently, the majority of nucleus-encoded mitochondrial proteins are unknown even for model plant Arabidopsis. We reported a computational framework for predicting Arabidopsis mitochondrial proteins based on a probabilistic model, called Naive Bayesian Network, which integrates disparate genomic data generated from eight bioinformatics tools, multiple orthologous mappings, protein domain properties and co-expression patterns using 1,027 microarray profiles. Through this approach, we predicted 2,311 candidate mitochondrial proteins with 84.67% accuracy and 2.53% FPR performances. Together with those experimental confirmed proteins, 2,585 mitochondria proteins (named CoreMitoP) were identified, we explored those proteins with unknown functions based on protein-protein interaction network (PIN) and annotated novel functions for 26.65% CoreMitoP proteins. Moreover, we found newly predicted mitochondrial proteins embedded in particular subnetworks of the PIN, mainly functioning in response to diverse environmental stresses, like salt, draught, cold, and wound etc. Candidate mitochondrial proteins involved in those physiological acitivites provide useful targets for further investigation. Assigned functions also provide comprehensive information for Arabidopsis mitochondrial proteome. PMID:21297957

CML24, Regulated in Expression by Diverse Stimuli, Encodes a Potential Ca2+ Sensor That Functions in Responses to Abscisic Acid, Daylength, and Ion Stress1

PubMed Central

Delk, Nikkí A.; Johnson, Keith A.; Chowdhury, Naweed I.; Braam, Janet

2005-01-01

Changes in intracellular calcium (Ca2+) levels serve to signal responses to diverse stimuli. Ca2+ signals are likely perceived through proteins that bind Ca2+, undergo conformation changes following Ca2+ binding, and interact with target proteins. The 50-member calmodulin-like (CML) Arabidopsis (Arabidopsis thaliana) family encodes proteins containing the predicted Ca2+-binding EF-hand motif. The functions of virtually all these proteins are unknown. CML24, also known as TCH2, shares over 40% amino acid sequence identity with calmodulin, has four EF hands, and undergoes Ca2+-dependent changes in hydrophobic interaction chromatography and migration rate through denaturing gel electrophoresis, indicating that CML24 binds Ca2+ and, as a consequence, undergoes conformational changes. CML24 expression occurs in all major organs, and transcript levels are increased from 2- to 15-fold in plants subjected to touch, darkness, heat, cold, hydrogen peroxide, abscisic acid (ABA), and indole-3-acetic acid. However, CML24 protein accumulation changes were not detectable. The putative CML24 regulatory region confers reporter expression at sites of predicted mechanical stress; in regions undergoing growth; in vascular tissues and various floral organs; and in stomata, trichomes, and hydathodes. CML24-underexpressing transgenics are resistant to ABA inhibition of germination and seedling growth, are defective in long-day induction of flowering, and have enhanced tolerance to CoCl2, molybdic acid, ZnSO4, and MgCl2. MgCl2 tolerance is not due to reduced uptake or to elevated Ca2+ accumulation. Together, these data present evidence that CML24, a gene expressed in diverse organs and responsive to diverse stimuli, encodes a potential Ca2+ sensor that may function to enable responses to ABA, daylength, and presence of various salts. PMID:16113225

Phyllosphere Metaproteomes of Trees from the Brazilian Atlantic Forest Show High Levels of Functional Redundancy.

PubMed

Lambais, M R; Barrera, S E; Santos, E C; Crowley, D E; Jumpponen, A

2017-01-01

The phyllosphere of the Brazilian Atlantic Forest has been estimated to contain several million bacterial species that are associated with approximately 20000 plant species. Despite the high bacterial diversity in the phyllosphere, the function of these microorganisms and the mechanisms driving their community assembly are largely unknown. In this study, we characterized the bacterial communities in the phyllospheres of four tree species of the Atlantic Forest (Mollinedia schottiana, Ocotea dispersa, Ocotea teleiandra, and Tabebuia serratifolia) and their metaproteomes to examine the basic protein functional groups expressed in the phyllosphere. Bacterial community analyses using 16S rRNA gene sequencing confirmed prior observations that plant species harbor distinct bacterial communities and that plants of the same taxon have more similar communities than more distantly related taxa. Using LC-ESI-Q-TOF, we identified 216 nonredundant proteins, based on 3503 peptide mass spectra. Most protein families were shared among the phyllosphere communities, suggesting functional redundancy despite differences in the species compositions of the bacterial communities. Proteins involved in glycolysis and anaerobic carbohydrate metabolism, solute transport, protein metabolism, cell motility, stress and antioxidant responses, nitrogen metabolism, and iron homeostasis were among the most frequently detected. In contrast to prior studies on crop plants and Arabidopsis, a low abundance of OTUs related to Methylobacterium and no proteins associated with the metabolism of one-carbon molecules were detected in the phyllospheres of the tree species studied here. Our data suggest that even though the phyllosphere bacterial communities of different tree species are phylogenetically diverse, their metaproteomes are functionally convergent with respect to traits required for survival on leaf surfaces.

Evolution of an intricate J-protein network driving protein disaggregation in eukaryotes.

PubMed

Nillegoda, Nadinath B; Stank, Antonia; Malinverni, Duccio; Alberts, Niels; Szlachcic, Anna; Barducci, Alessandro; De Los Rios, Paolo; Wade, Rebecca C; Bukau, Bernd

2017-05-15

Hsp70 participates in a broad spectrum of protein folding processes extending from nascent chain folding to protein disaggregation. This versatility in function is achieved through a diverse family of J-protein cochaperones that select substrates for Hsp70. Substrate selection is further tuned by transient complexation between different classes of J-proteins, which expands the range of protein aggregates targeted by metazoan Hsp70 for disaggregation. We assessed the prevalence and evolutionary conservation of J-protein complexation and cooperation in disaggregation. We find the emergence of a eukaryote-specific signature for interclass complexation of canonical J-proteins. Consistently, complexes exist in yeast and human cells, but not in bacteria, and correlate with cooperative action in disaggregation in vitro. Signature alterations exclude some J-proteins from networking, which ensures correct J-protein pairing, functional network integrity and J-protein specialization. This fundamental change in J-protein biology during the prokaryote-to-eukaryote transition allows for increased fine-tuning and broadening of Hsp70 function in eukaryotes.

sc-PDB: a database for identifying variations and multiplicity of 'druggable' binding sites in proteins.

PubMed

Meslamani, Jamel; Rognan, Didier; Kellenberger, Esther

2011-05-01

The sc-PDB database is an annotated archive of druggable binding sites extracted from the Protein Data Bank. It contains all-atoms coordinates for 8166 protein-ligand complexes, chosen for their geometrical and physico-chemical properties. The sc-PDB provides a functional annotation for proteins, a chemical description for ligands and the detailed intermolecular interactions for complexes. The sc-PDB now includes a hierarchical classification of all the binding sites within a functional class. The sc-PDB entries were first clustered according to the protein name indifferent of the species. For each cluster, we identified dissimilar sites (e.g. catalytic and allosteric sites of an enzyme). SCOPE AND APPLICATIONS: The classification of sc-PDB targets by binding site diversity was intended to facilitate chemogenomics approaches to drug design. In ligand-based approaches, it avoids comparing ligands that do not share the same binding site. In structure-based approaches, it permits to quantitatively evaluate the diversity of the binding site definition (variations in size, sequence and/or structure). The sc-PDB database is freely available at: http://bioinfo-pharma.u-strasbg.fr/scPDB.

The membrane fusion enigma: SNAREs, Sec1/Munc18 proteins, and their accomplices--guilty as charged?

PubMed

Rizo, Josep; Südhof, Thomas C

2012-01-01

Neurotransmitter release is governed by proteins that have homo-logs in most types of intracellular membrane fusion, including the Sec1/Munc18 protein Munc18-1 and the SNARE proteins syntaxin-1, synaptobrevin/VAMP, and SNAP-25. The SNAREs initiate fusion by forming tight SNARE complexes that bring the vesicle and plasma membranes together. SNARE maintenance in a functional state depends on two chaperone systems (Hsc70/αCSP/SGT and synuclein); defects in these systems lead to neurodegeneration. Munc18-1 binds to an autoinhibitory closed conformation of syntaxin-1, gating formation of SNARE complexes, and also binds to SNARE complexes, which likely underlies the crucial function of Munc18-1 in membrane fusion by an as-yet unclear mechanism. Syntaxin-1 opening is mediated by Munc13s through their MUN domain, which is homologous to diverse tethering factors and may also have a general role in fusion. MUN domain activity is likely modulated in diverse presynaptic plasticity processes that depend on Ca(2+) and RIM proteins, among others.

Intermediate Filaments and the Regulation of Cell Motility during Regeneration and Wound Healing.

PubMed

Cheng, Fang; Eriksson, John E

2017-09-01

SUMMARYIntermediate filaments (IFs) comprise a diverse group of flexible cytoskeletal structures, the assembly, dynamics, and functions of which are regulated by posttranslational modifications. Characteristically, the expression of IF proteins is specific for tissues, differentiation stages, cell types, and functional contexts. Recent research has rapidly expanded the knowledge of IF protein functions. From being regarded as primarily structural proteins, it is now well established that IFs act as powerful modulators of cell motility and migration, playing crucial roles in wound healing and tissue regeneration, as well as inflammatory and immune responses. Although many of these IF-associated functions are essential for tissue repair, the involvement of IF proteins has been established in many additional facets of tissue healing and regeneration. Here, we review the recent progress in understanding the multiple functions of cytoplasmic IFs that relate to cell motility in the context of wound healing, taking examples from studies on keratin, vimentin, and nestin. Wound healing and regeneration include orchestration of a broad range of cellular processes, including regulation of cell attachment and migration, proliferation, differentiation, immune responses, angiogenesis, and remodeling of the extracellular matrix. In this respect, IF proteins now emerge as multifactorial and tissue-specific integrators of tissue regeneration, thereby acting as essential guardian biopolymers at the interface between health and disease, the failing of which contributes to a diverse range of pathologies. Copyright © 2017 Cold Spring Harbor Laboratory Press; all rights reserved.

A frequency-based linguistic approach to protein decoding and design: Simple concepts, diverse applications, and the SCS Package

PubMed Central

Motomura, Kenta; Nakamura, Morikazu; Otaki, Joji M.

2013-01-01

Protein structure and function information is coded in amino acid sequences. However, the relationship between primary sequences and three-dimensional structures and functions remains enigmatic. Our approach to this fundamental biochemistry problem is based on the frequencies of short constituent sequences (SCSs) or words. A protein amino acid sequence is considered analogous to an English sentence, where SCSs are equivalent to words. Availability scores, which are defined as real SCS frequencies in the non-redundant amino acid database relative to their probabilistically expected frequencies, demonstrate the biological usage bias of SCSs. As a result, this frequency-based linguistic approach is expected to have diverse applications, such as secondary structure specifications by structure-specific SCSs and immunological adjuvants with rare or non-existent SCSs. Linguistic similarities (e.g., wide ranges of scale-free distributions) and dissimilarities (e.g., behaviors of low-rank samples) between proteins and the natural English language have been revealed in the rank-frequency relationships of SCSs or words. We have developed a web server, the SCS Package, which contains five applications for analyzing protein sequences based on the linguistic concept. These tools have the potential to assist researchers in deciphering structurally and functionally important protein sites, species-specific sequences, and functional relationships between SCSs. The SCS Package also provides researchers with a tool to construct amino acid sequences de novo based on the idiomatic usage of SCSs. PMID:24688703

A frequency-based linguistic approach to protein decoding and design: Simple concepts, diverse applications, and the SCS Package.

PubMed

Motomura, Kenta; Nakamura, Morikazu; Otaki, Joji M

2013-01-01

Protein structure and function information is coded in amino acid sequences. However, the relationship between primary sequences and three-dimensional structures and functions remains enigmatic. Our approach to this fundamental biochemistry problem is based on the frequencies of short constituent sequences (SCSs) or words. A protein amino acid sequence is considered analogous to an English sentence, where SCSs are equivalent to words. Availability scores, which are defined as real SCS frequencies in the non-redundant amino acid database relative to their probabilistically expected frequencies, demonstrate the biological usage bias of SCSs. As a result, this frequency-based linguistic approach is expected to have diverse applications, such as secondary structure specifications by structure-specific SCSs and immunological adjuvants with rare or non-existent SCSs. Linguistic similarities (e.g., wide ranges of scale-free distributions) and dissimilarities (e.g., behaviors of low-rank samples) between proteins and the natural English language have been revealed in the rank-frequency relationships of SCSs or words. We have developed a web server, the SCS Package, which contains five applications for analyzing protein sequences based on the linguistic concept. These tools have the potential to assist researchers in deciphering structurally and functionally important protein sites, species-specific sequences, and functional relationships between SCSs. The SCS Package also provides researchers with a tool to construct amino acid sequences de novo based on the idiomatic usage of SCSs.

Diversity in structure and function of tethering complexes: evidence for different mechanisms in vesicular transport regulation.

PubMed

Kümmel, D; Heinemann, U

2008-04-01

The term 'tethering factor' has been coined for a heterogeneous group of proteins that all are required for protein trafficking prior to vesicle docking and SNARE-mediated membrane fusion. Two groups of tethering factors can be distinguished, long coiled-coil proteins and multi-subunit complexes. To date, eight such protein complexes have been identified in yeast, and they are required for different trafficking steps. Homologous complexes are found in all eukaryotic organisms, but conservation seems to be less strict than for other components of the trafficking machinery. In fact, for most proposed multi-subunit tethers their ability to actually bridge two membranes remains to be shown. Here we discuss recent progress in the structural and functional characterization of tethering complexes and present the emerging view that the different complexes are quite diverse in their structure and the molecular mechanisms underlying their function. TRAPP and the exocyst are the structurally best characterized tethering complexes. Their comparison fails to reveal any similarity on a struc nottural level. Furthermore, the interactions with regulatory Rab GTPases vary, with TRAPP acting as a nucleotide exchange factor and the exocyst being an effector. Considering these differences among the tethering complexes as well as between their yeast and mammalian orthologs which is apparent from recent studies, we suggest that tethering complexes do not mediate a strictly conserved process in vesicular transport but are diverse regulators acting after vesicle budding and prior to membrane fusion.

Mitochondrial Biogenesis in Diverse Cauliflower Cultivars under Mild and Severe Drought. Impaired Coordination of Selected Transcript and Proteomic Responses, and Regulation of Various Multifunctional Proteins

PubMed Central

Rurek, Michał; Czołpińska, Magdalena; Staszak, Aleksandra Maria; Nowak, Witold; Krzesiński, Włodzimierz; Spiżewski, Tomasz

2018-01-01

Mitochondrial responses under drought within Brassica genus are poorly understood. The main goal of this study was to investigate mitochondrial biogenesis of three cauliflower (Brassica oleracea var. botrytis) cultivars with varying drought tolerance. Diverse quantitative changes (decreases in abundance mostly) in the mitochondrial proteome were assessed by two-dimensional gel electrophoresis (2D PAGE) coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS). Respiratory (e.g., complex II, IV (CII, CIV) and ATP synthase subunits), transporter (including diverse porin isoforms) and matrix multifunctional proteins (e.g., components of RNA editing machinery) were diversely affected in their abundance under two drought levels. Western immunoassays showed additional cultivar-specific responses of selected mitochondrial proteins. Dehydrin-related tryptic peptides (found in several 2D spots) immunopositive with dehydrin-specific antisera highlighted the relevance of mitochondrial dehydrin-like proteins for the drought response. The abundance of selected mRNAs participating in drought response was also determined. We conclude that mitochondrial biogenesis was strongly, but diversely affected in various cauliflower cultivars, and associated with drought tolerance at the proteomic and functional levels. However, discussed alternative oxidase (AOX) regulation at the RNA and protein level were largely uncoordinated due to the altered availability of transcripts for translation, mRNA/ribosome interactions, and/or miRNA impact on transcript abundance and translation. PMID:29642585

Quantitative Proteomics Reveals the Regulatory Networks of Circular RNA CDR1as in Hepatocellular Carcinoma Cells.

PubMed

Yang, Xue; Xiong, Qian; Wu, Ying; Li, Siting; Ge, Feng

2017-10-06

Circular RNAs (circRNAs), a class of widespread endogenous RNAs, play crucial roles in diverse biological processes and are potential biomarkers in diverse human diseases and cancers. Cerebellar-degeneration-related protein 1 antisense RNA (CDR1as), an oncogenic circRNA, is involved in human tumorigenesis and is dysregulated in hepatocellular carcinoma (HCC). However, the molecular mechanisms underlying CDR1as functions in HCC remain unclear. Here we explored the functions of CDR1as and searched for CDR1as-regulated proteins in HCC cells. A quantitative proteomics strategy was employed to globally identify CDR1as-regulated proteins in HCC cells. In total, we identified 330 differentially expressed proteins (DEPs) upon enhanced CDR1as expression in HepG2 cells, indicating that they could be proteins regulated by CDR1as. Bioinformatic analysis revealed that many DEPs were involved in cell proliferation and the cell cycle. Further functional studies of epidermal growth factor receptor (EGFR) found that CDR1as exerts its effects on cell proliferation at least in part through the regulation of EGFR expression. We further confirmed that CDR1as could inhibit the expression of microRNA-7 (miR-7). EGFR is a validated target of miR-7; therefore, CDR1as may exert its function by regulating EGFR expression via targeting miR-7 in HCC cells. Taken together, we revealed novel functions and underlying mechanisms of CDR1as in HCC cells. This study serves as the first proteome-wide analysis of a circRNA-regulated protein in cells and provides a reliable and highly efficient method for globally identifying circRNA-regulated proteins.

Methods for the Detection, Study, and Dynamic Profiling of O-GlcNAc Glycosylation.

PubMed

Thompson, John W; Griffin, Matthew E; Hsieh-Wilson, Linda C

2018-01-01

The addition of O-linked β-N-acetylglucosamine (O-GlcNAc) to serine/threonine residues of proteins is a ubiquitous posttranslational modification found in all multicellular organisms. Like phosphorylation, O-GlcNAc glycosylation (O-GlcNAcylation) is inducible and regulates a myriad of physiological and pathological processes. However, understanding the diverse functions of O-GlcNAcylation is often challenging due to the difficulty of detecting and quantifying the modification. Thus, robust methods to study O-GlcNAcylation are essential to elucidate its key roles in the regulation of individual proteins, complex cellular processes, and disease. In this chapter, we describe a set of chemoenzymatic labeling methods to (1) detect O-GlcNAcylation on proteins of interest, (2) monitor changes in both the total levels of O-GlcNAcylation and its stoichiometry on proteins of interest, and (3) enable mapping of O-GlcNAc to specific serine/threonine residues within proteins to facilitate functional studies. First, we outline a procedure for the expression and purification of a multiuse mutant galactosyltransferase enzyme (Y289L GalT). We then describe the use of Y289L GalT to modify O-GlcNAc residues with a functional handle, N-azidoacetylgalactosamine (GalNAz). Finally, we discuss several applications of the copper-catalyzed azide-alkyne cycloaddition "click" reaction to attach various alkyne-containing chemical probes to GalNAz and demonstrate how this functionalization of O-GlcNAc-modified proteins can be used to realize (1)-(3) above. Overall, these methods, which utilize commercially available reagents and standard protein analytical tools, will serve to advance our understanding of the diverse and important functions of O-GlcNAcylation. © 2018 Elsevier Inc. All rights reserved.

Identification of widespread adenosine nucleotide binding in Mycobacterium tuberculosis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ansong, Charles; Ortega, Corrie; Payne, Samuel H.

The annotation of protein function is almost completely performed by in silico approaches. However, computational prediction of protein function is frequently incomplete and error prone. In Mycobacterium tuberculosis (Mtb), ~25% of all genes have no predicted function and are annotated as hypothetical proteins. This lack of functional information severely limits our understanding of Mtb pathogenicity. Current tools for experimental functional annotation are limited and often do not scale to entire protein families. Here, we report a generally applicable chemical biology platform to functionally annotate bacterial proteins by combining activity-based protein profiling (ABPP) and quantitative LC-MS-based proteomics. As an example ofmore » this approach for high-throughput protein functional validation and discovery, we experimentally annotate the families of ATP-binding proteins in Mtb. Our data experimentally validate prior in silico predictions of >250 ATPases and adenosine nucleotide-binding proteins, and reveal 73 hypothetical proteins as novel ATP-binding proteins. We identify adenosine cofactor interactions with many hypothetical proteins containing a diversity of unrelated sequences, providing a new and expanded view of adenosine nucleotide binding in Mtb. Furthermore, many of these hypothetical proteins are both unique to Mycobacteria and essential for infection, suggesting specialized functions in mycobacterial physiology and pathogenicity. Thus, we provide a generally applicable approach for high throughput protein function discovery and validation, and highlight several ways in which application of activity-based proteomics data can improve the quality of functional annotations to facilitate novel biological insights.« less

Evolutionary Turnover of Kinetochore Proteins: A Ship of Theseus?

PubMed

Drinnenberg, Ines A; Henikoff, Steven; Malik, Harmit S

2016-07-01

The kinetochore is a multiprotein complex that mediates the attachment of a eukaryotic chromosome to the mitotic spindle. The protein composition of kinetochores is similar across species as divergent as yeast and human. However, recent findings have revealed an unexpected degree of compositional diversity in kinetochores. For example, kinetochore proteins that are essential in some species have been lost in others, whereas new kinetochore proteins have emerged in other lineages. Even in lineages with similar kinetochore composition, individual kinetochore proteins have functionally diverged to acquire either essential or redundant roles. Thus, despite functional conservation, the repertoire of kinetochore proteins has undergone recurrent evolutionary turnover. Copyright © 2016 Elsevier Ltd. All rights reserved.

FunSimMat: a comprehensive functional similarity database

PubMed Central

Schlicker, Andreas; Albrecht, Mario

2008-01-01

Functional similarity based on Gene Ontology (GO) annotation is used in diverse applications like gene clustering, gene expression data analysis, protein interaction prediction and evaluation. However, there exists no comprehensive resource of functional similarity values although such a database would facilitate the use of functional similarity measures in different applications. Here, we describe FunSimMat (Functional Similarity Matrix, http://funsimmat.bioinf.mpi-inf.mpg.de/), a large new database that provides several different semantic similarity measures for GO terms. It offers various precomputed functional similarity values for proteins contained in UniProtKB and for protein families in Pfam and SMART. The web interface allows users to efficiently perform both semantic similarity searches with GO terms and functional similarity searches with proteins or protein families. All results can be downloaded in tab-delimited files for use with other tools. An additional XML–RPC interface gives automatic online access to FunSimMat for programs and remote services. PMID:17932054

Cross-biome metagenomic analyses of soil microbial communities and their functional attributes.

PubMed

Fierer, Noah; Leff, Jonathan W; Adams, Byron J; Nielsen, Uffe N; Bates, Scott Thomas; Lauber, Christian L; Owens, Sarah; Gilbert, Jack A; Wall, Diana H; Caporaso, J Gregory

2012-12-26

For centuries ecologists have studied how the diversity and functional traits of plant and animal communities vary across biomes. In contrast, we have only just begun exploring similar questions for soil microbial communities despite soil microbes being the dominant engines of biogeochemical cycles and a major pool of living biomass in terrestrial ecosystems. We used metagenomic sequencing to compare the composition and functional attributes of 16 soil microbial communities collected from cold deserts, hot deserts, forests, grasslands, and tundra. Those communities found in plant-free cold desert soils typically had the lowest levels of functional diversity (diversity of protein-coding gene categories) and the lowest levels of phylogenetic and taxonomic diversity. Across all soils, functional beta diversity was strongly correlated with taxonomic and phylogenetic beta diversity; the desert microbial communities were clearly distinct from the nondesert communities regardless of the metric used. The desert communities had higher relative abundances of genes associated with osmoregulation and dormancy, but lower relative abundances of genes associated with nutrient cycling and the catabolism of plant-derived organic compounds. Antibiotic resistance genes were consistently threefold less abundant in the desert soils than in the nondesert soils, suggesting that abiotic conditions, not competitive interactions, are more important in shaping the desert microbial communities. As the most comprehensive survey of soil taxonomic, phylogenetic, and functional diversity to date, this study demonstrates that metagenomic approaches can be used to build a predictive understanding of how microbial diversity and function vary across terrestrial biomes.

Constraints imposed by non-functional protein–protein interactions on gene expression and proteome size

PubMed Central

Zhang, Jingshan; Maslov, Sergei; Shakhnovich, Eugene I

2008-01-01

Crowded intracellular environments present a challenge for proteins to form functional specific complexes while reducing non-functional interactions with promiscuous non-functional partners. Here we show how the need to minimize the waste of resources to non-functional interactions limits the proteome diversity and the average concentration of co-expressed and co-localized proteins. Using the results of high-throughput Yeast 2-Hybrid experiments, we estimate the characteristic strength of non-functional protein–protein interactions. By combining these data with the strengths of specific interactions, we assess the fraction of time proteins spend tied up in non-functional interactions as a function of their overall concentration. This allows us to sketch the phase diagram for baker's yeast cells using the experimentally measured concentrations and subcellular localization of their proteins. The positions of yeast compartments on the phase diagram are consistent with our hypothesis that the yeast proteome has evolved to operate closely to the upper limit of its size, whereas keeping individual protein concentrations sufficiently low to reduce non-functional interactions. These findings have implication for conceptual understanding of intracellular compartmentalization, multicellularity and differentiation. PMID:18682700

Elucidating the ensemble of functionally-relevant transitions in protein systems with a robotics-inspired method

PubMed Central

2013-01-01

Background Many proteins tune their biological function by transitioning between different functional states, effectively acting as dynamic molecular machines. Detailed structural characterization of transition trajectories is central to understanding the relationship between protein dynamics and function. Computational approaches that build on the Molecular Dynamics framework are in principle able to model transition trajectories at great detail but also at considerable computational cost. Methods that delay consideration of dynamics and focus instead on elucidating energetically-credible conformational paths connecting two functionally-relevant structures provide a complementary approach. Effective sampling-based path planning methods originating in robotics have been recently proposed to produce conformational paths. These methods largely model short peptides or address large proteins by simplifying conformational space. Methods We propose a robotics-inspired method that connects two given structures of a protein by sampling conformational paths. The method focuses on small- to medium-size proteins, efficiently modeling structural deformations through the use of the molecular fragment replacement technique. In particular, the method grows a tree in conformational space rooted at the start structure, steering the tree to a goal region defined around the goal structure. We investigate various bias schemes over a progress coordinate for balance between coverage of conformational space and progress towards the goal. A geometric projection layer promotes path diversity. A reactive temperature scheme allows sampling of rare paths that cross energy barriers. Results and conclusions Experiments are conducted on small- to medium-size proteins of length up to 214 amino acids and with multiple known functionally-relevant states, some of which are more than 13Å apart of each-other. Analysis reveals that the method effectively obtains conformational paths connecting structural states that are significantly different. A detailed analysis on the depth and breadth of the tree suggests that a soft global bias over the progress coordinate enhances sampling and results in higher path diversity. The explicit geometric projection layer that biases the exploration away from over-sampled regions further increases coverage, often improving proximity to the goal by forcing the exploration to find new paths. The reactive temperature scheme is shown effective in increasing path diversity, particularly in difficult structural transitions with known high-energy barriers. PMID:24565158

Transcriptome-based identification of ABC transporters in the western tarnished plant bug lygus hesperus

USDA-ARS?s Scientific Manuscript database

ATP-binding cassette (ABC) transporters are a large superfamily of proteins that mediate diverse physiological functions by coupling ATP hydrolysis with substrate transport across lipid membranes. In insects, these proteins play roles in metabolism, development, eye pigmentation, and xenobiotic cle...

N-terminal acetylation -an Essential Protein Modification Emerges as an Important Regulator of Stress Responses.

PubMed

Linster, Eric; Wirtz, Markus

2018-06-26

N-terminal acetylation (NTA) is a prevalent protein modification in eukaryotes. The majority of proteins is acetylated at their N-terminus in a co-translational manner by ribosome-associated N-terminal acetyltransferases (NAT). However, the recent discovery of Golgi-membrane localized NATs in metazoan, and plastid-localized NATs in plants challenged the dogma of static, co-translational imprinting of the proteome by NTA. Indeed, NTA by the cytosolic NatA is highly dynamic and under hormonal control in plants. Such active control has not been evidenced yet in other eukaryotes and might be an adaptation to the sessile lifestyle of plants forcing them to cope with diverse environmental challenges. The function of NTA for individual proteins is distinct and yet unpredictable. In yeast and humans, NTA has been shown to affect protein-protein interactions, subcellular localization, folding, aggregation, or degradation of a handful of proteins. In particular, the impact of NTA on the protein-turnover is documented by diverse examples in yeast. Consequently, NTA has recently dicovered to be a degradation signal in a distinct branch of the N-end rule pathway ubiquitin-mediated proteolysis. In this review, we summarize the current knowledge on the NAT machinery in higher plants and discuss the potential function of NTA during biotic and abiotic stresses.

The innate capacity of proteins to protect against reactive radical species.

PubMed

Hamdy, Omar M; Alizadeh, Arman; Julian, Ryan R

2015-08-07

Maintaining redox homeostasis, or the balance of oxidant and antioxidant forces, is essential for proper cellular functioning in biology. Although the antioxidant nature of many small molecules such as vitamin c and glutathione have been thoroughly investigated, contributions to redox homeostasis from larger biomolecules have received less attention. Evidence has shown that some proteins are antioxidant (in a non-catalytic sense), but large scale examination of this property for a diverse set of proteins has proven difficult. Herein, radical-directed dissociation mass spectrometry (RDD-MS) is used to examine the antioxidant capacity of a series of proteins with diverse biological roles, persistence intervals, and localizations. Digestion of these proteins reveals that all contain antioxidant peptide regions. Examination of the amino acid content of the antioxidant peptides does not reveal significant differences relative to normal peptides, suggesting that sequence may be more important than residue content. Sequence homology analysis across organisms reveals that antioxidant regions are frequently conserved, although many of these regions are also known to have other functions which may have influenced evolutionary pressure. Regardless of the origin, it is clear that many proteins may play secondary roles as sacrificial antioxidants within the cellular milieu.

Functional evolution and structural conservation in chimeric cytochromes p450: calibrating a structure-guided approach.

PubMed

Otey, Christopher R; Silberg, Jonathan J; Voigt, Christopher A; Endelman, Jeffrey B; Bandara, Geethani; Arnold, Frances H

2004-03-01

Recombination generates chimeric proteins whose ability to fold depends on minimizing structural perturbations that result when portions of the sequence are inherited from different parents. These chimeric sequences can display functional properties characteristic of the parents or acquire entirely new functions. Seventeen chimeras were generated from two CYP102 members of the functionally diverse cytochrome p450 family. Chimeras predicted to have limited structural disruption, as defined by the SCHEMA algorithm, displayed CO binding spectra characteristic of folded p450s. Even this small population exhibited significant functional diversity: chimeras displayed altered substrate specificities, a wide range in thermostabilities, up to a 40-fold increase in peroxidase activity, and ability to hydroxylate a substrate toward which neither parent heme domain shows detectable activity. These results suggest that SCHEMA-guided recombination can be used to generate diverse p450s for exploring function evolution within the p450 structural framework.

Mitochondrial Protein Interaction Mapping Identifies Regulators of Respiratory Chain Function.

PubMed

Floyd, Brendan J; Wilkerson, Emily M; Veling, Mike T; Minogue, Catie E; Xia, Chuanwu; Beebe, Emily T; Wrobel, Russell L; Cho, Holly; Kremer, Laura S; Alston, Charlotte L; Gromek, Katarzyna A; Dolan, Brendan K; Ulbrich, Arne; Stefely, Jonathan A; Bohl, Sarah L; Werner, Kelly M; Jochem, Adam; Westphall, Michael S; Rensvold, Jarred W; Taylor, Robert W; Prokisch, Holger; Kim, Jung-Ja P; Coon, Joshua J; Pagliarini, David J

2016-08-18

Mitochondria are essential for numerous cellular processes, yet hundreds of their proteins lack robust functional annotation. To reveal functions for these proteins (termed MXPs), we assessed condition-specific protein-protein interactions for 50 select MXPs using affinity enrichment mass spectrometry. Our data connect MXPs to diverse mitochondrial processes, including multiple aspects of respiratory chain function. Building upon these observations, we validated C17orf89 as a complex I (CI) assembly factor. Disruption of C17orf89 markedly reduced CI activity, and its depletion is found in an unresolved case of CI deficiency. We likewise discovered that LYRM5 interacts with and deflavinates the electron-transferring flavoprotein that shuttles electrons to coenzyme Q (CoQ). Finally, we identified a dynamic human CoQ biosynthetic complex involving multiple MXPs whose topology we map using purified components. Collectively, our data lend mechanistic insight into respiratory chain-related activities and prioritize hundreds of additional interactions for further exploration of mitochondrial protein function. Copyright © 2016 Elsevier Inc. All rights reserved.

New frontiers: discovering cilia-independent functions of cilia proteins.

PubMed

Vertii, Anastassiia; Bright, Alison; Delaval, Benedicte; Hehnly, Heidi; Doxsey, Stephen

2015-10-01

In most vertebrates, mitotic spindles and primary cilia arise from a common origin, the centrosome. In non-cycling cells, the centrosome is the template for primary cilia assembly and, thus, is crucial for their associated sensory and signaling functions. During mitosis, the duplicated centrosomes mature into spindle poles, which orchestrate mitotic spindle assembly, chromosome segregation, and orientation of the cell division axis. Intriguingly, both cilia and spindle poles are centrosome-based, functionally distinct structures that require the action of microtubule-mediated, motor-driven transport for their assembly. Cilia proteins have been found at non-cilia sites, where they have distinct functions, illustrating a diverse and growing list of cellular processes and structures that utilize cilia proteins for crucial functions. In this review, we discuss cilia-independent functions of cilia proteins and re-evaluate their potential contributions to "cilia" disorders. © 2015 The Authors.

Artificially Engineered Protein Polymers.

PubMed

Yang, Yun Jung; Holmberg, Angela L; Olsen, Bradley D

2017-06-07

Modern polymer science increasingly requires precise control over macromolecular structure and properties for engineering advanced materials and biomedical systems. The application of biological processes to design and synthesize artificial protein polymers offers a means for furthering macromolecular tunability, enabling polymers with dispersities of ∼1.0 and monomer-level sequence control. Taking inspiration from materials evolved in nature, scientists have created modular building blocks with simplified monomer sequences that replicate the function of natural systems. The corresponding protein engineering toolbox has enabled the systematic development of complex functional polymeric materials across areas as diverse as adhesives, responsive polymers, and medical materials. This review discusses the natural proteins that have inspired the development of key building blocks for protein polymer engineering and the function of these elements in material design. The prospects and progress for scalable commercialization of protein polymers are reviewed, discussing both technology needs and opportunities.

Hierarchy and extremes in selections from pools of randomized proteins

PubMed Central

Boyer, Sébastien; Biswas, Dipanwita; Kumar Soshee, Ananda; Scaramozzino, Natale; Nizak, Clément; Rivoire, Olivier

2016-01-01

Variation and selection are the core principles of Darwinian evolution, but quantitatively relating the diversity of a population to its capacity to respond to selection is challenging. Here, we examine this problem at a molecular level in the context of populations of partially randomized proteins selected for binding to well-defined targets. We built several minimal protein libraries, screened them in vitro by phage display, and analyzed their response to selection by high-throughput sequencing. A statistical analysis of the results reveals two main findings. First, libraries with the same sequence diversity but built around different “frameworks” typically have vastly different responses; second, the distribution of responses of the best binders in a library follows a simple scaling law. We show how an elementary probabilistic model based on extreme value theory rationalizes the latter finding. Our results have implications for designing synthetic protein libraries, estimating the density of functional biomolecules in sequence space, characterizing diversity in natural populations, and experimentally investigating evolvability (i.e., the potential for future evolution). PMID:26969726

Hierarchy and extremes in selections from pools of randomized proteins.

PubMed

Boyer, Sébastien; Biswas, Dipanwita; Kumar Soshee, Ananda; Scaramozzino, Natale; Nizak, Clément; Rivoire, Olivier

2016-03-29

Variation and selection are the core principles of Darwinian evolution, but quantitatively relating the diversity of a population to its capacity to respond to selection is challenging. Here, we examine this problem at a molecular level in the context of populations of partially randomized proteins selected for binding to well-defined targets. We built several minimal protein libraries, screened them in vitro by phage display, and analyzed their response to selection by high-throughput sequencing. A statistical analysis of the results reveals two main findings. First, libraries with the same sequence diversity but built around different "frameworks" typically have vastly different responses; second, the distribution of responses of the best binders in a library follows a simple scaling law. We show how an elementary probabilistic model based on extreme value theory rationalizes the latter finding. Our results have implications for designing synthetic protein libraries, estimating the density of functional biomolecules in sequence space, characterizing diversity in natural populations, and experimentally investigating evolvability (i.e., the potential for future evolution).

Identifying DNA-binding proteins using structural motifs and the electrostatic potential

PubMed Central

Shanahan, Hugh P.; Garcia, Mario A.; Jones, Susan; Thornton, Janet M.

2004-01-01

Robust methods to detect DNA-binding proteins from structures of unknown function are important for structural biology. This paper describes a method for identifying such proteins that (i) have a solvent accessible structural motif necessary for DNA-binding and (ii) a positive electrostatic potential in the region of the binding region. We focus on three structural motifs: helix–turn-helix (HTH), helix–hairpin–helix (HhH) and helix–loop–helix (HLH). We find that the combination of these variables detect 78% of proteins with an HTH motif, which is a substantial improvement over previous work based purely on structural templates and is comparable to more complex methods of identifying DNA-binding proteins. Similar true positive fractions are achieved for the HhH and HLH motifs. We see evidence of wide evolutionary diversity for DNA-binding proteins with an HTH motif, and much smaller diversity for those with an HhH or HLH motif. PMID:15356290

Detection of Diverse and High Molecular Weight Nesprin-1 and Nesprin-2 Isoforms Using Western Blotting.

PubMed

Carthew, James; Karakesisoglou, Iakowos

2016-01-01

Heavily utilized in cell and molecular biology, western blotting is considered a crucial technique for the detection and quantification of proteins within complex mixtures. In particular, the detection of members of the nesprin (nuclear envelope spectrin repeat protein) family has proven difficult to analyze due to their substantial isoform diversity, molecular weight variation, and the sheer size of both nesprin-1 and nesprin-2 giant protein variants (>800 kDa). Nesprin isoforms contain distinct domain signatures, perform differential cytoskeletal associations, occupy different subcellular compartments, and vary in their tissue expression profiles. This structural and functional variance highlights the need to distinguish between the full range of proteins within the nesprin protein family, allowing for greater understanding of their specific roles in cell biology and disease. Herein, we describe a western blotting protocol modified for the detection of low to high molecular weight (50-1000 kDa) nesprin proteins.

Telomere- and Telomerase-Associated Proteins and Their Functions in the Plant Cell

PubMed Central

Procházková Schrumpfová, Petra; Schořová, Šárka; Fajkus, Jiří

2016-01-01

Telomeres, as physical ends of linear chromosomes, are targets of a number of specific proteins, including primarily telomerase reverse transcriptase. Access of proteins to the telomere may be affected by a number of diverse factors, e.g., protein interaction partners, local DNA or chromatin structures, subcellular localization/trafficking, or simply protein modification. Knowledge of composition of the functional nucleoprotein complex of plant telomeres is only fragmentary. Moreover, the plant telomeric repeat binding proteins that were characterized recently appear to also be involved in non-telomeric processes, e.g., ribosome biogenesis. This interesting finding was not totally unexpected since non-telomeric functions of yeast or animal telomeric proteins, as well as of telomerase subunits, have been reported for almost a decade. Here we summarize known facts about the architecture of plant telomeres and compare them with the well-described composition of telomeres in other organisms. PMID:27446102

ELMO Domains, Evolutionary and Functional Characterization of a Novel GTPase-activating Protein (GAP) Domain for Arf Protein Family GTPases*

PubMed Central

East, Michael P.; Bowzard, J. Bradford; Dacks, Joel B.; Kahn, Richard A.

2012-01-01

The human family of ELMO domain-containing proteins (ELMODs) consists of six members and is defined by the presence of the ELMO domain. Within this family are two subclassifications of proteins, based on primary sequence conservation, protein size, and domain architecture, deemed ELMOD and ELMO. In this study, we used homology searching and phylogenetics to identify ELMOD family homologs in genomes from across eukaryotic diversity. This demonstrated not only that the protein family is ancient but also that ELMOs are potentially restricted to the supergroup Opisthokonta (Metazoa and Fungi), whereas proteins with the ELMOD organization are found in diverse eukaryotes and thus were likely the form present in the last eukaryotic common ancestor. The segregation of the ELMO clade from the larger ELMOD group is consistent with their contrasting functions as unconventional Rac1 guanine nucleotide exchange factors and the Arf family GTPase-activating proteins, respectively. We used unbiased, phylogenetic sorting and sequence alignments to identify the most highly conserved residues within the ELMO domain to identify a putative GAP domain within the ELMODs. Three independent but complementary assays were used to provide an initial characterization of this domain. We identified a highly conserved arginine residue critical for both the biochemical and cellular GAP activity of ELMODs. We also provide initial evidence of the function of human ELMOD1 as an Arf family GAP at the Golgi. These findings provide the basis for the future study of the ELMOD family of proteins and a new avenue for the study of Arf family GTPases. PMID:23014990

Functional diversity of secreted cestode Kunitz proteins: Inhibition of serine peptidases and blockade of cation channels

PubMed Central

Fló, Martín; Margenat, Mariana; Pellizza, Leonardo; Durán, Rosario; Salceda, Emilio; Alvarez, Beatriz

2017-01-01

We previously reported a multigene family of monodomain Kunitz proteins from Echinococcus granulosus (EgKU-1-EgKU-8), and provided evidence that some EgKUs are secreted by larval worms to the host interface. In addition, functional studies and homology modeling suggested that, similar to monodomain Kunitz families present in animal venoms, the E. granulosus family could include peptidase inhibitors as well as channel blockers. Using enzyme kinetics and whole-cell patch-clamp, we now demonstrate that the EgKUs are indeed functionally diverse. In fact, most of them behaved as high affinity inhibitors of either chymotrypsin (EgKU-2-EgKU-3) or trypsin (EgKU-5-EgKU-8). In contrast, the close paralogs EgKU-1 and EgKU-4 blocked voltage-dependent potassium channels (Kv); and also pH-dependent sodium channels (ASICs), while showing null (EgKU-1) or marginal (EgKU-4) peptidase inhibitory activity. We also confirmed the presence of EgKUs in secretions from other parasite stages, notably from adult worms and metacestodes. Interestingly, data from genome projects reveal that at least eight additional monodomain Kunitz proteins are encoded in the genome; that particular EgKUs are up-regulated in various stages; and that analogous Kunitz families exist in other medically important cestodes, but not in trematodes. Members of this expanded family of secreted cestode proteins thus have the potential to block, through high affinity interactions, the function of host counterparts (either peptidases or cation channels) and contribute to the establishment and persistence of infection. From a more general perspective, our results confirm that multigene families of Kunitz inhibitors from parasite secretions and animal venoms display a similar functional diversity and thus, that host-parasite co-evolution may also drive the emergence of a new function associated with the Kunitz scaffold. PMID:28192542

Structure and function of seed storage proteins in faba bean (Vicia faba L.).

PubMed

Liu, Yujiao; Wu, Xuexia; Hou, Wanwei; Li, Ping; Sha, Weichao; Tian, Yingying

2017-05-01

The protein subunit is the most important basic unit of protein, and its study can unravel the structure and function of seed storage proteins in faba bean. In this study, we identified six specific protein subunits in Faba bean (cv. Qinghai 13) combining liquid chromatography (LC), liquid chromatography-electronic spray ionization mass (LC-ESI-MS/MS) and bio-information technology. The results suggested a diversity of seed storage proteins in faba bean, and a total of 16 proteins (four GroEL molecular chaperones and 12 plant-specific proteins) were identified from 97-, 96-, 64-, 47-, 42-, and 38-kD-specific protein subunits in faba bean based on the peptide sequence. We also analyzed the composition and abundance of the amino acids, the physicochemical characteristics, secondary structure, three-dimensional structure, transmembrane domain, and possible subcellular localization of these identified proteins in faba bean seed, and finally predicted function and structure. The three-dimensional structures were generated based on homologous modeling, and the protein function was analyzed based on the annotation from the non-redundant protein database (NR database, NCBI) and function analysis of optimal modeling. The objective of this study was to identify the seed storage proteins in faba bean and confirm the structure and function of these proteins. Our results can be useful for the study of protein nutrition and achieve breeding goals for optimal protein quality in faba bean.

Hydrophobicity diversity in globular and nonglobular proteins measured with the Gini index.

PubMed

Carugo, Oliviero

2017-12-01

Amino acids and their properties are variably distributed in proteins and different compositions determine all protein features, ranging from solubility to stability and functionality. Gini index, a tool to estimate distribution uniformity, is widely used in macroeconomics and has numerous statistical applications. Here, Gini index is used to analyze the distribution of hydrophobicity in proteins and to compare hydrophobicity distribution in globular and intrinsically disordered proteins. Based on the analysis of carefully selected high-quality data sets of proteins extracted from the Protein Data Bank (http://www.rcsb.org) and from the DisProt database (http://www.disprot.org/), it is observed that hydrophobicity is distributed in a more diverse way in intrinsically disordered proteins than in folded and soluble globular proteins. This correlates with the observation that the amino acid composition deviates from the uniformity (estimate with the Shannon and the Gini-Simpson indices) more in intrinsically disordered proteins than in globular and soluble proteins. Although statistical tools tike the Gini index have received little attention in molecular biology, these results show that they allow one to estimate sequence diversity and that they are useful to delineate trends that can hardly be described, otherwise, in simple and concise ways. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Significant expansion of exon-bordering protein domains during animal proteome evolution

PubMed Central

Liu, Mingyi; Walch, Heiko; Wu, Shaoping; Grigoriev, Andrei

2005-01-01

We present evidence of remarkable genome-wide mobility and evolutionary expansion for a class of protein domains whose borders locate close to the borders of their encoding exons. These exon-bordering domains are more numerous and widely distributed in the human genome than other domains. They also co-occur with more diverse domains to form a larger variety of domain architectures in human proteins. A systematic comparison of nine animal genomes from nematodes to mammals revealed that exon-bordering domains expanded faster than other protein domains in both abundance and distribution, as well as the diversity of co-occurring domains and the domain architectures of harboring proteins. Furthermore, exon-bordering domains exhibited a particularly strong preference for class 1-1 intron phase. Our findings suggest that exon-bordering domains were amplified and interchanged within a genome more often and/or more successfully than other domains during evolution, probably the result of extensive exon shuffling and gene duplication events. The diverse biological functions of these domains underscore the important role they play in the expansion and diversification of animal proteomes. PMID:15640447

Biologically active LIL proteins built with minimal chemical diversity

PubMed Central

Heim, Erin N.; Marston, Jez L.; Federman, Ross S.; Edwards, Anne P. B.; Karabadzhak, Alexander G.; Petti, Lisa M.; Engelman, Donald M.; DiMaio, Daniel

2015-01-01

We have constructed 26-amino acid transmembrane proteins that specifically transform cells but consist of only two different amino acids. Most proteins are long polymers of amino acids with 20 or more chemically distinct side-chains. The artificial transmembrane proteins reported here are the simplest known proteins with specific biological activity, consisting solely of an initiating methionine followed by specific sequences of leucines and isoleucines, two hydrophobic amino acids that differ only by the position of a methyl group. We designate these proteins containing leucine (L) and isoleucine (I) as LIL proteins. These proteins functionally interact with the transmembrane domain of the platelet-derived growth factor β-receptor and specifically activate the receptor to transform cells. Complete mutagenesis of these proteins identified individual amino acids required for activity, and a protein consisting solely of leucines, except for a single isoleucine at a particular position, transformed cells. These surprisingly simple proteins define the minimal chemical diversity sufficient to construct proteins with specific biological activity and change our view of what can constitute an active protein in a cellular context. PMID:26261320

Effects of the NO/soluble guanylate cyclase/cGMP system on the functions of human platelets.

PubMed

Makhoul, Stephanie; Walter, Elena; Pagel, Oliver; Walter, Ulrich; Sickmann, Albert; Gambaryan, Stepan; Smolenski, Albert; Zahedi, René P; Jurk, Kerstin

2018-06-01

Platelets are circulating sentinels of vascular integrity and are activated, inhibited, or modulated by multiple hormones, vasoactive substances or drugs. Endothelium- or drug-derived NO strongly inhibits platelet activation via activation of the soluble guanylate cyclase (sGC) and cGMP elevation, often in synergy with cAMP-elevation by prostacyclin. However, the molecular mechanisms and diversity of cGMP effects in platelets are poorly understood and sometimes controversial. Recently, we established the quantitative human platelet proteome, the iloprost/prostacyclin/cAMP/protein kinase A (PKA)-regulated phosphoproteome, and the interactions of the ADP- and iloprost/prostacyclin-affected phosphoproteome. We also showed that the sGC stimulator riociguat is in vitro a highly specific inhibitor, via cGMP, of various functions of human platelets. Here, we review the regulatory role of the cGMP/protein kinase G (PKG) system in human platelet function, and our current approaches to establish and analyze the phosphoproteome after selective stimulation of the sGC/cGMP pathway by NO donors and riociguat. Present data indicate an extensive and diverse NO/riociguat/cGMP phosphoproteome, which has to be compared with the cAMP phosphoproteome. In particular, sGC/cGMP-regulated phosphorylation of many membrane proteins, G-proteins and their regulators, signaling molecules, protein kinases, and proteins involved in Ca 2+ regulation, suggests that the sGC/cGMP system targets multiple signaling networks rather than a limited number of PKG substrate proteins. Copyright © 2018 Elsevier Inc. All rights reserved.

Defining the human deubiquitinating enzyme interaction landscape.

PubMed

Sowa, Mathew E; Bennett, Eric J; Gygi, Steven P; Harper, J Wade

2009-07-23

Deubiquitinating enzymes (Dubs) function to remove covalently attached ubiquitin from proteins, thereby controlling substrate activity and/or abundance. For most Dubs, their functions, targets, and regulation are poorly understood. To systematically investigate Dub function, we initiated a global proteomic analysis of Dubs and their associated protein complexes. This was accomplished through the development of a software platform called CompPASS, which uses unbiased metrics to assign confidence measurements to interactions from parallel nonreciprocal proteomic data sets. We identified 774 candidate interacting proteins associated with 75 Dubs. Using Gene Ontology, interactome topology classification, subcellular localization, and functional studies, we link Dubs to diverse processes, including protein turnover, transcription, RNA processing, DNA damage, and endoplasmic reticulum-associated degradation. This work provides the first glimpse into the Dub interaction landscape, places previously unstudied Dubs within putative biological pathways, and identifies previously unknown interactions and protein complexes involved in this increasingly important arm of the ubiquitin-proteasome pathway.

Defining the Human Deubiquitinating Enzyme Interaction Landscape

PubMed Central

Sowa, Mathew E.; Bennett, Eric J.; Gygi, Steven P.; Harper, J. Wade

2009-01-01

Summary Deubiquitinating enzymes (Dubs) function to remove covalently attached ubiquitin from proteins, thereby controlling substrate activity and/or abundance. For most Dubs, their functions, targets, and regulation are poorly understood. To systematically investigate Dub function, we initiated a global proteomic analysis of Dubs and their associated protein complexes. This was accomplished through the development of a software platform, called CompPASS, which uses unbiased metrics to assign confidence measurements to interactions from parallel non-reciprocal proteomic datasets. We identified 774 candidate interacting proteins associated with 75 Dubs. Using Gene Ontology, interactome topology classification, sub-cellular localization and functional studies, we link Dubs to diverse processes, including protein turnover, transcription, RNA processing, DNA damage, and endoplasmic reticulum-associated degradation. This work provides the first glimpse into the Dub interaction landscape, places previously unstudied Dubs within putative biological pathways, and identifies previously unknown interactions and protein complexes involved in this increasingly important arm of the ubiquitin-proteasome pathway. PMID:19615732

Insights into Structural and Mechanistic Features of Viral IRES Elements

PubMed Central

Martinez-Salas, Encarnacion; Francisco-Velilla, Rosario; Fernandez-Chamorro, Javier; Embarek, Azman M.

2018-01-01

Internal ribosome entry site (IRES) elements are cis-acting RNA regions that promote internal initiation of protein synthesis using cap-independent mechanisms. However, distinct types of IRES elements present in the genome of various RNA viruses perform the same function despite lacking conservation of sequence and secondary RNA structure. Likewise, IRES elements differ in host factor requirement to recruit the ribosomal subunits. In spite of this diversity, evolutionarily conserved motifs in each family of RNA viruses preserve sequences impacting on RNA structure and RNA–protein interactions important for IRES activity. Indeed, IRES elements adopting remarkable different structural organizations contain RNA structural motifs that play an essential role in recruiting ribosomes, initiation factors and/or RNA-binding proteins using different mechanisms. Therefore, given that a universal IRES motif remains elusive, it is critical to understand how diverse structural motifs deliver functions relevant for IRES activity. This will be useful for understanding the molecular mechanisms beyond cap-independent translation, as well as the evolutionary history of these regulatory elements. Moreover, it could improve the accuracy to predict IRES-like motifs hidden in genome sequences. This review summarizes recent advances on the diversity and biological relevance of RNA structural motifs for viral IRES elements. PMID:29354113

Structural and functional diversity of cadherin at the adherens junction

PubMed Central

2011-01-01

Adhesion between cells is essential to the evolution of multicellularity. Indeed, morphogenesis in animals requires firm but flexible intercellular adhesions that are mediated by subcellular structures like the adherens junction (AJ). A key component of AJs is classical cadherins, a group of transmembrane proteins that maintain dynamic cell–cell associations in many animal species. An evolutionary reconstruction of cadherin structure and function provides a comprehensive framework with which to appreciate the diversity of morphogenetic mechanisms in animals. PMID:21708975

Inter-individual variation in expression: a missing link in biomarker biology?

PubMed

Little, Peter F R; Williams, Rohan B H; Wilkins, Marc R

2009-01-01

The past decade has seen an explosion of variation data demonstrating that diversity of both protein-coding sequences and of regulatory elements of protein-coding genes is common and of functional importance. In this article, we argue that genetic diversity can no longer be ignored in studies of human biology, even research projects without explicit genetic experimental design, and that this knowledge can, and must, inform research. By way of illustration, we focus on the potential role of genetic data in case-control studies to identify and validate cancer protein biomarkers. We argue that a consideration of genetics, in conjunction with proteomic biomarker discovery projects, should improve the proportion of biomarkers that can accurately classify patients.

Chromosome ends: different sequences may provide conserved functions.

PubMed

Louis, Edward J; Vershinin, Alexander V

2005-07-01

The structures of specific chromosome regions, centromeres and telomeres, present a number of puzzles. As functions performed by these regions are ubiquitous and essential, their DNA, proteins and chromatin structure are expected to be conserved. Recent studies of centromeric DNA from human, Drosophila and plant species have demonstrated that a hidden universal centromere-specific sequence is highly unlikely. The DNA of telomeres is more conserved consisting of a tandemly repeated 6-8 bp Arabidopsis-like sequence in a majority of organisms as diverse as protozoan, fungi, mammals and plants. However, there are alternatives to short DNA repeats at the ends of chromosomes and for telomere elongation by telomerase. Here we focus on the similarities and diversity that exist among the structural elements, DNA sequences and proteins, that make up terminal domains (telomeres and subtelomeres), and how organisms use these in different ways to fulfil the functions of end-replication and end-protection. Copyright (c) 2005 Wiley Periodicals, Inc.

The mGluR theory of fragile X mental retardation.

PubMed

Bear, Mark F; Huber, Kimberly M; Warren, Stephen T

2004-07-01

Many of the diverse functional consequences of activating group 1 metabotropic glutamate receptors require translation of pre-existing mRNA near synapses. One of these consequences is long-term depression (LTD) of transmission at hippocampal synapses. Loss of fragile X mental retardation protein (FMRP), the defect responsible for fragile X syndrome in humans, increases LTD in mouse hippocampus. This finding is consistent with the growing evidence that FMRP normally functions as a repressor of translation of specific mRNAs. Here we present a theory that can account for diverse neurological and psychiatric aspects of fragile X syndrome, based on the assumption that many of the protein-synthesis-dependent functions of metabotropic receptors are exaggerated in fragile X syndrome. The theory suggests new directions for basic research as well as novel therapeutic approaches for the treatment of humans with fragile X, the most frequent inherited cause of mental retardation and an identified cause of autism.

From hatching to dispatching: the multiple cellular roles of the Hsp70 molecular chaperone machinery.

PubMed

Meimaridou, Eirini; Gooljar, Sakina B; Chapple, J Paul

2009-01-01

Molecular chaperones are best recognized for their roles in de novo protein folding and the cellular response to stress. However, many molecular chaperones, and in particular the Hsp70 chaperone machinery, have multiple diverse cellular functions. At the molecular level, chaperones are mediators of protein conformational change. To facilitate conformational change of client/substrate proteins, in manifold contexts, chaperone power must be closely regulated and harnessed to specific cellular locales--this is controlled by cochaperones. This review considers specialized functions of the Hsp70 chaperone machinery mediated by its cochaperones. We focus on vesicular trafficking, protein degradation and a potential role in G protein-coupled receptor processing.

Drosophila Pelle phosphorylates Dichaete protein and influences its subcellular distribution in developing oocytes.

PubMed

Mutsuddi, Mousumi; Mukherjee, Ashim; Shen, Baohe; Manley, James L; Nambu, John R

2010-01-01

The Drosophila Dichaete gene encodes a member of the Sox family of high mobility group (HMG) domain proteins that have crucial gene regulatory functions in diverse developmental processes. The subcellular localization and transcriptional regulatory activities of Sox proteins can be regulated by several post-translational modifications. To identify genes that functionally interact with Dichaete, we undertook a genetic modifier screen based on a Dichaete gain-of-function phenotype in the adult eye. Mutations in several genes, including decapentaplegic, engrailed and pelle, behaved as dominant modifiers of this eye phenotype. Further analysis of pelle mutants revealed that loss of pelle function results in alterations in the distinctive cytoplasmic distribution of Dichaete protein within the developing oocyte, as well as defects in the elaboration of individual egg chambers. The death domain-containing region of the Pelle protein kinase was found to associate with both Dichaete and mouse Sox2 proteins, and Pelle can phosphorylate Dichaete protein in vitro. Overall, these findings reveal that maternal functions of pelle are essential for proper localization of Dichaete protein in the oocyte and normal egg chamber formation. Dichaete appears to be a novel phosphorylation substrate for Pelle and may function in a Pelle-dependent signaling pathway during oogenesis.

Rapid directed evolution of stabilized proteins with cellular high-throughput encapsulation solubilization and screening (CHESS).

PubMed

Yong, K J; Scott, D J

2015-03-01

Directed evolution is a powerful method for engineering proteins towards user-defined goals and has been used to generate novel proteins for industrial processes, biological research and drug discovery. Typical directed evolution techniques include cellular display, phage display, ribosome display and water-in-oil compartmentalization, all of which physically link individual members of diverse gene libraries to their translated proteins. This allows the screening or selection for a desired protein function and subsequent isolation of the encoding gene from diverse populations. For biotechnological and industrial applications there is a need to engineer proteins that are functional under conditions that are not compatible with these techniques, such as high temperatures and harsh detergents. Cellular High-throughput Encapsulation Solubilization and Screening (CHESS), is a directed evolution method originally developed to engineer detergent-stable G proteins-coupled receptors (GPCRs) for structural biology. With CHESS, library-transformed bacterial cells are encapsulated in detergent-resistant polymers to form capsules, which serve to contain mutant genes and their encoded proteins upon detergent mediated solubilization of cell membranes. Populations of capsules can be screened like single cells to enable rapid isolation of genes encoding detergent-stable protein mutants. To demonstrate the general applicability of CHESS to other proteins, we have characterized the stability and permeability of CHESS microcapsules and employed CHESS to generate thermostable, sodium dodecyl sulfate (SDS) resistant green fluorescent protein (GFP) mutants, the first soluble proteins to be engineered using CHESS. © 2014 Wiley Periodicals, Inc.

Pipeline for inferring protein function from dynamics using coarse-grained molecular mechanics forcefield.

PubMed

Bhadra, Pratiti; Pal, Debnath

2017-04-01

Dynamics is integral to the function of proteins, yet the use of molecular dynamics (MD) simulation as a technique remains under-explored for molecular function inference. This is more important in the context of genomics projects where novel proteins are determined with limited evolutionary information. Recently we developed a method to match the query protein's flexible segments to infer function using a novel approach combining analysis of residue fluctuation-graphs and auto-correlation vectors derived from coarse-grained (CG) MD trajectory. The method was validated on a diverse dataset with sequence identity between proteins as low as 3%, with high function-recall rates. Here we share its implementation as a publicly accessible web service, named DynFunc (Dynamics Match for Function) to query protein function from ≥1 µs long CG dynamics trajectory information of protein subunits. Users are provided with the custom-developed coarse-grained molecular mechanics (CGMM) forcefield to generate the MD trajectories for their protein of interest. On upload of trajectory information, the DynFunc web server identifies specific flexible regions of the protein linked to putative molecular function. Our unique application does not use evolutionary information to infer molecular function from MD information and can, therefore, work for all proteins, including moonlighting and the novel ones, whenever structural information is available. Our pipeline is expected to be of utility to all structural biologists working with novel proteins and interested in moonlighting functions. Copyright © 2017 Elsevier Ltd. All rights reserved.

Evolution at protein ends: major contribution of alternative transcription initiation and termination to the transcriptome and proteome diversity in mammals

PubMed Central

Shabalina, Svetlana A.; Ogurtsov, Aleksey Y.; Spiridonov, Nikolay A.; Koonin, Eugene V.

2014-01-01

Alternative splicing (AS), alternative transcription initiation (ATI) and alternative transcription termination (ATT) create the extraordinary complexity of transcriptomes and make key contributions to the structural and functional diversity of mammalian proteomes. Analysis of mammalian genomic and transcriptomic data shows that contrary to the traditional view, the joint contribution of ATI and ATT to the transcriptome and proteome diversity is quantitatively greater than the contribution of AS. Although the mean numbers of protein-coding constitutive and alternative nucleotides in gene loci are nearly identical, their distribution along the transcripts is highly non-uniform. On average, coding exons in the variable 5′ and 3′ transcript ends that are created by ATI and ATT contain approximately four times more alternative nucleotides than core protein-coding regions that diversify exclusively via AS. Short upstream exons that encompass alternative 5′-untranslated regions and N-termini of proteins evolve under strong nucleotide-level selection whereas in 3′-terminal exons that encode protein C-termini, protein-level selection is significantly stronger. The groups of genes that are subject to ATI and ATT show major differences in biological roles, expression and selection patterns. PMID:24792168

TULIPs: tunable, light-controlled interacting protein tags for cell biology.

PubMed

Strickland, Devin; Lin, Yuan; Wagner, Elizabeth; Hope, C Matthew; Zayner, Josiah; Antoniou, Chloe; Sosnick, Tobin R; Weiss, Eric L; Glotzer, Michael

2012-03-04

Naturally photoswitchable proteins offer a means of directly manipulating the formation of protein complexes that drive a diversity of cellular processes. We developed tunable light-inducible dimerization tags (TULIPs) based on a synthetic interaction between the LOV2 domain of Avena sativa phototropin 1 (AsLOV2) and an engineered PDZ domain (ePDZ). TULIPs can recruit proteins to diverse structures in living yeast and mammalian cells, either globally or with precise spatial control using a steerable laser. The equilibrium binding and kinetic parameters of the interaction are tunable by mutation, making TULIPs readily adaptable to signaling pathways with varying sensitivities and response times. We demonstrate the utility of TULIPs by conferring light sensitivity to functionally distinct components of the yeast mating pathway and by directing the site of cell polarization.

Pharmacoperone drugs: targeting misfolded proteins causing lysosomal storage-, ion channels-, and G protein-coupled receptors-associated conformational disorders.

PubMed

Hou, Zhi-Shuai; Ulloa-Aguirre, Alfredo; Tao, Ya-Xiong

2018-06-01

Conformational diseases are caused by structurally abnormal proteins that cannot fold properly and achieve their native conformation. Misfolded proteins frequently originate from genetic mutations that may lead to loss-of-function diseases involving a variety of structurally diverse proteins including enzymes, ion channels, and membrane receptors. Pharmacoperones are small molecules that cross the cell surface plasma membrane and reach their target proteins within the cell, serving as molecular scaffolds to stabilize the native conformation of misfolded or well-folded but destabilized proteins, to prevent their degradation and promote correct trafficking to their functional site of action. Because of their high specificity toward the target protein, pharmacoperones are currently the focus of intense investigation as therapy for several conformational diseases. Areas covered: This review summarizes data on the mechanisms leading to protein misfolding and the use of pharmacoperone drugs as an experimental approach to rescue function of distinct misfolded/misrouted proteins associated with a variety of diseases, such as lysosomal storage diseases, channelopathies, and G protein-coupled receptor misfolding diseases. Expert commentary: The fact that many misfolded proteins may retain function, offers a unique therapeutic opportunity to cure disease by directly correcting misrouting through administering pharmacoperone drugs thereby rescuing function of disease-causing, conformationally abnormal proteins.

Peroxisome biogenesis, protein targeting mechanisms and PEX gene functions in plants.

PubMed

Cross, Laura L; Ebeed, Heba Talat; Baker, Alison

2016-05-01

Peroxisomes play diverse and important roles in plants. The functions of peroxisomes are dependent upon their steady state protein composition which in turn reflects the balance of formation and turnover of the organelle. Protein import and turnover of constituent peroxisomal proteins are controlled by the state of cell growth and environment. The evolutionary origin of the peroxisome and the role of the endoplasmic reticulum in peroxisome biogenesis are discussed, as informed by studies of the trafficking of peroxisome membrane proteins. The process of matrix protein import in plants and its similarities and differences with peroxisomes in other organisms is presented and discussed in the context of peroxin distribution across the green plants. Copyright © 2015 Elsevier B.V. All rights reserved.

Candida albicans Agglutinin-Like Sequence (Als) Family Vignettes: A Review of Als Protein Structure and Function

PubMed Central

Hoyer, Lois L.; Cota, Ernesto

2016-01-01

Approximately two decades have passed since the description of the first gene in the Candida albicans ALS (agglutinin-like sequence) family. Since that time, much has been learned about the composition of the family and the function of its encoded cell-surface glycoproteins. Solution of the structure of the Als adhesive domain provides the opportunity to evaluate the molecular basis for protein function. This review article is formatted as a series of fundamental questions and explores the diversity of the Als proteins, as well as their role in ligand binding, aggregative effects, and attachment to abiotic surfaces. Interaction of Als proteins with each other, their functional equivalence, and the effects of protein abundance on phenotypic conclusions are also examined. Structural features of Als proteins that may facilitate invasive function are considered. Conclusions that are firmly supported by the literature are presented while highlighting areas that require additional investigation to reveal basic features of the Als proteins, their relatedness to each other, and their roles in C. albicans biology. PMID:27014205

Tuning the Protein Corona of Hydrogel Nanoparticles: The Synthesis of Abiotic Protein and Peptide Affinity Reagents.

PubMed

O'Brien, Jeffrey; Shea, Kenneth J

2016-06-21

Nanomaterials, when introduced into a complex, protein-rich environment, rapidly acquire a protein corona. The type and amount of proteins that constitute the corona depend significantly on the synthetic identity of the nanomaterial. For example, hydrogel nanoparticles (NPs) such as poly(N-isopropylacrylamide) (NIPAm) have little affinity for plasma proteins; in contrast, carboxylated poly(styrene) NPs acquire a dense protein corona. This range of protein adsorption suggests that the protein corona might be "tuned" by controlling the chemical composition of the NP. In this Account, we demonstrate that small libraries of synthetic polymer NPs incorporating a diverse pool of functional monomers can be screened for candidates with high affinity and selectivity to targeted biomacromolecules. Through directed synthetic evolution of NP compositions, one can tailor the protein corona to create synthetic organic hydrogel polymer NPs with high affinity and specificity to peptide toxins, enzymes, and other functional proteins, as well as to specific domains of large proteins. In addition, many NIPAm NPs undergo a change in morphology as a function of temperature. This transformation often correlates with a significant change in NP-biomacromolecule affinity, resulting in a temperature-dependent protein corona. This temperature dependence has been used to develop NP hydrogels with autonomous affinity switching for the protection of proteins from thermal stress and as a method of biomacromolecule purification through a selective thermally induced catch and release. In addition to temperature, changes in pH or buffer can also alter a NP protein corona composition, a property that has been exploited for protein purification. Finally, synthetic polymer nanoparticles with low nanomolar affinity for a peptide toxin were shown to capture and neutralize the toxin in the bloodstream of living mice. While the development of synthetic polymer alternatives to protein affinity reagents is in its early stages, these recent successes using only small libraries of functional monomers are most encouraging. It is likely that by expanding the chemical diversity of functional hydrogels and other polymers, a much broader range of NP-biomacromolecule affinity pairs will result. Since these robust, nontoxic polymers are readily synthesized in the chemistry laboratory, we believe the results presented in this Account offer a promising future for the development of low cost alternatives to more traditional protein affinity reagents such as antibodies.

Shigeru Tsuiki: a pioneer in the research fields of complex carbohydrates and protein phosphatases.

PubMed

Miyagi, Taeko; Kikuchi, Kunimi; Tamura, Shinri

2011-11-01

Dr Tsuiki made three major contributions during his illustrious career as a biochemist. First, he developed the procedure for mucin isolation from bovine submaxillary glands. His work became the basis for mucin biochemistry. Second, he identified four distinct molecular species of mammalian sialidase. Subsequent studies based on his work led to the discovery that sialidase plays a unique role as an intracellular signalling factor involved in the regulation of a variety of cellular functions. Finally, he established the molecular basis for the diversity of mammalian protein phosphatases through protein purification and molecular cloning. His work prompted the functional studies of protein phosphatases.

Functional microdomains in bacterial membranes.

PubMed

López, Daniel; Kolter, Roberto

2010-09-01

The membranes of eukaryotic cells harbor microdomains known as lipid rafts that contain a variety of signaling and transport proteins. Here we show that bacterial membranes contain microdomains functionally similar to those of eukaryotic cells. These membrane microdomains from diverse bacteria harbor homologs of Flotillin-1, a eukaryotic protein found exclusively in lipid rafts, along with proteins involved in signaling and transport. Inhibition of lipid raft formation through the action of zaragozic acid--a known inhibitor of squalene synthases--impaired biofilm formation and protein secretion but not cell viability. The orchestration of physiological processes in microdomains may be a more widespread feature of membranes than previously appreciated.

Playing RNase P evolution: swapping the RNA catalyst for a protein reveals functional uniformity of highly divergent enzyme forms.

PubMed

Weber, Christoph; Hartig, Andreas; Hartmann, Roland K; Rossmanith, Walter

2014-08-01

The RNase P family is a diverse group of endonucleases responsible for the removal of 5' extensions from tRNA precursors. The diversity of enzyme forms finds its extremes in the eukaryal nucleus where RNA-based catalysis by complex ribonucleoproteins in some organisms contrasts with single-polypeptide enzymes in others. Such structural contrast suggests associated functional differences, and the complexity of the ribonucleoprotein was indeed proposed to broaden the enzyme's functionality beyond tRNA processing. To explore functional overlap and differences between most divergent forms of RNase P, we replaced the nuclear RNase P of Saccharomyces cerevisiae, a 10-subunit ribonucleoprotein, with Arabidopsis thaliana PRORP3, a single monomeric protein. Surprisingly, the RNase P-swapped yeast strains were viable, displayed essentially unimpaired growth under a wide variety of conditions, and, in a certain genetic background, their fitness even slightly exceeded that of the wild type. The molecular analysis of the RNase P-swapped strains showed a minor disturbance in tRNA metabolism, but did not point to any RNase P substrates or functions beyond that. Altogether, these results indicate the full functional exchangeability of the highly dissimilar enzymes. Our study thereby establishes the RNase P family, with its combination of structural diversity and functional uniformity, as an extreme case of convergent evolution. It moreover suggests that the apparently gratuitous complexity of some RNase P forms is the result of constructive neutral evolution rather than reflecting increased functional versatility.

STRIPAK complexes: structure, biological function, and involvement in human diseases.

PubMed

Hwang, Juyeon; Pallas, David C

2014-02-01

The mammalian striatin family consists of three proteins, striatin, S/G2 nuclear autoantigen, and zinedin. Striatin family members have no intrinsic catalytic activity, but rather function as scaffolding proteins. Remarkably, they organize multiple diverse, large signaling complexes that participate in a variety of cellular processes. Moreover, they appear to be regulatory/targeting subunits for the major eukaryotic serine/threonine protein phosphatase 2A. In addition, striatin family members associate with germinal center kinase III kinases as well as other novel components, earning these assemblies the name striatin-interacting phosphatase and kinase (STRIPAK) complexes. Recently, there has been a great increase in functional and mechanistic studies aimed at identifying and understanding the roles of STRIPAK and STRIPAK-like complexes in cellular processes of multiple organisms. These studies have identified novel STRIPAK and STRIPAK-like complexes and have explored their roles in specific signaling pathways. Together, the results of these studies have sparked increased interest in striatin family complexes because they have revealed roles in signaling, cell cycle control, apoptosis, vesicular trafficking, Golgi assembly, cell polarity, cell migration, neural and vascular development, and cardiac function. Moreover, STRIPAK complexes have been connected to clinical conditions, including cardiac disease, diabetes, autism, and cerebral cavernous malformation. In this review, we discuss the expression, localization, and protein domain structure of striatin family members. Then we consider the diverse complexes these proteins and their homologs form in various organisms, emphasizing what is known regarding function and regulation. Finally, we explore possible roles of striatin family complexes in disease, especially cerebral cavernous malformation. Copyright © 2013 Elsevier Ltd. All rights reserved.

Polycomb Group (PcG) Proteins and Human Cancers: Multifaceted Functions and Therapeutic Implications

PubMed Central

Wang, Wei; Qin, Jiang-Jiang; Voruganti, Sukesh; Nag, Subhasree; Zhou, Jianwei; Zhang, Ruiwen

2016-01-01

Polycomb group (PcG) proteins are transcriptional repressors that regulate several crucial developmental and physiological processes in the cell. More recently, they have been found to play important roles in human carcinogenesis and cancer development and progression. The deregulation and dysfunction of PcG proteins often lead to blocking or inappropriate activation of developmental pathways, enhancing cellular proliferation, inhibiting apoptosis, and increasing the cancer stem cell population. Genetic and molecular investigations of PcG proteins have long been focused on their PcG functions. However, PcG proteins have recently been shown to exert non-polycomb functions, contributing to the regulation of diverse cellular functions. We and others have demonstrated that PcG proteins regulate the expression and function of several oncogenes and tumor suppressor genes in a PcG-independent manner, and PcG proteins are associated with the survival of patients with cancer. In this review, we summarize the recent advances in the research on PcG proteins, including both the polycomb-repressive and non-polycomb functions. We specifically focus on the mechanisms by which PcG proteins play roles in cancer initiation, development, and progression. Finally, we discuss the potential value of PcG proteins as molecular biomarkers for the diagnosis and prognosis of cancer, and as molecular targets for cancer therapy. PMID:26227500

Plant lectins: the ties that bind in root symbiosis and plant defense.

PubMed

De Hoff, Peter L; Brill, Laurence M; Hirsch, Ann M

2009-07-01

Lectins are a diverse group of carbohydrate-binding proteins that are found within and associated with organisms from all kingdoms of life. Several different classes of plant lectins serve a diverse array of functions. The most prominent of these include participation in plant defense against predators and pathogens and involvement in symbiotic interactions between host plants and symbiotic microbes, including mycorrhizal fungi and nitrogen-fixing rhizobia. Extensive biological, biochemical, and molecular studies have shed light on the functions of plant lectins, and a plethora of uncharacterized lectin genes are being revealed at the genomic scale, suggesting unexplored and novel diversity in plant lectin structure and function. Integration of the results from these different types of research is beginning to yield a more detailed understanding of the function of lectins in symbiosis, defense, and plant biology in general.

Comprehensive Phylogenetic Analysis Sheds Light on the Diversity and Origin of the MLO Family of Integral Membrane Proteins

PubMed Central

Kusch, Stefan; Pesch, Lina; Panstruga, Ralph

2016-01-01

Mildew resistance Locus O (MLO) proteins are polytopic integral membrane proteins that have long been considered as plant-specific and being primarily involved in plant–powdery mildew interactions. However, research in the past decade has revealed that MLO proteins diverged into a family with several clades whose members are associated with different physiological processes. We provide a largely increased dataset of MLO amino acid sequences, comprising nearly all major land plant lineages. Based on this comprehensive dataset, we defined seven phylogenetic clades and reconstructed the likely evolution of the MLO family in embryophytes. We further identified several MLO peptide motifs that are either conserved in all MLO proteins or confined to one or several clades, supporting the notion that clade-specific diversification of MLO functions is associated with particular sequence motifs. In baker’s yeast, some of these motifs are functionally linked to transmembrane (TM) transport of organic molecules and ions. In addition, we attempted to define the evolutionary origin of the MLO family and found that MLO-like proteins with highly diverse membrane topologies are present in green algae, but also in the distinctly related red algae (Rhodophyta), Amoebozoa, and Chromalveolata. Finally, we discovered several instances of putative fusion events between MLO proteins and different kinds of proteins. Such Rosetta stone-type hybrid proteins might be instructive for future analysis of potential MLO functions. Our findings suggest that MLO is an ancient protein that possibly evolved in unicellular photosynthetic eukaryotes, and consolidated in land plants with a conserved topology, comprising seven TM domains and an intrinsically unstructured C-terminus. PMID:26893454

Direct protein interaction underlies gene-for-gene specificity and coevolution of the flax resistance genes and flax rust avirulence genes

PubMed Central

Dodds, Peter N.; Lawrence, Gregory J.; Catanzariti, Ann-Maree; Teh, Trazel; Wang, Ching-I. A.; Ayliffe, Michael A.; Kobe, Bostjan; Ellis, Jeffrey G.

2006-01-01

Plant resistance proteins (R proteins) recognize corresponding pathogen avirulence (Avr) proteins either indirectly through detection of changes in their host protein targets or through direct R–Avr protein interaction. Although indirect recognition imposes selection against Avr effector function, pathogen effector molecules recognized through direct interaction may overcome resistance through sequence diversification rather than loss of function. Here we show that the flax rust fungus AvrL567 genes, whose products are recognized by the L5, L6, and L7 R proteins of flax, are highly diverse, with 12 sequence variants identified from six rust strains. Seven AvrL567 variants derived from Avr alleles induce necrotic responses when expressed in flax plants containing corresponding resistance genes (R genes), whereas five variants from avr alleles do not. Differences in recognition specificity between AvrL567 variants and evidence for diversifying selection acting on these genes suggest they have been involved in a gene-specific arms race with the corresponding flax R genes. Yeast two-hybrid assays indicate that recognition is based on direct R–Avr protein interaction and recapitulate the interaction specificity observed in planta. Biochemical analysis of Escherichia coli-produced AvrL567 proteins shows that variants that escape recognition nevertheless maintain a conserved structure and stability, suggesting that the amino acid sequence differences directly affect the R–Avr protein interaction. We suggest that direct recognition associated with high genetic diversity at corresponding R and Avr gene loci represents an alternative outcome of plant–pathogen coevolution to indirect recognition associated with simple balanced polymorphisms for functional and nonfunctional R and Avr genes. PMID:16731621

Functions of tissue-resident eosinophils.

PubMed

Weller, Peter F; Spencer, Lisa A

2017-12-01

Eosinophils are a prominent cell type in particular host responses such as the response to helminth infection and allergic disease. Their effector functions have been attributed to their capacity to release cationic proteins stored in cytoplasmic granules by degranulation. However, eosinophils are now being recognized for more varied functions in previously underappreciated diverse tissue sites, based on the ability of eosinophils to release cytokines (often preformed) that mediate a broad range of activities into the local environment. In this Review, we consider evolving insights into the tissue distribution of eosinophils and their functional immunobiology, which enable eosinophils to secrete in a selective manner cytokines and other mediators that have diverse, 'non-effector' functions in health and disease.

INK4 proteins, a family of mammalian CDK inhibitors with novel biological functions.

PubMed

Cánepa, Eduardo T; Scassa, María E; Ceruti, Julieta M; Marazita, Mariela C; Carcagno, Abel L; Sirkin, Pablo F; Ogara, María F

2007-07-01

The cyclin D-Cdk4-6/INK4/Rb/E2F pathway plays a key role in controlling cell growth by integrating multiple mitogenic and antimitogenic stimuli. The members of INK4 family, comprising p16(INK4a), p15(INK4b), p18(INK4c), and p19(INK4d), block the progression of the cell cycle by binding to either Cdk4 or Cdk6 and inhibiting the action of cyclin D. These INK4 proteins share a similar structure dominated by several ankyrin repeats. Although they appear to be structurally redundant and equally potent as inhibitors, the INK4 family members are differentially expressed during mouse development. The striking diversity in the pattern of expression of INK4 genes suggested that this family of cell cycle inhibitors might have cell lineage-specific or tissue-specific functions. The INK4 proteins are commonly lost or inactivated by mutations in diverse types of cancer, and they represent established or candidate tumor suppressors. Apart from their capacity to arrest cells in the G1-phase of the cell cycle they have been shown to participate in an increasing number of cellular processes. Given their emerging roles in fundamental physiological as well as pathological processes, it is interesting to explore the diverse roles for the individual INK4 family members in different functions other than cell cycle regulation. Extensive studies, over the past few years, uncover the involvement of INK4 proteins in senescence, apoptosis, DNA repair, and multistep oncogenesis. We will focus the discussion here on these unexpected issues.

The targeting of plant cellular systems by injected type III effector proteins.

PubMed

Lewis, Jennifer D; Guttman, David S; Desveaux, Darrell

2009-12-01

The battle between phytopathogenic bacteria and their plant hosts has revealed a diverse suite of strategies and mechanisms employed by the pathogen or the host to gain the higher ground. Pathogens continually evolve tactics to acquire host resources and dampen host defences. Hosts must evolve surveillance and defence systems that are sensitive enough to rapidly respond to a diverse range of pathogens, while reducing costly and damaging inappropriate misexpression. The primary virulence mechanism employed by many bacteria is the type III secretion system, which secretes and translocates effector proteins directly into the cells of their plant hosts. Effectors have diverse enzymatic functions and can target specific components of plant systems. While these effectors should favour bacterial fitness, the host may be able to thwart infection by recognizing the activity or presence of these foreign molecules and initiating retaliatory immune measures. We review the diverse host cellular systems exploited by bacterial effectors, with particular focus on plant proteins directly targeted by effectors. Effector-host interactions reveal different stages of the battle between pathogen and host, as well as the diverse molecular strategies employed by bacterial pathogens to hijack eukaryotic cellular systems.

Electric-field-stimulated protein mechanics

PubMed Central

Hekstra, Doeke R.; White, K. Ian; Socolich, Michael A.; Henning, Robert W.; Šrajer, Vukica; Ranganathan, Rama

2017-01-01

The internal mechanics of proteins—the coordinated motions of amino acids and the pattern of forces constraining these motions—connects protein structure to function. Here we describe a new method combining the application of strong electric field pulses to protein crystals with time-resolved X-ray crystallography to observe conformational changes in spatial and temporal detail. Using a human PDZ domain (LNX2PDZ2) as a model system, we show that protein crystals tolerate electric field pulses strong enough to drive concerted motions on the sub-microsecond timescale. The induced motions are subtle, involve diverse physical mechanisms, and occur throughout the protein structure. The global pattern of electric-field-induced motions is consistent with both local and allosteric conformational changes naturally induced by ligand binding, including at conserved functional sites in the PDZ domain family. This work lays the foundation for comprehensive experimental study of the mechanical basis of protein function. PMID:27926732

Evolution of an ancient protein function involved in organized multicellularity in animals.

PubMed

Anderson, Douglas P; Whitney, Dustin S; Hanson-Smith, Victor; Woznica, Arielle; Campodonico-Burnett, William; Volkman, Brian F; King, Nicole; Thornton, Joseph W; Prehoda, Kenneth E

2016-01-07

To form and maintain organized tissues, multicellular organisms orient their mitotic spindles relative to neighboring cells. A molecular complex scaffolded by the GK protein-interaction domain (GKPID) mediates spindle orientation in diverse animal taxa by linking microtubule motor proteins to a marker protein on the cell cortex localized by external cues. Here we illuminate how this complex evolved and commandeered control of spindle orientation from a more ancient mechanism. The complex was assembled through a series of molecular exploitation events, one of which - the evolution of GKPID's capacity to bind the cortical marker protein - can be recapitulated by reintroducing a single historical substitution into the reconstructed ancestral GKPID. This change revealed and repurposed an ancient molecular surface that previously had a radically different function. We show how the physical simplicity of this binding interface enabled the evolution of a new protein function now essential to the biological complexity of many animals.

Characterization and expression analysis of genes involved in SUMOylation during embryogenesis in rainbow trout (Oncorhynchus mykiss)

USDA-ARS?s Scientific Manuscript database

SUMOylation is the post-translational modification of proteins by the addition of the small ubiquitin-like modifier (SUMO), which plays an important role in various cellular processes. It has been reported that SUMO and its related proteins are important in diverse reproductive functions such as ovu...

LRAT-specific domain facilitates vitamin A metabolism by domain swapping in HRASLS3

DOE PAGES

Golczak, Marcin; Sears, Avery E.; Kiser, Philip D.; ...

2014-11-10

Cellular uptake of vitamin A, production of visual chromophore and triglyceride homeostasis in adipocytes depend on two representatives of the vertebrate N1pC/P60 protein family, lecithin:retinol acyltransferase (LRAT) and HRAS-like tumor suppressor 3 (HRASLS3). Both proteins function as lipid-metabolizing enzymes but differ in their substrate preferences and dominant catalytic activity. The mechanism of this catalytic diversity is not understood. In this paper, by using a gain-of-function approach, we identified a specific sequence responsible for the substrate specificity of N1pC/P60 proteins. A 2.2-Å crystal structure of the HRASLS3-LRAT chimeric enzyme in a thioester catalytic intermediate state revealed a major structural rearrangement accompaniedmore » by three-dimensional domain swapping dimerization not observed in native HRASLS proteins. Structural changes affecting the active site environment contributed to slower hydrolysis of the catalytic intermediate, supporting efficient acyl transfer. Finally, these findings reveal structural adaptation that facilitates selective catalysis and mechanism responsible for diverse substrate specificity within the LRAT-like enzyme family.« less

Tinkering with Translation: Protein Synthesis in Virus-Infected Cells

PubMed Central

Walsh, Derek; Mathews, Michael B.; Mohr, Ian

2013-01-01

Viruses are obligate intracellular parasites, and their replication requires host cell functions. Although the size, composition, complexity, and functions encoded by their genomes are remarkably diverse, all viruses rely absolutely on the protein synthesis machinery of their host cells. Lacking their own translational apparatus, they must recruit cellular ribosomes in order to translate viral mRNAs and produce the protein products required for their replication. In addition, there are other constraints on viral protein production. Crucially, host innate defenses and stress responses capable of inactivating the translation machinery must be effectively neutralized. Furthermore, the limited coding capacity of the viral genome needs to be used optimally. These demands have resulted in complex interactions between virus and host that exploit ostensibly virus-specific mechanisms and, at the same time, illuminate the functioning of the cellular protein synthesis apparatus. PMID:23209131

Nutritional composition of honey bee food stores vary with floral composition.

PubMed

Donkersley, Philip; Rhodes, Glenn; Pickup, Roger W; Jones, Kevin C; Power, Eileen F; Wright, Geraldine A; Wilson, Kenneth

2017-12-01

Sufficiently diverse and abundant resources are essential for generalist consumers, and form an important part of a suite of conservation strategies for pollinators. Honey bees are generalist foragers and are dependent on diverse forage to adequately meet their nutritional needs. Through analysis of stored pollen (bee bread) samples obtained from 26 honey bee (Apis mellifera L.) hives across NW-England, we quantified bee bread nutritional content and the plant species that produced these stores from pollen. Protein was the most abundant nutrient by mass (63%), followed by carbohydrates (26%). Protein and lipid content (but not carbohydrate) contributed significantly to ordinations of floral diversity, linking dietary quality with forage composition. DNA sequencing of the ITS2 region of the nuclear ribosomal DNA gene identified pollen from 89 distinct plant genera, with each bee bread sample containing between 6 and 35 pollen types. Dominant genera included dandelion (Taraxacum), which was positively correlated with bee bread protein content, and cherry (Prunus), which was negatively correlated with the amount of protein. In addition, proportions of amino acids (e.g. histidine and valine) varied as a function of floral species composition. These results also quantify the effects of individual plant genera on the nutrition of honey bees. We conclude that pollens of different plants act synergistically to influence host nutrition; the pollen diversity of bee bread is linked to its nutrient content. Diverse environments compensate for the loss of individual forage plants, and diversity loss may, therefore, destabilize consumer communities due to restricted access to alternative resources.

A Robust and Versatile Method of Combinatorial Chemical Synthesis of Gene Libraries via Hierarchical Assembly of Partially Randomized Modules

PubMed Central

Popova, Blagovesta; Schubert, Steffen; Bulla, Ingo; Buchwald, Daniela; Kramer, Wilfried

2015-01-01

A major challenge in gene library generation is to guarantee a large functional size and diversity that significantly increases the chances of selecting different functional protein variants. The use of trinucleotides mixtures for controlled randomization results in superior library diversity and offers the ability to specify the type and distribution of the amino acids at each position. Here we describe the generation of a high diversity gene library using tHisF of the hyperthermophile Thermotoga maritima as a scaffold. Combining various rational criteria with contingency, we targeted 26 selected codons of the thisF gene sequence for randomization at a controlled level. We have developed a novel method of creating full-length gene libraries by combinatorial assembly of smaller sub-libraries. Full-length libraries of high diversity can easily be assembled on demand from smaller and much less diverse sub-libraries, which circumvent the notoriously troublesome long-term archivation and repeated proliferation of high diversity ensembles of phages or plasmids. We developed a generally applicable software tool for sequence analysis of mutated gene sequences that provides efficient assistance for analysis of library diversity. Finally, practical utility of the library was demonstrated in principle by assessment of the conformational stability of library members and isolating protein variants with HisF activity from it. Our approach integrates a number of features of nucleic acids synthetic chemistry, biochemistry and molecular genetics to a coherent, flexible and robust method of combinatorial gene synthesis. PMID:26355961

A Robust and Versatile Method of Combinatorial Chemical Synthesis of Gene Libraries via Hierarchical Assembly of Partially Randomized Modules.

PubMed

Popova, Blagovesta; Schubert, Steffen; Bulla, Ingo; Buchwald, Daniela; Kramer, Wilfried

2015-01-01

A major challenge in gene library generation is to guarantee a large functional size and diversity that significantly increases the chances of selecting different functional protein variants. The use of trinucleotides mixtures for controlled randomization results in superior library diversity and offers the ability to specify the type and distribution of the amino acids at each position. Here we describe the generation of a high diversity gene library using tHisF of the hyperthermophile Thermotoga maritima as a scaffold. Combining various rational criteria with contingency, we targeted 26 selected codons of the thisF gene sequence for randomization at a controlled level. We have developed a novel method of creating full-length gene libraries by combinatorial assembly of smaller sub-libraries. Full-length libraries of high diversity can easily be assembled on demand from smaller and much less diverse sub-libraries, which circumvent the notoriously troublesome long-term archivation and repeated proliferation of high diversity ensembles of phages or plasmids. We developed a generally applicable software tool for sequence analysis of mutated gene sequences that provides efficient assistance for analysis of library diversity. Finally, practical utility of the library was demonstrated in principle by assessment of the conformational stability of library members and isolating protein variants with HisF activity from it. Our approach integrates a number of features of nucleic acids synthetic chemistry, biochemistry and molecular genetics to a coherent, flexible and robust method of combinatorial gene synthesis.

Deep Illumina-Based Shotgun Sequencing Reveals Dietary Effects on the Structure and Function of the Fecal Microbiome of Growing Kittens

PubMed Central

Deusch, Oliver; O’Flynn, Ciaran; Colyer, Alison; Morris, Penelope; Allaway, David; Jones, Paul G.; Swanson, Kelly S.

2014-01-01

Background Previously, we demonstrated that dietary protein:carbohydrate ratio dramatically affects the fecal microbial taxonomic structure of kittens using targeted 16S gene sequencing. The present study, using the same fecal samples, applied deep Illumina shotgun sequencing to identify the diet-associated functional potential and analyze taxonomic changes of the feline fecal microbiome. Methodology & Principal Findings Fecal samples from kittens fed one of two diets differing in protein and carbohydrate content (high–protein, low–carbohydrate, HPLC; and moderate-protein, moderate-carbohydrate, MPMC) were collected at 8, 12 and 16 weeks of age (n = 6 per group). A total of 345.3 gigabases of sequence were generated from 36 samples, with 99.75% of annotated sequences identified as bacterial. At the genus level, 26% and 39% of reads were annotated for HPLC- and MPMC-fed kittens, with HPLC-fed cats showing greater species richness and microbial diversity. Two phyla, ten families and fifteen genera were responsible for more than 80% of the sequences at each taxonomic level for both diet groups, consistent with the previous taxonomic study. Significantly different abundances between diet groups were observed for 324 genera (56% of all genera identified) demonstrating widespread diet-induced changes in microbial taxonomic structure. Diversity was not affected over time. Functional analysis identified 2,013 putative enzyme function groups were different (p<0.000007) between the two dietary groups and were associated to 194 pathways, which formed five discrete clusters based on average relative abundance. Of those, ten contained more (p<0.022) enzyme functions with significant diet effects than expected by chance. Six pathways were related to amino acid biosynthesis and metabolism linking changes in dietary protein with functional differences of the gut microbiome. Conclusions These data indicate that feline feces-derived microbiomes have large structural and functional differences relating to the dietary protein:carbohydrate ratio and highlight the impact of diet early in life. PMID:25010839

ERK and p38 MAPK-Activated Protein Kinases: a Family of Protein Kinases with Diverse Biological Functions

PubMed Central

Roux, Philippe P.; Blenis, John

2004-01-01

Conserved signaling pathways that activate the mitogen-activated protein kinases (MAPKs) are involved in relaying extracellular stimulations to intracellular responses. The MAPKs coordinately regulate cell proliferation, differentiation, motility, and survival, which are functions also known to be mediated by members of a growing family of MAPK-activated protein kinases (MKs; formerly known as MAPKAP kinases). The MKs are related serine/threonine kinases that respond to mitogenic and stress stimuli through proline-directed phosphorylation and activation of the kinase domain by extracellular signal-regulated kinases 1 and 2 and p38 MAPKs. There are currently 11 vertebrate MKs in five subfamilies based on primary sequence homology: the ribosomal S6 kinases, the mitogen- and stress-activated kinases, the MAPK-interacting kinases, MAPK-activated protein kinases 2 and 3, and MK5. In the last 5 years, several MK substrates have been identified, which has helped tremendously to identify the biological role of the members of this family. Together with data from the study of MK-knockout mice, the identities of the MK substrates indicate that they play important roles in diverse biological processes, including mRNA translation, cell proliferation and survival, and the nuclear genomic response to mitogens and cellular stresses. In this article, we review the existing data on the MKs and discuss their physiological functions based on recent discoveries. PMID:15187187

Bypassing Evolutionary Roadblocks: Phenotypic Diversity in Isogenic Population Bridges Tradeoff in Evolution of a New Function

NASA Astrophysics Data System (ADS)

Petrie, K. L.; Meyer, J. R.

2017-07-01

A novel mechanism of innovation bridges fitness valleys by violating the one gene-one phenotype dogma. Protein products of a single gene partition into populations, some of which carry out a new function and some the old, avoiding tradeoffs.

Network-based function prediction and interactomics: the case for metabolic enzymes.

PubMed

Janga, S C; Díaz-Mejía, J Javier; Moreno-Hagelsieb, G

2011-01-01

As sequencing technologies increase in power, determining the functions of unknown proteins encoded by the DNA sequences so produced becomes a major challenge. Functional annotation is commonly done on the basis of amino-acid sequence similarity alone. Long after sequence similarity becomes undetectable by pair-wise comparison, profile-based identification of homologs can often succeed due to the conservation of position-specific patterns, important for a protein's three dimensional folding and function. Nevertheless, prediction of protein function from homology-driven approaches is not without problems. Homologous proteins might evolve different functions and the power of homology detection has already started to reach its maximum. Computational methods for inferring protein function, which exploit the context of a protein in cellular networks, have come to be built on top of homology-based approaches. These network-based functional inference techniques provide both a first hand hint into a proteins' functional role and offer complementary insights to traditional methods for understanding the function of uncharacterized proteins. Most recent network-based approaches aim to integrate diverse kinds of functional interactions to boost both coverage and confidence level. These techniques not only promise to solve the moonlighting aspect of proteins by annotating proteins with multiple functions, but also increase our understanding on the interplay between different functional classes in a cell. In this article we review the state of the art in network-based function prediction and describe some of the underlying difficulties and successes. Given the volume of high-throughput data that is being reported the time is ripe to employ these network-based approaches, which can be used to unravel the functions of the uncharacterized proteins accumulating in the genomic databases. © 2010 Elsevier Inc. All rights reserved.

Endozoicomonas genomes reveal functional adaptation and plasticity in bacterial strains symbiotically associated with diverse marine hosts

PubMed Central

Neave, Matthew J.; Michell, Craig T.; Apprill, Amy; Voolstra, Christian R.

2017-01-01

Endozoicomonas bacteria are globally distributed and often abundantly associated with diverse marine hosts including reef-building corals, yet their function remains unknown. In this study we generated novel Endozoicomonas genomes from single cells and metagenomes obtained directly from the corals Stylophora pistillata, Pocillopora verrucosa, and Acropora humilis. We then compared these culture-independent genomes to existing genomes of bacterial isolates acquired from a sponge, sea slug, and coral to examine the functional landscape of this enigmatic genus. Sequencing and analysis of single cells and metagenomes resulted in four novel genomes with 60–76% and 81–90% genome completeness, respectively. These data also confirmed that Endozoicomonas genomes are large and are not streamlined for an obligate endosymbiotic lifestyle, implying that they have free-living stages. All genomes show an enrichment of genes associated with carbon sugar transport and utilization and protein secretion, potentially indicating that Endozoicomonas contribute to the cycling of carbohydrates and the provision of proteins to their respective hosts. Importantly, besides these commonalities, the genomes showed evidence for differential functional specificity and diversification, including genes for the production of amino acids. Given this metabolic diversity of Endozoicomonas we propose that different genotypes play disparate roles and have diversified in concert with their hosts. PMID:28094347

Characterizing Conformational Dynamics of Proteins Using Evolutionary Couplings.

PubMed

Feng, Jiangyan; Shukla, Diwakar

2018-01-25

Understanding of protein conformational dynamics is essential for elucidating molecular origins of protein structure-function relationship. Traditionally, reaction coordinates, i.e., some functions of protein atom positions and velocities have been used to interpret the complex dynamics of proteins obtained from experimental and computational approaches such as molecular dynamics simulations. However, it is nontrivial to identify the reaction coordinates a priori even for small proteins. Here, we evaluate the power of evolutionary couplings (ECs) to capture protein dynamics by exploring their use as reaction coordinates, which can efficiently guide the sampling of a conformational free energy landscape. We have analyzed 10 diverse proteins and shown that a few ECs are sufficient to characterize complex conformational dynamics of proteins involved in folding and conformational change processes. With the rapid strides in sequencing technology, we expect that ECs could help identify reaction coordinates a priori and enhance the sampling of the slow dynamical process associated with protein folding and conformational change.

Implementing a rational and consistent nomenclature for serine/arginine-rich protein splicing factors (SR proteins) in plants.

PubMed

Barta, Andrea; Kalyna, Maria; Reddy, Anireddy S N

2010-09-01

Growing interest in alternative splicing in plants and the extensive sequencing of new plant genomes necessitate more precise definition and classification of genes coding for splicing factors. SR proteins are a family of RNA binding proteins, which function as essential factors for constitutive and alternative splicing. We propose a unified nomenclature for plant SR proteins, taking into account the newly revised nomenclature of the mammalian SR proteins and a number of plant-specific properties of the plant proteins. We identify six subfamilies of SR proteins in Arabidopsis thaliana and rice (Oryza sativa), three of which are plant specific. The proposed subdivision of plant SR proteins into different subfamilies will allow grouping of paralogous proteins and simple assignment of newly discovered SR orthologs from other plant species and will promote functional comparisons in diverse plant species.

Rapid functional diversification in the structurally conserved ELAV family of neuronal RNA binding proteins

PubMed Central

Samson, Marie-Laure

2008-01-01

Background The Drosophila gene embryonic lethal abnormal visual system (elav) is the prototype of a gene family present in all metazoans. Its members encode structurally conserved neuronal proteins with three RNA Recognition Motifs (RRM) but they paradoxically act at diverse levels of post-transcriptional regulation. In an attempt to understand the history of this family, we searched for orthologs in eleven completely sequenced genomes, including those of humans, D. melanogaster and C. elegans, for which cDNAs are available. Results We analyzed 23 orthologs/paralogs of elav, and found evidence of gain/loss of gene copy number. For one set of genes, including elav itself, the coding sequences are free of introns and their products most resemble ELAV. The remaining genes show remarkable conservation of their exon organization, and their products most resemble FNE and RBP9, proteins encoded by the two elav paralogs of Drosophila. Remarkably, three of the conserved exon junctions are both close to structural elements, involved respectively in protein-RNA interactions and in the regulation of sub-cellular localization, and in the vicinity of diverse sequence variations. Conclusion The data indicate that the essential elav gene of Drosophila is newly emerged, restricted to dipterans and of retrotransposed origin. We propose that the conserved exon junctions constitute potential sites for sequence/function modifications, and that RRM binding proteins, whose function relies upon plastic RNA-protein interactions, may have played an important role in brain evolution. PMID:18715504

Exploring Fold Space Preferences of New-born and Ancient Protein Superfamilies

PubMed Central

Edwards, Hannah; Abeln, Sanne; Deane, Charlotte M.

2013-01-01

The evolution of proteins is one of the fundamental processes that has delivered the diversity and complexity of life we see around ourselves today. While we tend to define protein evolution in terms of sequence level mutations, insertions and deletions, it is hard to translate these processes to a more complete picture incorporating a polypeptide's structure and function. By considering how protein structures change over time we can gain an entirely new appreciation of their long-term evolutionary dynamics. In this work we seek to identify how populations of proteins at different stages of evolution explore their possible structure space. We use an annotation of superfamily age to this space and explore the relationship between these ages and a diverse set of properties pertaining to a superfamily's sequence, structure and function. We note several marked differences between the populations of newly evolved and ancient structures, such as in their length distributions, secondary structure content and tertiary packing arrangements. In particular, many of these differences suggest a less elaborate structure for newly evolved superfamilies when compared with their ancient counterparts. We show that the structural preferences we report are not a residual effect of a more fundamental relationship with function. Furthermore, we demonstrate the robustness of our results, using significant variation in the algorithm used to estimate the ages. We present these age estimates as a useful tool to analyse protein populations. In particularly, we apply this in a comparison of domains containing greek key or jelly roll motifs. PMID:24244135

Activation and Function of the MAPKs and Their Substrates, the MAPK-Activated Protein Kinases

PubMed Central

Cargnello, Marie; Roux, Philippe P.

2011-01-01

Summary: The mitogen-activated protein kinases (MAPKs) regulate diverse cellular programs by relaying extracellular signals to intracellular responses. In mammals, there are more than a dozen MAPK enzymes that coordinately regulate cell proliferation, differentiation, motility, and survival. The best known are the conventional MAPKs, which include the extracellular signal-regulated kinases 1 and 2 (ERK1/2), c-Jun amino-terminal kinases 1 to 3 (JNK1 to -3), p38 (α, β, γ, and δ), and ERK5 families. There are additional, atypical MAPK enzymes, including ERK3/4, ERK7/8, and Nemo-like kinase (NLK), which have distinct regulation and functions. Together, the MAPKs regulate a large number of substrates, including members of a family of protein Ser/Thr kinases termed MAPK-activated protein kinases (MAPKAPKs). The MAPKAPKs are related enzymes that respond to extracellular stimulation through direct MAPK-dependent activation loop phosphorylation and kinase activation. There are five MAPKAPK subfamilies: the p90 ribosomal S6 kinase (RSK), the mitogen- and stress-activated kinase (MSK), the MAPK-interacting kinase (MNK), the MAPK-activated protein kinase 2/3 (MK2/3), and MK5 (also known as p38-regulated/activated protein kinase [PRAK]). These enzymes have diverse biological functions, including regulation of nucleosome and gene expression, mRNA stability and translation, and cell proliferation and survival. Here we review the mechanisms of MAPKAPK activation by the different MAPKs and discuss their physiological roles based on established substrates and recent discoveries. PMID:21372320

GOLabeler: Improving Sequence-based Large-scale Protein Function Prediction by Learning to Rank.

PubMed

You, Ronghui; Zhang, Zihan; Xiong, Yi; Sun, Fengzhu; Mamitsuka, Hiroshi; Zhu, Shanfeng

2018-03-07

Gene Ontology (GO) has been widely used to annotate functions of proteins and understand their biological roles. Currently only <1% of more than 70 million proteins in UniProtKB have experimental GO annotations, implying the strong necessity of automated function prediction (AFP) of proteins, where AFP is a hard multilabel classification problem due to one protein with a diverse number of GO terms. Most of these proteins have only sequences as input information, indicating the importance of sequence-based AFP (SAFP: sequences are the only input). Furthermore homology-based SAFP tools are competitive in AFP competitions, while they do not necessarily work well for so-called difficult proteins, which have <60% sequence identity to proteins with annotations already. Thus the vital and challenging problem now is how to develop a method for SAFP, particularly for difficult proteins. The key of this method is to extract not only homology information but also diverse, deep- rooted information/evidence from sequence inputs and integrate them into a predictor in a both effective and efficient manner. We propose GOLabeler, which integrates five component classifiers, trained from different features, including GO term frequency, sequence alignment, amino acid trigram, domains and motifs, and biophysical properties, etc., in the framework of learning to rank (LTR), a paradigm of machine learning, especially powerful for multilabel classification. The empirical results obtained by examining GOLabeler extensively and thoroughly by using large-scale datasets revealed numerous favorable aspects of GOLabeler, including significant performance advantage over state-of-the-art AFP methods. http://datamining-iip.fudan.edu.cn/golabeler. zhusf@fudan.edu.cn. Supplementary data are available at Bioinformatics online.

A Val85Met Mutation in Melanocortin-1 Receptor Is Associated with Reductions in Eumelanic Pigmentation and Cell Surface Expression in Domestic Rock Pigeons (Columba livia)

PubMed Central

Guernsey, Michael W.; Ritscher, Lars; Miller, Matthew A.; Smith, Daniel A.; Schöneberg, Torsten; Shapiro, Michael D.

2013-01-01

Variation in the melanocortin-1 receptor (Mc1r) is associated with pigmentation diversity in wild and domesticated populations of vertebrates, including several species of birds. Among domestic bird species, pigmentation variation in the rock pigeon ( Columba livia ) is particularly diverse. To determine the potential contribution of Mc1r variants to pigment diversity in pigeons, we sequenced Mc1r in a wide range of pigeon breeds and identified several single nucleotide polymorphisms, including a variant that codes for an amino acid substitution (Val85Met). In contrast to the association between Val85Met and eumelanism in other avian species, this change was associated with pheomelanism in pigeons. In vitro cAMP accumulation and protein expression assays revealed that Val85Met leads to decreased receptor function and reduced cell surface expression of the mutant protein. The reduced in vitro function is consistent with the observed association with reduced eumelanic pigmentation. Comparative genetic and cellular studies provide important insights about the range of mechanisms underlying diversity among vertebrates, including different phenotypic associations with similar mutations in different species. PMID:23977400

Ligand binding by repeat proteins: natural and designed

PubMed Central

Grove, Tijana Z; Cortajarena, Aitziber L; Regan, Lynne

2012-01-01

Repeat proteins contain tandem arrays of small structural motifs. As a consequence of this architecture, they adopt non-globular, extended structures that present large, highly specific surfaces for ligand binding. Here we discuss recent advances toward understanding the functional role of this unique modular architecture. We showcase specific examples of natural repeat proteins interacting with diverse ligands and also present examples of designed repeat protein–ligand interactions. PMID:18602006

Beyond the Eye: Molecular Evolution of Extraocular Photoreception.

PubMed

Porter, Megan L

2016-11-01

The molecular mechanisms used by biological systems to detect light are diverse, with at least 10 classes of photosensor proteins and additional photosensitive domains characterized. At least six of these protein classes-Type I microbial opsins, Type II animal opsins, cryptochromes, gustatory-related receptors (GRRs), transient receptor potential A1 ion channels, and euglenoid photoactivated adenylyl cylases-can be considered as playing a role in extraocular systems (e.g., expressed outside of the eye in organisms with a visual system). These six classes of extraocular photosensor proteins consist of four broad groups: (1) seven transmembrane proteins, (2) cryptochromes, (3) ion channels, and (4) adenylyl cyclases. The light-driven functions of these extraocular photoreceptors are diverse, ranging from circadian entrainment to phototactic behavior. There are surprising similarities in structural motifs, with at least three independent families-the GRRs and Types I and II opsins-evolving a seven transmembrane helical tertiary structure for light sensing. When considering all of the photosensitive proteins, particularly those in microbial lineages, an image of evolutionary flexibility is emerging, with examples of fusion proteins from multiple types of photosensors and photosensitive domains shared among diverse arrays of proteins. In general, large questions remain for most of these photosensor proteins about exactly how the protein evolved light sensitivity, how light interacts with the protein, and how the photosensitive protein is transducing the signal. © The Author 2016. Published by Oxford University Press on behalf of the Society for Integrative and Comparative Biology. All rights reserved. For permissions please email: journals.permissions@oup.com.

Alternative cytoskeletal landscapes: cytoskeletal novelty and evolution in basal excavate protists

PubMed Central

Dawson, Scott C.; Paredez, Alexander R.

2016-01-01

Microbial eukaryotes encompass the majority of eukaryotic evolutionary and cytoskeletal diversity. The cytoskeletal complexity observed in multicellular organisms appears to be an expansion of components present in genomes of diverse microbial eukaryotes such as the basal lineage of flagellates, the Excavata. Excavate protists have complex and diverse cytoskeletal architectures and life cycles – essentially alternative cytoskeletal “landscapes” – yet still possess conserved microtubule- and actin-associated proteins. Comparative genomic analyses have revealed that a subset of excavates, however, lack many canonical actin-binding proteins central to actin cytoskeleton function in other eukaryotes. Overall, excavates possess numerous uncharacterized and “hypothetical” genes, and may represent an undiscovered reservoir of novel cytoskeletal genes and cytoskeletal mechanisms. The continued development of molecular genetic tools in these complex microbial eukaryotes will undoubtedly contribute to our overall understanding of cytoskeletal diversity and evolution. PMID:23312067

The secretory pathway: exploring yeast diversity.

PubMed

Delic, Marizela; Valli, Minoska; Graf, Alexandra B; Pfeffer, Martin; Mattanovich, Diethard; Gasser, Brigitte

2013-11-01

Protein secretion is an essential process for living organisms. In eukaryotes, this encompasses numerous steps mediated by several hundred cellular proteins. The core functions of translocation through the endoplasmic reticulum membrane, primary glycosylation, folding and quality control, and vesicle-mediated secretion are similar from yeasts to higher eukaryotes. However, recent research has revealed significant functional differences between yeasts and mammalian cells, and even among diverse yeast species. This review provides a current overview of the canonical protein secretion pathway in the model yeast Saccharomyces cerevisiae, highlighting differences to mammalian cells as well as currently unresolved questions, and provides a genomic comparison of the S. cerevisiae pathway to seven other yeast species where secretion has been investigated due to their attraction as protein production platforms, or for their relevance as pathogens. The analysis of Candida albicans, Candida glabrata, Kluyveromyces lactis, Pichia pastoris, Hansenula polymorpha, Yarrowia lipolytica, and Schizosaccharomyces pombe reveals that many - but not all - secretion steps are more redundant in S. cerevisiae due to duplicated genes, while some processes are even absent in this model yeast. Recent research obviates that even where homologous genes are present, small differences in protein sequence and/or differences in the regulation of gene expression may lead to quite different protein secretion phenotypes. © 2013 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

Malonylome Analysis Reveals the Involvement of Lysine Malonylation in Metabolism and Photosynthesis in Cyanobacteria.

PubMed

Ma, Yanyan; Yang, Mingkun; Lin, Xiaohuang; Liu, Xin; Huang, Hui; Ge, Feng

2017-05-05

As a recently validated reversible post translational modification, lysine malonylation regulates diverse cellular processes from bacteria to mammals, but its existence and function in photosynthetic organisms remain unknown. Cyanobacteria are the most ancient group of photosynthetic prokaryotes and contribute about 50% of the total primary production on Earth. Previously, we reported the lysine acetylome in the model cyanobacterium Synechocystis sp. PCC 6803 (Synechocystis). Here we performed the first proteomic survey of lysine malonylation in Synechocystis using highly accurate tandem mass spectrometry in combination with affinity purification. We identified 598 lysine malonylation sites on 339 proteins with high confidence in total. A bioinformatic analysis suggested that these malonylated proteins may play various functions and were distributed in diverse subcellular compartments. Among them, many malonylated proteins were involved in cellular metabolism. The functional significance of lysine malonylation in the metabolic enzyme activity of phosphoglycerate kinase (PGK) was determined by site-specific mutagenesis and biochemical studies. Interestingly, 27 proteins involved in photosynthesis were found to be malonylated for the first time, suggesting that lysine malonylation may be involved in photosynthesis. Thus our results provide the first lysine malonylome in a photosynthetic organism and suggest a previously unexplored role of lysine malonylation in the regulation of metabolic processes and photosynthesis in Synechocystis as well as in other photosynthetic organisms.

Omnipresence of the polyproline II helix in fibrous and globular proteins.

PubMed

Esipova, Natalia G; Tumanyan, Vladimir G

2017-02-01

Left-handed helical conformation of a polypeptide chain (PPII) is the third type of the protein backbone structure. This conformation universally exists in fibrous, globular proteins, and biologically active peptides. It has unique physical and chemical properties determining a wide range of biological functions, from the protein folding to the tissue differentiation. New examples of the structure have been appearing in spite of difficulties in their detection and investigation. The annotation and prediction of the PPII was also a challenging task. Recently, many PPII motifs with new and/or unexpected functions are being accumulated in databases. In this review we describe the major structural and dynamic forms of PPII, the diversity of its functions, and the role in different biological processes. Copyright © 2016 Elsevier Ltd. All rights reserved.

Chlorite dismutases, DyPs, and EfeB: 3 microbial heme enzyme families comprise the CDE structural superfamily

PubMed Central

Goblirsch, Brandon; Kurker, Richard C.; Streit, Bennett R.; Wilmot, Carrie M.; DuBois, Jennifer L.

2011-01-01

Heme proteins are extremely diverse, widespread, and versatile biocatalysts, sensors, and molecular transporters. The chlorite dismutase family of hemoproteins received its name due to the ability of the first-isolated members to detoxify anthropogenic ClO2−, a function believed to have evolved only in the last few decades. Family members have since been found in fifteen bacterial and archaeal genera, suggesting ancient roots. A structure- and sequence-based examination of the family is presented, in which key sequence and structural motifs are identified and possible functions for family proteins are proposed. Newly identified structural homologies moreover demonstrate clear connections to two other large, ancient, and functionally mysterious protein families. We propose calling them collectively the CDE superfamily of heme proteins. PMID:21354424

Orthologs, paralogs and genome comparisons

NASA Technical Reports Server (NTRS)

Gogarten, J. P.; Olendzenski, L.

1999-01-01

During the past decade, ancient gene duplications were recognized as one of the main forces in the generation of diverse gene families and the creation of new functional capabilities. New tools developed to search data banks for homologous sequences, and an increased availability of reliable three-dimensional structural information led to the recognition that proteins with diverse functions can belong to the same superfamily. Analyses of the evolution of these superfamilies promises to provide insights into early evolution but are complicated by several important evolutionary processes. Horizontal transfer of genes can lead to a vertical spread of innovations among organisms, therefore finding a certain property in some descendants of an ancestor does not guarantee that it was present in that ancestor. Complete or partial gene conversion between duplicated genes can yield phylogenetic trees with several, apparently independent gene duplications, suggesting an often surprising parallelism in the evolution of independent lineages. Additionally, the breakup of domains within a protein and the fusion of domains into multifunctional proteins makes the delineation of superfamilies a task that remains difficult to automate.

The bioactive acidic serine- and aspartate-rich motif peptide.

PubMed

Minamizaki, Tomoko; Yoshiko, Yuji

2015-01-01

The organic component of the bone matrix comprises 40% dry weight of bone. The organic component is mostly composed of type I collagen and small amounts of non-collagenous proteins (NCPs) (10-15% of the total bone protein content). The small integrin-binding ligand N-linked glycoprotein (SIBLING) family, a NCP, is considered to play a key role in bone mineralization. SIBLING family of proteins share common structural features and includes the arginine-glycine-aspartic acid (RGD) motif and acidic serine- and aspartic acid-rich motif (ASARM). Clinical manifestations of gene mutations and/or genetically modified mice indicate that SIBLINGs play diverse roles in bone and extraskeletal tissues. ASARM peptides might not be primary responsible for the functional diversity of SIBLINGs, but this motif is suggested to be a key domain of SIBLINGs. However, the exact function of ASARM peptides is poorly understood. In this article, we discuss the considerable progress made in understanding the role of ASARM as a bioactive peptide.

Evolution and Diversity of the Human Hepatitis D Virus Genome

PubMed Central

Huang, Chi-Ruei; Lo, Szecheng J.

2010-01-01

Human hepatitis delta virus (HDV) is the smallest RNA virus in genome. HDV genome is divided into a viroid-like sequence and a protein-coding sequence which could have originated from different resources and the HDV genome was eventually constituted through RNA recombination. The genome subsequently diversified through accumulation of mutations selected by interactions between the mutated RNA and proteins with host factors to successfully form the infectious virions. Therefore, we propose that the conservation of HDV nucleotide sequence is highly related with its functionality. Genome analysis of known HDV isolates shows that the C-terminal coding sequences of large delta antigen (LDAg) are the highest diversity than other regions of protein-coding sequences but they still retain biological functionality to interact with the heavy chain of clathrin can be selected and maintained. Since viruses interact with many host factors, including escaping the host immune response, how to design a program to predict RNA genome evolution is a great challenging work. PMID:20204073

Emerging roles for the BAI1 protein family in the regulation of phagocytosis, synaptogenesis, neurovasculature, and tumor development

PubMed Central

Cork, Sarah M.

2011-01-01

While G-protein-coupled receptors (GPCRs) have received considerable attention for their biological activity in a diversity of physiological functions and have become targets for therapeutic intervention in many diseases, the function of the cell adhesion subfamily of GPCRs remains poorly understood. Within this group, the family of brain angiogenesis inhibitor molecules (BAI1-3) has become increasingly appreciated for their diverse roles in biology and disease. In particular, recent findings suggest emerging roles for BAI1 in the regulation of phenomena including phagocytosis, synaptogenesis, and the inhibition of tumor growth and angiogenesis via the processing of its extracellular domain into secreted vasculostatins. Here we summarize the known biological features of the BAI proteins, including their structure, proteolysis events, and interacting partners, and their recently identified ability to regulate certain signaling pathways. Finally, we discuss the potential of the BAIs as therapeutics or targets for diseases as varied as cancer, stroke, and schizophrenia. PMID:21509575

Bacterial microcompartment assembly: The key role of encapsulation peptides

DOE PAGES

Aussignargues, Clément; Paasch, Bradley C.; Gonzalez-Esquer, Raul; ...

2015-06-23

Bacterial microcompartments (BMCs) are proteinaceous organelles used by a broad range of bacteria to segregate and optimize metabolic reactions. Their functions are diverse, and can be divided into anabolic (carboxysome) and catabolic (metabolosomes) processes, depending on their cargo enzymes. The assembly pathway for the β-carboxysome has been characterized, revealing that biogenesis proceeds from the inside out. The enzymes coalesce into a procarboxysome, followed by encapsulation in a protein shell that is recruited to the procarboxysome by a short (~17 amino acids) extension on the C-terminus of one of the encapsulated proteins. A similar extension is also found on the N-more » or C-termini of a subset of metabolosome core enzymes. These encapsulation peptides (EPs) are characterized by a primary structure predicted to form an amphipathic α-helix that interacts with shell proteins. In this study, we review the features, function and widespread occurrence of EPs among metabolosomes, and propose an expanded role for EPs in the assembly of diverse BMCs.« less

The Plant Peptidome: An Expanding Repertoire of Structural Features and Biological Functions[OPEN

PubMed Central

Tavormina, Patrizia; De Coninck, Barbara; Nikonorova, Natalia; De Smet, Ive; Cammue, Bruno P.A.

2015-01-01

Peptides fulfill a plethora of functions in plant growth, development, and stress responses. They act as key components of cell-to-cell communication, interfere with signaling and response pathways, or display antimicrobial activity. Strikingly, both the diversity and amount of plant peptides have been largely underestimated. Most characterized plant peptides to date acting as small signaling peptides or antimicrobial peptides are derived from nonfunctional precursor proteins. However, evidence is emerging on peptides derived from a functional protein, directly translated from small open reading frames (without the involvement of a precursor) or even encoded by primary transcripts of microRNAs. These novel types of peptides further add to the complexity of the plant peptidome, even though their number is still limited and functional characterization as well as translational evidence are often controversial. Here, we provide a comprehensive overview of the reported types of plant peptides, including their described functional and structural properties. We propose a novel, unifying peptide classification system to emphasize the enormous diversity in peptide synthesis and consequent complexity of the still expanding knowledge on the plant peptidome. PMID:26276833

Alternative intronic promoters in development and disease.

PubMed

Vacik, Tomas; Raska, Ivan

2017-05-01

Approximately 20,000 mammalian genes are estimated to encode between 250 thousand and 1 million different proteins. This enormous diversity of the mammalian proteome is caused by the ability of a single-gene locus to encode multiple protein isoforms. Protein isoforms encoded by one gene locus can be functionally distinct, and they can even have antagonistic functions. One of the mechanisms involved in creating this proteome complexity is alternative promoter usage. Alternative intronic promoters are located downstream from their canonical counterparts and drive the expression of alternative RNA isoforms that lack upstream exons. These upstream exons can encode some important functional domains, and proteins encoded by alternative mRNA isoforms can be thus functionally distinct from the full-length protein encoded by canonical mRNA isoforms. Since any misbalance of functionally distinct protein isoforms is likely to have detrimental consequences for the cell and the whole organism, their expression must be precisely regulated. Misregulation of alternative intronic promoters is frequently associated with various developmental defects and diseases including cancer, and it is becoming increasingly clear that this phenomenon deserves more attention.

Compounds that correct F508del-CFTR trafficking can also correct other protein trafficking diseases: an in vitro study using cell lines

PubMed Central

2013-01-01

Background Many genetic diseases are due to defects in protein trafficking where the mutant protein is recognized by the quality control systems, retained in the endoplasmic reticulum (ER), and degraded by the proteasome. In many cases, the mutant protein retains function if it can be trafficked to its proper cellular location. We have identified structurally diverse correctors that restore the trafficking and function of the most common mutation causing cystic fibrosis, F508del-CFTR. Most of these correctors do not act directly as ligands of CFTR, but indirectly on other pathways to promote folding and correction. We hypothesize that these proteostasis regulators may also correct other protein trafficking diseases. Methods To test our hypothesis, we used stable cell lines or transient transfection to express 2 well-studied trafficking disease mutations in each of 3 different proteins: the arginine-vasopressin receptor 2 (AVPR2, also known as V2R), the human ether-a-go-go-related gene (KCNH2, also known as hERG), and finally the sulfonylurea receptor 1 (ABCC8, also known as SUR1). We treated cells expressing these mutant proteins with 9 structurally diverse F508del-CFTR correctors that function through different cellular mechanisms and assessed whether correction occurred via immunoblotting and functional assays. Results were deemed significantly different from controls by a one-way ANOVA (p < 0.05). Results Here we show that F508del-CFTR correctors RDR1, KM60 and KM57 also correct some mutant alleles of other protein trafficking diseases. We also show that one corrector, the cardiac glycoside ouabain, was found to alter the glycosylation of all mutant alleles tested. Conclusions Correctors of F508del-CFTR trafficking might have broader applications to other protein trafficking diseases. PMID:23316740

Diverse structures, functions and uses of FK506 binding proteins.

PubMed

Bonner, Julia Maeve; Boulianne, Gabrielle L

2017-10-01

FK506 (Tacrolimus), isolated from Streptomyces tsukubaenis is a powerful immunosuppressant shown to inhibit T cell activation. FK506 mediated immunosuppression requires the formation of a complex between FK506, a FK506 binding protein (FKBP) and calcineurin. Numerous FKBPs have been identified in a wide range of species, from single celled organisms to humans. FKBPs show peptidylprolyl cis/trans isomerase (PPIase) activity and have been shown to affect a wide range of cellular processes including protein folding, receptor signaling and apoptosis. FKBPs also affect numerous biological functions in addition to immunosuppression including regulation of cardiac function, neuronal function and development and have been implicated in several diseases including cardiac disease, cancer and neurodegenerative diseases such as Alzheimer's disease. More recently, FKBPs have proven useful as molecular tools for studying protein interactions, localization and functions. This review provides an overview of the current state of knowledge of FKBPs and their numerous biological functions and uses. Copyright © 2017 Elsevier Inc. All rights reserved.

A carrot leucine-rich-repeat protein that inhibits ice recrystallization.

PubMed

Worrall, D; Elias, L; Ashford, D; Smallwood, M; Sidebottom, C; Lillford, P; Telford, J; Holt, C; Bowles, D

1998-10-02

Many organisms adapted to live at subzero temperatures express antifreeze proteins that improve their tolerance to freezing. Although structurally diverse, all antifreeze proteins interact with ice surfaces, depress the freezing temperature of aqueous solutions, and inhibit ice crystal growth. A protein purified from carrot shares these functional features with antifreeze proteins of fish. Expression of the carrot complementary DNA in tobacco resulted in the accumulation of antifreeze activity in the apoplast of plants grown at greenhouse temperatures. The sequence of carrot antifreeze protein is similar to that of polygalacturonase inhibitor proteins and contains leucine-rich repeats.

The prokaryotic antecedents of the ubiquitin-signaling system and the early evolution of ubiquitin-like β-grasp domains

PubMed Central

Iyer, Lakshminarayan M; Burroughs, A Maxwell; Aravind, L

2006-01-01

Background Ubiquitin (Ub)-mediated signaling is one of the hallmarks of all eukaryotes. Prokaryotic homologs of Ub (ThiS and MoaD) and E1 ligases have been studied in relation to sulfur incorporation reactions in thiamine and molybdenum/tungsten cofactor biosynthesis. However, there is no evidence for entire protein modification systems with Ub-like proteins and deconjugation by deubiquitinating enzymes in prokaryotes. Hence, the evolutionary assembly of the eukaryotic Ub-signaling apparatus remains unclear. Results We systematically analyzed prokaryotic Ub-related β-grasp fold proteins using sensitive sequence profile searches and structural analysis. Consequently, we identified novel Ub-related proteins beyond the characterized ThiS, MoaD, TGS, and YukD domains. To understand their functional associations, we sought and recovered several conserved gene neighborhoods and domain architectures. These included novel associations involving diverse sulfur metabolism proteins, siderophore biosynthesis and the gene encoding the transfer mRNA binding protein SmpB, as well as domain fusions between Ub-like domains and PIN-domain related RNAses. Most strikingly, we found conserved gene neighborhoods in phylogenetically diverse bacteria combining genes for JAB domains (the primary de-ubiquitinating isopeptidases of the proteasomal complex), along with E1-like adenylating enzymes and different Ub-related proteins. Further sequence analysis of other conserved genes in these neighborhoods revealed several Ub-conjugating enzyme/E2-ligase related proteins. Genes for an Ub-like protein and a JAB domain peptidase were also found in the tail assembly gene cluster of certain caudate bacteriophages. Conclusion These observations imply that members of the Ub family had already formed strong functional associations with E1-like proteins, UBC/E2-related proteins, and JAB peptidases in the bacteria. Several of these Ub-like proteins and the associated protein families are likely to function together in signaling systems just as in eukaryotes. PMID:16859499

Functional analysis of proteins and protein species using shotgun proteomics and linear mathematics.

PubMed

Hoehenwarter, Wolfgang; Chen, Yanmei; Recuenco-Munoz, Luis; Wienkoop, Stefanie; Weckwerth, Wolfram

2011-07-01

Covalent post-translational modification of proteins is the primary modulator of protein function in the cell. It greatly expands the functional potential of the proteome compared to the genome. In the past few years shotgun proteomics-based research, where the proteome is digested into peptides prior to mass spectrometric analysis has been prolific in this area. It has determined the kinetics of tens of thousands of sites of covalent modification on an equally large number of proteins under various biological conditions and uncovered a transiently active regulatory network that extends into diverse branches of cellular physiology. In this review, we discuss this work in light of the concept of protein speciation, which emphasizes the entire post-translationally modified molecule and its interactions and not just the modification site as the functional entity. Sometimes, particularly when considering complex multisite modification, all of the modified molecular species involved in the investigated condition, the protein species must be completely resolved for full understanding. We present a mathematical technique that delivers a good approximation for shotgun proteomics data.

SITEX 2.0: Projections of protein functional sites on eukaryotic genes. Extension with orthologous genes.

PubMed

Medvedeva, Irina V; Demenkov, Pavel S; Ivanisenko, Vladimir A

2017-04-01

Functional sites define the diversity of protein functions and are the central object of research of the structural and functional organization of proteins. The mechanisms underlying protein functional sites emergence and their variability during evolution are distinguished by duplication, shuffling, insertion and deletion of the exons in genes. The study of the correlation between a site structure and exon structure serves as the basis for the in-depth understanding of sites organization. In this regard, the development of programming resources that allow the realization of the mutual projection of exon structure of genes and primary and tertiary structures of encoded proteins is still the actual problem. Previously, we developed the SitEx system that provides information about protein and gene sequences with mapped exon borders and protein functional sites amino acid positions. The database included information on proteins with known 3D structure. However, data with respect to orthologs was not available. Therefore, we added the projection of sites positions to the exon structures of orthologs in SitEx 2.0. We implemented a search through database using site conservation variability and site discontinuity through exon structure. Inclusion of the information on orthologs allowed to expand the possibilities of SitEx usage for solving problems regarding the analysis of the structural and functional organization of proteins. Database URL: http://www-bionet.sscc.ru/sitex/ .

On the conservative nature of intragenic recombination

PubMed Central

Drummond, D. Allan; Silberg, Jonathan J.; Meyer, Michelle M.; Wilke, Claus O.; Arnold, Frances H.

2005-01-01

Intragenic recombination rapidly creates protein sequence diversity compared with random mutation, but little is known about the relative effects of recombination and mutation on protein function. Here, we compare recombination of the distantly related β-lactamases PSE-4 and TEM-1 to mutation of PSE-4. We show that, among β-lactamase variants containing the same number of amino acid substitutions, variants created by recombination retain function with a significantly higher probability than those generated by random mutagenesis. We present a simple model that accurately captures the differing effects of mutation and recombination in real and simulated proteins with only four parameters: (i) the amino acid sequence distance between parents, (ii) the number of substitutions, (iii) the average probability that random substitutions will preserve function, and (iv) the average probability that substitutions generated by recombination will preserve function. Our results expose a fundamental functional enrichment in regions of protein sequence space accessible by recombination and provide a framework for evaluating whether the relative rates of mutation and recombination observed in nature reflect the underlying imbalance in their effects on protein function. PMID:15809422

On the conservative nature of intragenic recombination.

PubMed

Drummond, D Allan; Silberg, Jonathan J; Meyer, Michelle M; Wilke, Claus O; Arnold, Frances H

2005-04-12

Intragenic recombination rapidly creates protein sequence diversity compared with random mutation, but little is known about the relative effects of recombination and mutation on protein function. Here, we compare recombination of the distantly related beta-lactamases PSE-4 and TEM-1 to mutation of PSE-4. We show that, among beta-lactamase variants containing the same number of amino acid substitutions, variants created by recombination retain function with a significantly higher probability than those generated by random mutagenesis. We present a simple model that accurately captures the differing effects of mutation and recombination in real and simulated proteins with only four parameters: (i) the amino acid sequence distance between parents, (ii) the number of substitutions, (iii) the average probability that random substitutions will preserve function, and (iv) the average probability that substitutions generated by recombination will preserve function. Our results expose a fundamental functional enrichment in regions of protein sequence space accessible by recombination and provide a framework for evaluating whether the relative rates of mutation and recombination observed in nature reflect the underlying imbalance in their effects on protein function.

AKAP-scaffolding proteins and regulation of cardiac physiology

PubMed Central

Mauban, JRH; O'Donnell, M; Warrier, S; Manni, S; Bond, M

2009-01-01

A kinase anchoring proteins (AKAPs) compose a growing list of diverse but functionally related proteins defined by their ability to bind to the regulatory subunit of protein kinase A. AKAPs perform an integral role in the spatiotemporal modulation of a multitude of cellular signaling pathways. This review highlights the extensive role of AKAPs in cardiac excitation/contraction coupling and cardiac physiology. The literature shows that particular AKAPs are involved in cardiac Ca2+ influx, release, re-uptake, and myocyte repolarization. Studies have also suggested roles for AKAPs in cardiac remodeling. Transgenic studies show functional effects of AKAPs, not only in the cardiovascular system, but in other organ systems as well. PMID:19364910

Modeling of DNA and Protein Organization Levels with Cn3D Software

ERIC Educational Resources Information Center

Stasinakis, Panagiotis K.; Nicolaou, Despoina

2017-01-01

The molecular structure of living organisms and the complex interactions amongst its components are the basis for the diversity observed at the macroscopic level. Proteins and nucleic acids are some of the major molecular components, and play a key role in several biological functions, such as those of development and evolution. This article…

Multiple structure-intrinsic disorder interactions regulate and coordinate Hox protein function

NASA Astrophysics Data System (ADS)

Bondos, Sarah

During animal development, Hox transcription factors determine fate of developing tissues to generate diverse organs and appendages. Hox proteins are famous for their bizarre mutant phenotypes, such as replacing antennae with legs. Clearly, the functions of individual Hox proteins must be distinct and reliable in vivo, or the organism risks malformation or death. However, within the Hox protein family, the DNA-binding homeodomains are highly conserved and the amino acids that contact DNA are nearly invariant. These observations raise the question: How do different Hox proteins correctly identify their distinct target genes using a common DNA binding domain? One possible means to modulate DNA binding is through the influence of the non-homeodomain protein regions, which differ significantly among Hox proteins. However genetic approaches never detected intra-protein interactions, and early biochemical attempts were hindered because the special features of ``intrinsically disordered'' sequences were not appreciated. We propose the first-ever structural model of a Hox protein to explain how specific contacts between distant, intrinsically disordered regions of the protein and the homeodomain regulate DNA binding and coordinate this activity with other Hox molecular functions.

Intermediate filament protein evolution and protists.

PubMed

Preisner, Harald; Habicht, Jörn; Garg, Sriram G; Gould, Sven B

2018-03-23

Metazoans evolved from a single protist lineage. While all eukaryotes share a conserved actin and tubulin-based cytoskeleton, it is commonly perceived that intermediate filaments (IFs), including lamin, vimentin or keratin among many others, are restricted to metazoans. Actin and tubulin proteins are conserved enough to be detectable across all eukaryotic genomes using standard phylogenetic methods, but IF proteins, in contrast, are notoriously difficult to identify by such means. Since the 1950s, dozens of cytoskeletal proteins in protists have been identified that seemingly do not belong to any of the IF families described for metazoans, yet, from a structural and functional perspective fit criteria that define metazoan IF proteins. Here, we briefly review IF protein discovery in metazoans and the implications this had for the definition of this protein family. We argue that the many cytoskeletal and filament-forming proteins of protists should be incorporated into a more comprehensive picture of IF evolution by aligning it with the recent identification of lamins across the phylogenetic diversity of eukaryotic supergroups. This then brings forth the question of how the diversity of IF proteins has unfolded. The evolution of IF proteins likely represents an example of convergent evolution, which, in combination with the speed with which these cytoskeletal proteins are evolving, generated their current diversity. IF proteins did not first emerge in metazoa, but in protists. Only the emergence of cytosolic IF proteins that appear to stem from a nuclear lamin is unique to animals and coincided with the emergence of true animal multicellularity. © 2018 Wiley Periodicals, Inc.

GalaxyDock BP2 score: a hybrid scoring function for accurate protein-ligand docking

NASA Astrophysics Data System (ADS)

Baek, Minkyung; Shin, Woong-Hee; Chung, Hwan Won; Seok, Chaok

2017-07-01

Protein-ligand docking is a useful tool for providing atomic-level understanding of protein functions in nature and design principles for artificial ligands or proteins with desired properties. The ability to identify the true binding pose of a ligand to a target protein among numerous possible candidate poses is an essential requirement for successful protein-ligand docking. Many previously developed docking scoring functions were trained to reproduce experimental binding affinities and were also used for scoring binding poses. However, in this study, we developed a new docking scoring function, called GalaxyDock BP2 Score, by directly training the scoring power of binding poses. This function is a hybrid of physics-based, empirical, and knowledge-based score terms that are balanced to strengthen the advantages of each component. The performance of the new scoring function exhibits significant improvement over existing scoring functions in decoy pose discrimination tests. In addition, when the score is used with the GalaxyDock2 protein-ligand docking program, it outperformed other state-of-the-art docking programs in docking tests on the Astex diverse set, the Cross2009 benchmark set, and the Astex non-native set. GalaxyDock BP2 Score and GalaxyDock2 with this score are freely available at http://galaxy.seoklab.org/softwares/galaxydock.html.

Tandem-repeat protein domains across the tree of life.

PubMed

Jernigan, Kristin K; Bordenstein, Seth R

2015-01-01

Tandem-repeat protein domains, composed of repeated units of conserved stretches of 20-40 amino acids, are required for a wide array of biological functions. Despite their diverse and fundamental functions, there has been no comprehensive assessment of their taxonomic distribution, incidence, and associations with organismal lifestyle and phylogeny. In this study, we assess for the first time the abundance of armadillo (ARM) and tetratricopeptide (TPR) repeat domains across all three domains in the tree of life and compare the results to our previous analysis on ankyrin (ANK) repeat domains in this journal. All eukaryotes and a majority of the bacterial and archaeal genomes analyzed have a minimum of one TPR and ARM repeat. In eukaryotes, the fraction of ARM-containing proteins is approximately double that of TPR and ANK-containing proteins, whereas bacteria and archaea are enriched in TPR-containing proteins relative to ARM- and ANK-containing proteins. We show in bacteria that phylogenetic history, rather than lifestyle or pathogenicity, is a predictor of TPR repeat domain abundance, while neither phylogenetic history nor lifestyle predicts ARM repeat domain abundance. Surprisingly, pathogenic bacteria were not enriched in TPR-containing proteins, which have been associated within virulence factors in certain species. Taken together, this comparative analysis provides a newly appreciated view of the prevalence and diversity of multiple types of tandem-repeat protein domains across the tree of life. A central finding of this analysis is that tandem repeat domain-containing proteins are prevalent not just in eukaryotes, but also in bacterial and archaeal species.

A chemical proteomics approach for global analysis of lysine monomethylome profiling.

PubMed

Wu, Zhixiang; Cheng, Zhongyi; Sun, Mingwei; Wan, Xuelian; Liu, Ping; He, Tieming; Tan, Minjia; Zhao, Yingming

2015-02-01

Methylation of lysine residues on histone proteins is known to play an important role in chromatin structure and function. However, non-histone protein substrates of this modification remain largely unknown. An effective approach for system-wide analysis of protein lysine methylation, particularly lysine monomethylation, is lacking. Here we describe a chemical proteomics approach for global screening for monomethyllysine substrates, involving chemical propionylation of monomethylated lysine, affinity enrichment of the modified monomethylated peptides, and HPLC/MS/MS analysis. Using this approach, we identified with high confidence 446 lysine monomethylation sites in 398 proteins, including three previously unknown histone monomethylation marks, representing the largest data set of protein lysine monomethylation described to date. Our data not only confirms previously discovered lysine methylation substrates in the nucleus and spliceosome, but also reveals new substrates associated with diverse biological processes. This method hence offers a powerful approach for dynamic study of protein lysine monomethylation under diverse cellular conditions and in human diseases. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

CPI motif interaction is necessary for capping protein function in cells

PubMed Central

Edwards, Marc; McConnell, Patrick; Schafer, Dorothy A.; Cooper, John A.

2015-01-01

Capping protein (CP) has critical roles in actin assembly in vivo and in vitro. CP binds with high affinity to the barbed end of actin filaments, blocking the addition and loss of actin subunits. Heretofore, models for actin assembly in cells generally assumed that CP is constitutively active, diffusing freely to find and cap barbed ends. However, CP can be regulated by binding of the ‘capping protein interaction' (CPI) motif, found in a diverse and otherwise unrelated set of proteins that decreases, but does not abolish, the actin-capping activity of CP and promotes uncapping in biochemical experiments. Here, we report that CP localization and the ability of CP to function in cells requires interaction with a CPI-motif-containing protein. Our discovery shows that cells target and/or modulate the capping activity of CP via CPI motif interactions in order for CP to localize and function in cells. PMID:26412145

The Development of Target-Specific Pose Filter Ensembles To Boost Ligand Enrichment for Structure-Based Virtual Screening.

PubMed

Xia, Jie; Hsieh, Jui-Hua; Hu, Huabin; Wu, Song; Wang, Xiang Simon

2017-06-26

Structure-based virtual screening (SBVS) has become an indispensable technique for hit identification at the early stage of drug discovery. However, the accuracy of current scoring functions is not high enough to confer success to every target and thus remains to be improved. Previously, we had developed binary pose filters (PFs) using knowledge derived from the protein-ligand interface of a single X-ray structure of a specific target. This novel approach had been validated as an effective way to improve ligand enrichment. Continuing from it, in the present work we attempted to incorporate knowledge collected from diverse protein-ligand interfaces of multiple crystal structures of the same target to build PF ensembles (PFEs). Toward this end, we first constructed a comprehensive data set to meet the requirements of ensemble modeling and validation. This set contains 10 diverse targets, 118 well-prepared X-ray structures of protein-ligand complexes, and large benchmarking actives/decoys sets. Notably, we designed a unique workflow of two-layer classifiers based on the concept of ensemble learning and applied it to the construction of PFEs for all of the targets. Through extensive benchmarking studies, we demonstrated that (1) coupling PFE with Chemgauss4 significantly improves the early enrichment of Chemgauss4 itself and (2) PFEs show greater consistency in boosting early enrichment and larger overall enrichment than our prior PFs. In addition, we analyzed the pairwise topological similarities among cognate ligands used to construct PFEs and found that it is the higher chemical diversity of the cognate ligands that leads to the improved performance of PFEs. Taken together, the results so far prove that the incorporation of knowledge from diverse protein-ligand interfaces by ensemble modeling is able to enhance the screening competence of SBVS scoring functions.

Polycomb Group (PcG) Proteins and Human Cancers: Multifaceted Functions and Therapeutic Implications.

PubMed

Wang, Wei; Qin, Jiang-Jiang; Voruganti, Sukesh; Nag, Subhasree; Zhou, Jianwei; Zhang, Ruiwen

2015-11-01

Polycomb group (PcG) proteins are transcriptional repressors that regulate several crucial developmental and physiological processes in the cell. More recently, they have been found to play important roles in human carcinogenesis and cancer development and progression. The deregulation and dysfunction of PcG proteins often lead to blocking or inappropriate activation of developmental pathways, enhancing cellular proliferation, inhibiting apoptosis, and increasing the cancer stem cell population. Genetic and molecular investigations of PcG proteins have long been focused on their PcG functions. However, PcG proteins have recently been shown to exert non-classical-Pc-functions, contributing to the regulation of diverse cellular functions. We and others have demonstrated that PcG proteins regulate the expression and function of several oncogenes and tumor suppressor genes in a PcG-independent manner, and PcG proteins are associated with the survival of patients with cancer. In this review, we summarize the recent advances in the research on PcG proteins, including both the Pc-repressive and non-classical-Pc-functions. We specifically focus on the mechanisms by which PcG proteins play roles in cancer initiation, development, and progression. Finally, we discuss the potential value of PcG proteins as molecular biomarkers for the diagnosis and prognosis of cancer, and as molecular targets for cancer therapy. © 2015 Wiley Periodicals, Inc.

Comparative proteomics reveal diverse functions and dynamic changes of Bombyx mori silk proteins spun from different development stages.

PubMed

Dong, Zhaoming; Zhao, Ping; Wang, Chen; Zhang, Yan; Chen, Jianping; Wang, Xin; Lin, Ying; Xia, Qingyou

2013-11-01

Silkworms (Bombyx mori) produce massive amounts of silk proteins to make cocoons during the final stages of larval development. Although the major components, fibroin and sericin, have been the focus for a long time, few researchers have realized the complexity of the silk proteome. We collected seven kinds of silk fibers spun by silkworm larvae at different developmental stages: the silks spun by new hatched larvae, second instar day 0 larvae, third instar day 0 larvae, fourth instar day 0 larvae, and fourth instar molting larvae, the scaffold silk used to attach the cocoon to the substrate and the cocoon silk. Analysis by liquid chromatography-tandem mass spectrometry identified 500 proteins from the seven silks. In addition to the expected fibroins, sericins, and some known protease inhibitors, we also identified further protease inhibitors, enzymes, proteins of unknown function, and other proteins. Unsurprisingly, our quantitative results showed fibroins and sericins were the most abundant proteins in all seven silks. Except for fibroins and sericins, protease inhibitors, enzymes, and proteins of unknown function were more abundant than other proteins. We found significant change in silk protein compositions through development, being consistent with their different biological functions and complicated formation.

Mining on scorpion venom biodiversity.

PubMed

Rodríguez de la Vega, Ricardo C; Schwartz, Elisabeth F; Possani, Lourival D

2010-12-15

Scorpion venoms are complex mixtures of dozens or even hundreds of distinct proteins, many of which are inter-genome active elements. Fifty years after the first scorpion toxin sequences were determined, chromatography-assisted purification followed by automated protein sequencing or gene cloning, on a case-by-case basis, accumulated nearly 250 amino acid sequences of scorpion venom components. A vast majority of the available sequences correspond to proteins adopting a common three-dimensional fold, whose ion channel modulating functions have been firmly established or could be confidently inferred. However, the actual molecular diversity contained in scorpion venoms -as revealed by bioassay-driven purification, some unexpected activities of "canonical" neurotoxins and even serendipitous discoveries- is much larger than those "canonical" toxin types. In the last few years mining into the molecular diversity contained in scorpion has been assisted by high-throughput Mass Spectrometry techniques and large-scale DNA sequencing, collectively accounting for the more than twofold increase in the number of known sequences of scorpion venom components (now reaching 500 unique sequences). This review, from a comparative perspective, deals with recent data obtained by proteomic and transcriptomic studies on scorpion venoms and venom glands. Altogether, these studies reveal a large contribution of non canonical venom components, which would account for more than half of the total protein diversity of any scorpion venom. On top of aiding at the better understanding of scorpion venom biology, whether in the context of venom function or within the venom gland itself, these "novel" venom components certainly are an interesting source of bioactive proteins, whose characterization is worth pursuing. Copyright © 2009 Elsevier Ltd. All rights reserved.

Protein kinase C isoforms at the neuromuscular junction: localization and specific roles in neurotransmission and development.

PubMed

Lanuza, Maria A; Santafe, Manel M; Garcia, Neus; Besalduch, Núria; Tomàs, Marta; Obis, Teresa; Priego, Mercedes; Nelson, Phillip G; Tomàs, Josep

2014-01-01

The protein kinase C family (PKC) regulates a variety of neural functions including neurotransmitter release. The selective activation of a wide range of PKC isoforms in different cells and domains is likely to contribute to the functional diversity of PKC phosphorylating activity. In this review, we describe the isoform localization, phosphorylation function, regulation and signalling of the PKC family at the neuromuscular junction. Data show the involvement of the PKC family in several important functions at the neuromuscular junction and in particular in the maturation of the synapse and the modulation of neurotransmission in the adult. © 2013 Anatomical Society.

Abundance, diversity and domain architecture variability in prokaryotic DNA-binding transcription factors.

PubMed

Perez-Rueda, Ernesto; Hernandez-Guerrero, Rafael; Martinez-Nuñez, Mario Alberto; Armenta-Medina, Dagoberto; Sanchez, Israel; Ibarra, J Antonio

2018-01-01

Gene regulation at the transcriptional level is a central process in all organisms, and DNA-binding transcription factors, known as TFs, play a fundamental role. This class of proteins usually binds at specific DNA sequences, activating or repressing gene expression. In general, TFs are composed of two domains: the DNA-binding domain (DBD) and an extra domain, which in this work we have named "companion domain" (CD). This latter could be involved in one or more functions such as ligand binding, protein-protein interactions or even with enzymatic activity. In contrast to DBDs, which have been widely characterized both experimentally and bioinformatically, information on the abundance, distribution, variability and possible role of the CDs is scarce. Here, we investigated these issues associated with the domain architectures of TFs in prokaryotic genomes. To this end, 19 families of TFs in 761 non-redundant bacterial and archaeal genomes were evaluated. In this regard we found four main groups based on the abundance and distribution in the analyzed genomes: i) LysR and TetR/AcrR; ii) AraC/XylS, SinR, and others; iii) Lrp, Fis, ArsR, and others; and iv) a group that included only two families, ArgR and BirA. Based on a classification of the organisms according to the life-styles, a major abundance of regulatory families in free-living organisms, in contrast with pathogenic, extremophilic or intracellular organisms, was identified. Finally, the protein architecture diversity associated to the 19 families considering a weight score for domain promiscuity evidenced which regulatory families were characterized by either a large diversity of CDs, here named as "promiscuous" families given the elevated number of variable domains found in those TFs, or a low diversity of CDs. Altogether this information helped us to understand the diversity and distribution of the 19 Prokaryotes TF families. Moreover, initial steps were taken to comprehend the variability of the extra domain in those TFs, which eventually might assist in evolutionary and functional studies.

Oxidative Modulation of Voltage-Gated Potassium Channels

PubMed Central

Sahoo, Nirakar; Hoshi, Toshinori

2014-01-01

Abstract Significance: Voltage-gated K+ channels are a large family of K+-selective ion channel protein complexes that open on membrane depolarization. These K+ channels are expressed in diverse tissues and their function is vital for numerous physiological processes, in particular of neurons and muscle cells. Potentially reversible oxidative regulation of voltage-gated K+ channels by reactive species such as reactive oxygen species (ROS) represents a contributing mechanism of normal cellular plasticity and may play important roles in diverse pathologies including neurodegenerative diseases. Recent Advances: Studies using various protocols of oxidative modification, site-directed mutagenesis, and structural and kinetic modeling provide a broader phenomenology and emerging mechanistic insights. Critical Issues: Physicochemical mechanisms of the functional consequences of oxidative modifications of voltage-gated K+ channels are only beginning to be revealed. In vivo documentation of oxidative modifications of specific amino-acid residues of various voltage-gated K+ channel proteins, including the target specificity issue, is largely absent. Future Directions: High-resolution chemical and proteomic analysis of ion channel proteins with respect to oxidative modification combined with ongoing studies on channel structure and function will provide a better understanding of how the function of voltage-gated K+ channels is tuned by ROS and the corresponding reducing enzymes to meet cellular needs. Antioxid. Redox Signal. 21, 933–952. PMID:24040918

Mapping transcription factor interactome networks using HaloTag protein arrays.

PubMed

Yazaki, Junshi; Galli, Mary; Kim, Alice Y; Nito, Kazumasa; Aleman, Fernando; Chang, Katherine N; Carvunis, Anne-Ruxandra; Quan, Rosa; Nguyen, Hien; Song, Liang; Alvarez, José M; Huang, Shao-Shan Carol; Chen, Huaming; Ramachandran, Niroshan; Altmann, Stefan; Gutiérrez, Rodrigo A; Hill, David E; Schroeder, Julian I; Chory, Joanne; LaBaer, Joshua; Vidal, Marc; Braun, Pascal; Ecker, Joseph R

2016-07-19

Protein microarrays enable investigation of diverse biochemical properties for thousands of proteins in a single experiment, an unparalleled capacity. Using a high-density system called HaloTag nucleic acid programmable protein array (HaloTag-NAPPA), we created high-density protein arrays comprising 12,000 Arabidopsis ORFs. We used these arrays to query protein-protein interactions for a set of 38 transcription factors and transcriptional regulators (TFs) that function in diverse plant hormone regulatory pathways. The resulting transcription factor interactome network, TF-NAPPA, contains thousands of novel interactions. Validation in a benchmarked in vitro pull-down assay revealed that a random subset of TF-NAPPA validated at the same rate of 64% as a positive reference set of literature-curated interactions. Moreover, using a bimolecular fluorescence complementation (BiFC) assay, we confirmed in planta several interactions of biological interest and determined the interaction localizations for seven pairs. The application of HaloTag-NAPPA technology to plant hormone signaling pathways allowed the identification of many novel transcription factor-protein interactions and led to the development of a proteome-wide plant hormone TF interactome network.

Crystal Structure of a Plant Multidrug and Toxic Compound Extrusion Family Protein.

PubMed

Tanaka, Yoshiki; Iwaki, Shigehiro; Tsukazaki, Tomoya

2017-09-05

The multidrug and toxic compound extrusion (MATE) family of proteins consists of transporters responsible for multidrug resistance in prokaryotes. In plants, a number of MATE proteins were identified by recent genomic and functional studies, which imply that the proteins have substrate-specific transport functions instead of multidrug extrusion. The three-dimensional structure of eukaryotic MATE proteins, including those of plants, has not been reported, preventing a better understanding of the molecular mechanism of these proteins. Here, we describe the crystal structure of a MATE protein from the plant Camelina sativa at 2.9 Å resolution. Two sets of six transmembrane α helices, assembled pseudo-symmetrically, possess a negatively charged internal pocket with an outward-facing shape. The crystal structure provides insight into the diversity of plant MATE proteins and their substrate recognition and transport through the membrane. Copyright © 2017 Elsevier Ltd. All rights reserved.

The TRP channel superfamily: insights into how structure, protein-lipid interactions and localization influence function.

PubMed

Reaves, B J; Wolstenholme, A J

2007-02-01

TRP (transient receptor potential) cationic channels are key molecules that are involved in a variety of diverse biological processes ranging from fertility to osmosensation and nociception. Increasing our knowledge of these channels will help us to understand a range of physiological and pathogenic processes, as well as highlighting potential therapeutic drug targets. The founding members of the TRP family, Drosophila TRP and TRPL (TRP-like) proteins, were identified within the last two decades and there has been a subsequent explosion in the number and type of TRP channel described. Although information is accumulating as to the function of some of the TRP channels, the activation and inactivation mechanisms, structure, and interacting proteins of many, if not most, are awaiting elucidation. The Cell and Molecular Biology of TRP Channels Meeting held at the University of Bath included speakers working on a number of the different subfamilies of TRP channels and provided a basis for highlighting both similarities and differences between these groups. As the TRP channels mediate diverse functions, this meeting also brought together an audience with wide-ranging research interests, including biochemistry, cell biology, physiology and neuroscience, and inspired lively discussion on the issues reviewed herein.

Critical protein GAPDH and its regulatory mechanisms in cancer cells

PubMed Central

Zhang, Jin-Ying; Zhang, Fan; Hong, Chao-Qun; Giuliano, Armando E.; Cui, Xiao-Jiang; Zhou, Guang-Ji; Zhang, Guo-Jun; Cui, Yu-Kun

2015-01-01

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH), initially identified as a glycolytic enzyme and considered as a housekeeping gene, is widely used as an internal control in experiments on proteins, mRNA, and DNA. However, emerging evidence indicates that GAPDH is implicated in diverse functions independent of its role in energy metabolism; the expression status of GAPDH is also deregulated in various cancer cells. One of the most common effects of GAPDH is its inconsistent role in the determination of cancer cell fate. Furthermore, studies have described GAPDH as a regulator of cell death; other studies have suggested that GAPDH participates in tumor progression and serves as a new therapeutic target. However, related regulatory mechanisms of its numerous cellular functions and deregulated expression levels remain unclear. GAPDH is tightly regulated at transcriptional and posttranscriptional levels, which are involved in the regulation of diverse GAPDH functions. Several cancer-related factors, such as insulin, hypoxia inducible factor-1 (HIF-1), p53, nitric oxide (NO), and acetylated histone, not only modulate GAPDH gene expression but also affect protein functions via common pathways. Moreover, posttranslational modifications (PTMs) occurring in GAPDH in cancer cells result in new activities unrelated to the original glycolytic function of GAPDH. In this review, recent findings related to GAPDH transcriptional regulation and PTMs are summarized. Mechanisms and pathways involved in GAPDH regulation and its different roles in cancer cells are also described. PMID:25859407

Similarity in Shape Dictates Signature Intrinsic Dynamics Despite No Functional Conservation in TIM Barrel Enzymes

PubMed Central

Tiwari, Sandhya P.; Reuter, Nathalie

2016-01-01

The conservation of the intrinsic dynamics of proteins emerges as we attempt to understand the relationship between sequence, structure and functional conservation. We characterise the conservation of such dynamics in a case where the structure is conserved but function differs greatly. The triosephosphate isomerase barrel fold (TBF), renowned for its 8 β-strand-α-helix repeats that close to form a barrel, is one of the most diverse and abundant folds found in known protein structures. Proteins with this fold have diverse enzymatic functions spanning five of six Enzyme Commission classes, and we have picked five different superfamily candidates for our analysis using elastic network models. We find that the overall shape is a large determinant in the similarity of the intrinsic dynamics, regardless of function. In particular, the β-barrel core is highly rigid, while the α-helices that flank the β-strands have greater relative mobility, allowing for the many possibilities for placement of catalytic residues. We find that these elements correlate with each other via the loops that link them, as opposed to being directly correlated. We are also able to analyse the types of motions encoded by the normal mode vectors of the α-helices. We suggest that the global conservation of the intrinsic dynamics in the TBF contributes greatly to its success as an enzymatic scaffold both through evolution and enzyme design. PMID:27015412

R4 RGS Proteins: Regulation of G Protein Signaling and Beyond

PubMed Central

Bansal, Geetanjali; Druey, Kirk M.; Xie, Zhihui

2007-01-01

The Regulators of G protein Signaling (RGS) proteins were initially characterized as inhibitors of signal transduction cascades initiated by G-protein-coupled receptors (GPCRs) because of their ability to increase the intrinsic GTPase activity of heterotrimeric G proteins. This GTPase accelerating (GAP) activity enhances G protein deactivation and promotes desensitization. However, in addition to this signature trait, emerging data have revealed an expanding network of proteins, lipids, and ions that interact with RGS proteins and confer additional regulatory functions. This review highlights recent advances in our understanding of the physiological functions of one subfamily of RGS proteins with a high degree of homology (B/R4) gleaned from recent studies of knockout mice or cells with reduced RGS expression. We also discuss some of the newly-appreciated interactions of RGS proteins with cellular factors that suggest RGS control of several components of G-protein-mediated pathways as well as a diverse array of non-GPCR-mediated biological responses. PMID:18006065

Disentangling the role of management, vegetation structure, and plant quality for Orthoptera in lowland meadows.

PubMed

Schirmel, Jens; Gerlach, Rebekka; Buhk, Constanze

2017-08-17

Seminatural grasslands provide habitats for various species and are important for biodiversity conservation. The understanding of the diverse responses of species and traits to different grassland management methods is therefore urgently needed. We disentangled the role of grassland management (fertilization and irrigation), vegetation structure (biomass, sward height) and plant quality (protein and fiber content) for Orthoptera communities in lowland hay meadows in Germany. We found vegetation structure to be the most important environmental category in explaining community structure of Orthoptera (species richness, total individuals, functional diversity and species composition). Intensively used meadows (fertilized, irrigated, high plant biomass) were characterized by assemblages with few species, low functional diversity, and low conservation value. Thereby, the relatively moderate fertilizer inputs in our study system of up to ∼75 kg N/ha/year reduced functional diversity of Orthoptera, while this negative effect of fertilization was not detectable when solely considering taxonomic aspects. We found strong support for a prominent role of plant quality in shaping Orthoptera communities and especially the trait composition. Our findings demonstrate the usefulness of considering both taxonomic and functional components (functional diversity) in biodiversity research and we suggest a stronger involvement of plant quality measures in Orthoptera studies. © 2017 Institute of Zoology, Chinese Academy of Sciences.

A-Kinase Anchoring Proteins That Regulate Cardiac Remodeling

PubMed Central

Carnegie, Graeme K.; Burmeister, Brian T.

2012-01-01

In response to injury or stress, the adult heart undergoes maladaptive changes, collectively defined as pathological cardiac remodeling. Here, we focus on the role of A-kinase anchoring proteins (AKAPs) in 3 main areas associated with cardiac remodeling and the progression of heart failure: excitation–contraction coupling, sarcomeric regulation, and induction of pathological hypertrophy. AKAPs are a diverse family of scaffold proteins that form multi-protein complexes, integrating cAMP signaling with protein kinases, phosphatases, and other effector proteins. Many AKAPs have been characterized in the heart, where they play a critical role in modulating cardiac function. PMID:22075671

MicRhoDE: a curated database for the analysis of microbial rhodopsin diversity and evolution

PubMed Central

Boeuf, Dominique; Audic, Stéphane; Brillet-Guéguen, Loraine; Caron, Christophe; Jeanthon, Christian

2015-01-01

Microbial rhodopsins are a diverse group of photoactive transmembrane proteins found in all three domains of life and in viruses. Today, microbial rhodopsin research is a flourishing research field in which new understandings of rhodopsin diversity, function and evolution are contributing to broader microbiological and molecular knowledge. Here, we describe MicRhoDE, a comprehensive, high-quality and freely accessible database that facilitates analysis of the diversity and evolution of microbial rhodopsins. Rhodopsin sequences isolated from a vast array of marine and terrestrial environments were manually collected and curated. To each rhodopsin sequence are associated related metadata, including predicted spectral tuning of the protein, putative activity and function, taxonomy for sequences that can be linked to a 16S rRNA gene, sampling date and location, and supporting literature. The database currently covers 7857 aligned sequences from more than 450 environmental samples or organisms. Based on a robust phylogenetic analysis, we introduce an operational classification system with multiple phylogenetic levels ranging from superclusters to species-level operational taxonomic units. An integrated pipeline for online sequence alignment and phylogenetic tree construction is also provided. With a user-friendly interface and integrated online bioinformatics tools, this unique resource should be highly valuable for upcoming studies of the biogeography, diversity, distribution and evolution of microbial rhodopsins. Database URL: http://micrhode.sb-roscoff.fr. PMID:26286928

MicRhoDE: a curated database for the analysis of microbial rhodopsin diversity and evolution.

PubMed

Boeuf, Dominique; Audic, Stéphane; Brillet-Guéguen, Loraine; Caron, Christophe; Jeanthon, Christian

2015-01-01

Microbial rhodopsins are a diverse group of photoactive transmembrane proteins found in all three domains of life and in viruses. Today, microbial rhodopsin research is a flourishing research field in which new understandings of rhodopsin diversity, function and evolution are contributing to broader microbiological and molecular knowledge. Here, we describe MicRhoDE, a comprehensive, high-quality and freely accessible database that facilitates analysis of the diversity and evolution of microbial rhodopsins. Rhodopsin sequences isolated from a vast array of marine and terrestrial environments were manually collected and curated. To each rhodopsin sequence are associated related metadata, including predicted spectral tuning of the protein, putative activity and function, taxonomy for sequences that can be linked to a 16S rRNA gene, sampling date and location, and supporting literature. The database currently covers 7857 aligned sequences from more than 450 environmental samples or organisms. Based on a robust phylogenetic analysis, we introduce an operational classification system with multiple phylogenetic levels ranging from superclusters to species-level operational taxonomic units. An integrated pipeline for online sequence alignment and phylogenetic tree construction is also provided. With a user-friendly interface and integrated online bioinformatics tools, this unique resource should be highly valuable for upcoming studies of the biogeography, diversity, distribution and evolution of microbial rhodopsins. Database URL: http://micrhode.sb-roscoff.fr. © The Author(s) 2015. Published by Oxford University Press.

Bacterial Inclusion Bodies: Discovering Their Better Half.

PubMed

Rinas, Ursula; Garcia-Fruitós, Elena; Corchero, José Luis; Vázquez, Esther; Seras-Franzoso, Joaquin; Villaverde, Antonio

2017-09-01

Bacterial inclusion bodies (IBs) are functional, non-toxic amyloids occurring in recombinant bacteria showing analogies with secretory granules of the mammalian endocrine system. The scientific interest in these mesoscale protein aggregates has been historically masked by their status as a hurdle in recombinant protein production. However, progressive understanding of how the cell handles the quality of recombinant polypeptides and the main features of their intriguing molecular organization has stimulated the interest in inclusion bodies and spurred their use in diverse technological fields. The engineering and tailoring of IBs as functional protein particles for materials science and biomedicine is a good example of how formerly undesired bacterial byproducts can be rediscovered as promising functional materials for a broad spectrum of applications. Copyright © 2017 Elsevier Ltd. All rights reserved.

Affinity resins as new tools for identifying target proteins of ascorbic acid.

PubMed

Iwaoka, Yuji; Nishino, Kohei; Ishikawa, Takahiro; Ito, Hideyuki; Sawa, Yoshihiro; Tai, Akihiro

2018-02-12

l-Ascorbic acid (AA) has diverse physiological functions, but little is known about the functional mechanisms of AA. In this study, we synthesized two types of affinity resin on which AA is immobilized in a stable form to identify new AA-targeted proteins, which can provide important clues for elucidating unknown functional mechanisms of AA. To our knowledge, an affinity resin on which AA as a ligand is immobilized has not been prepared, because AA is very unstable and rapidly degraded in an aqueous solution. By using the affinity resins, cytochrome c (cyt c) was identified as an AA-targeted protein, and we showed that oxidized cyt c exhibits specific affinity for AA. These results suggest that two kinds of AA-affinity resin can be powerful tools to identify new target proteins of AA.

Structural Insight into the Assembly of TRPV Channels

PubMed Central

Huynh, Kevin W.; Cohen, Matthew R.; Chakrapani, Sudha; Holdaway, Heather A.; Stewart, Phoebe L.; Moiseenkova-Bell, Vera Y.

2017-01-01

SUMMARY Transient receptor potential (TRP) proteins are a large family of polymodal nonselective cation channels. The TRP vanilloid (TRPV) subfamily consists of six homologous members with diverse functions. TRPV1–TRPV4 are nonselective cation channels proposed to play a role in nociception, while TRPV5 and TRPV6 are involved in epithelial Ca2+ homeostasis. Here we present the cryo-electron microscopy (cryo-EM) structure of functional, full-length TRPV2 at 13.6 Å resolution. The map reveals that the TRPV2 cytoplasmic domain displays a 4-fold petal-like shape in which high-resolution N-terminal ankyrin repeat domain (ARD) structures can be unambiguously fitted. Fitting of the available ARD structures for other TRPV subfamily members into the TRPV2 EM map suggests that TRPV subfamily members have highly homologous structural topologies. These results allowed us to postulate a structural explanation for the functional diversity among TRPV channels and their differential regulation by proteins and ligands. PMID:24373766

Biased signaling of the proton-sensing receptor OGR1 by benzodiazepines.

PubMed

Pera, Tonio; Deshpande, Deepak A; Ippolito, Michael; Wang, Bin; Gavrila, Adelina; Michael, James V; Nayak, Ajay P; Tompkins, Eric; Farrell, Eleni; Kroeze, Wesley K; Roth, Bryan L; Panettieri, Reynold A; Benovic, Jeffrey L; An, Steven S; Dulin, Nickolai O; Penn, Raymond B

2018-02-01

GPCRs have diverse signaling capabilities, based on their ability to assume various conformations. Moreover, it is now appreciated that certain ligands can promote distinct receptor conformations and thereby bias signaling toward a specific pathway to differentially affect cell function. The recently deorphanized G protein-coupled receptor OGR1 [ovarian cancer G protein-coupled receptor 1 ( GPR68)] exhibits diverse signaling events when stimulated by reductions in extracellular pH. We recently demonstrated airway smooth muscle cells transduce multiple signaling events, reflecting a diverse capacity to couple to multiple G proteins. Moreover, we recently discovered that the benzodiazepine lorazepam, more commonly recognized as an agonist of the γ-aminobutyric acid A (GABA A ) receptor, can function as an allosteric modulator of OGR1 and, similarly, can promote multiple signaling events. In this study, we demonstrated that different benzodiazepines exhibit a range of biases for OGR1, with sulazepam selectively activating the canonical Gs of the G protein signaling pathway, in heterologous expression systems, as well as in several primary cell types. These findings highlight the potential power of biased ligand pharmacology for manipulating receptor signaling qualitatively, to preferentially activate pathways that are therapeutically beneficial.-Pera, T., Deshpande, D. A., Ippolito, M., Wang, B., Gavrila, A., Michael, J. V., Nayak, A. P., Tompkins, E., Farrell, E., Kroeze, W. K., Roth, B. L., Panettieri, R. A. Jr Benovic, J. L., An, S. S., Dulin, N. O., Penn, R. B. Biased signaling of the proton-sensing receptor OGR1 by benzodiazepines.

Exploiting a wheat EST database to assess genetic diversity

PubMed Central

2010-01-01

Expressed sequence tag (EST) markers have been used to assess variety and genetic diversity in wheat (Triticum aestivum). In this study, 1549 ESTs from wheat infested with yellow rust were used to examine the genetic diversity of six susceptible and resistant wheat cultivars. The aim of using these cultivars was to improve the competitiveness of public wheat breeding programs through the intensive use of modern, particularly marker-assisted, selection technologies. The F2 individuals derived from cultivar crosses were screened for resistance to yellow rust at the seedling stage in greenhouses and adult stage in the field to identify DNA markers genetically linked to resistance. Five hundred and sixty ESTs were assembled into 136 contigs and 989 singletons. BlastX search results showed that 39 (29%) contigs and 96 (10%) singletons were homologous to wheat genes. The database-matched contigs and singletons were assigned to eight functional groups related to protein synthesis, photosynthesis, metabolism and energy, stress proteins, transporter proteins, protein breakdown and recycling, cell growth and division and reactive oxygen scavengers. PCR analyses with primers based on the contigs and singletons showed that the most polymorphic functional categories were photosynthesis (contigs) and metabolism and energy (singletons). EST analysis revealed considerable genetic variability among the Turkish wheat cultivars resistant and susceptible to yellow rust disease and allowed calculation of the mean genetic distance between cultivars, with the greatest similarity (0.725) being between Harmankaya99 and Sönmez2001, and the lowest (0.622) between Aytin98 and Izgi01. PMID:21637582

Exploiting a wheat EST database to assess genetic diversity.

PubMed

Karakas, Ozge; Gurel, Filiz; Uncuoglu, Ahu Altinkut

2010-10-01

Expressed sequence tag (EST) markers have been used to assess variety and genetic diversity in wheat (Triticum aestivum). In this study, 1549 ESTs from wheat infested with yellow rust were used to examine the genetic diversity of six susceptible and resistant wheat cultivars. The aim of using these cultivars was to improve the competitiveness of public wheat breeding programs through the intensive use of modern, particularly marker-assisted, selection technologies. The F(2) individuals derived from cultivar crosses were screened for resistance to yellow rust at the seedling stage in greenhouses and adult stage in the field to identify DNA markers genetically linked to resistance. Five hundred and sixty ESTs were assembled into 136 contigs and 989 singletons. BlastX search results showed that 39 (29%) contigs and 96 (10%) singletons were homologous to wheat genes. The database-matched contigs and singletons were assigned to eight functional groups related to protein synthesis, photosynthesis, metabolism and energy, stress proteins, transporter proteins, protein breakdown and recycling, cell growth and division and reactive oxygen scavengers. PCR analyses with primers based on the contigs and singletons showed that the most polymorphic functional categories were photosynthesis (contigs) and metabolism and energy (singletons). EST analysis revealed considerable genetic variability among the Turkish wheat cultivars resistant and susceptible to yellow rust disease and allowed calculation of the mean genetic distance between cultivars, with the greatest similarity (0.725) being between Harmankaya99 and Sönmez2001, and the lowest (0.622) between Aytin98 and Izgi01.

A nonpolymorphic major histocompatibility complex class Ib molecule binds a large array of diverse self-peptides

PubMed Central

1994-01-01

Unlike the highly polymorphic major histocompatibility complex (MHC) class Ia molecules, which present a wide variety of peptides to T cells, it is generally assumed that the nonpolymorphic MHC class Ib molecules may have evolved to function as highly specialized receptors for the presentation of structurally unique peptides. However, a thorough biochemical analysis of one class Ib molecule, the soluble isoform of Qa-2 antigen (H-2SQ7b), has revealed that it binds a diverse array of structurally similar peptides derived from intracellular proteins in much the same manner as the classical antigen-presenting molecules. Specifically, we find that SQ7b molecules are heterodimers of heavy and light chains complexed with nonameric peptides in a 1:1:1 ratio. These peptides contain a conserved hydrophobic residue at the COOH terminus and a combination of one or more conserved residue(s) at P7 (histidine), P2 (glutamine/leucine), and/or P3 (leucine/asparagine) as anchors for binding SQ7b. 2 of 18 sequenced peptides matched cytosolic proteins (cofilin and L19 ribosomal protein), suggesting an intracellular source of the SQ7b ligands. Minimal estimates of the peptide repertoire revealed that at least 200 different naturally processed self-peptides can bind SQ7b molecules. Since Qa-2 molecules associate with a diverse array of peptides, we suggest that they function as effective presenting molecules of endogenously synthesized proteins like the class Ia molecules. PMID:8294869

Protein Assembly and Building Blocks: Beyond the Limits of the LEGO Brick Metaphor.

PubMed

Levy, Yaakov

2017-09-26

Proteins, like other biomolecules, have a modular and hierarchical structure. Various building blocks are used to construct proteins of high structural complexity and diverse functionality. In multidomain proteins, for example, domains are fused to each other in different combinations to achieve different functions. Although the LEGO brick metaphor is justified as a means of simplifying the complexity of three-dimensional protein structures, several fundamental properties (such as allostery or the induced-fit mechanism) make deviation from it necessary to respect the plasticity, softness, and cross-talk that are essential to protein function. In this work, we illustrate recently reported protein behavior in multidomain proteins that deviates from the LEGO brick analogy. While earlier studies showed that a protein domain is often unaffected by being fused to another domain or becomes more stable following the formation of a new interface between the tethered domains, destabilization due to tethering has been reported for several systems. We illustrate that tethering may sometimes result in a multidomain protein behaving as "less than the sum of its parts". We survey these cases for which structure additivity does not guarantee thermodynamic additivity. Protein destabilization due to fusion to other domains may be linked in some cases to biological function and should be taken into account when designing large assemblies.

Evaluation of Several Two-Step Scoring Functions Based on Linear Interaction Energy, Effective Ligand Size, and Empirical Pair Potentials for Prediction of Protein-Ligand Binding Geometry and Free Energy

PubMed Central

Rahaman, Obaidur; Estrada, Trilce P.; Doren, Douglas J.; Taufer, Michela; Brooks, Charles L.; Armen, Roger S.

2011-01-01

The performance of several two-step scoring approaches for molecular docking were assessed for their ability to predict binding geometries and free energies. Two new scoring functions designed for “step 2 discrimination” were proposed and compared to our CHARMM implementation of the linear interaction energy (LIE) approach using the Generalized-Born with Molecular Volume (GBMV) implicit solvation model. A scoring function S1 was proposed by considering only “interacting” ligand atoms as the “effective size” of the ligand, and extended to an empirical regression-based pair potential S2. The S1 and S2 scoring schemes were trained and five-fold cross validated on a diverse set of 259 protein-ligand complexes from the Ligand Protein Database (LPDB). The regression-based parameters for S1 and S2 also demonstrated reasonable transferability in the CSARdock 2010 benchmark using a new dataset (NRC HiQ) of diverse protein-ligand complexes. The ability of the scoring functions to accurately predict ligand geometry was evaluated by calculating the discriminative power (DP) of the scoring functions to identify native poses. The parameters for the LIE scoring function with the optimal discriminative power (DP) for geometry (step 1 discrimination) were found to be very similar to the best-fit parameters for binding free energy over a large number of protein-ligand complexes (step 2 discrimination). Reasonable performance of the scoring functions in enrichment of active compounds in four different protein target classes established that the parameters for S1 and S2 provided reasonable accuracy and transferability. Additional analysis was performed to definitively separate scoring function performance from molecular weight effects. This analysis included the prediction of ligand binding efficiencies for a subset of the CSARdock NRC HiQ dataset where the number of ligand heavy atoms ranged from 17 to 35. This range of ligand heavy atoms is where improved accuracy of predicted ligand efficiencies is most relevant to real-world drug design efforts. PMID:21644546

Evaluation of several two-step scoring functions based on linear interaction energy, effective ligand size, and empirical pair potentials for prediction of protein-ligand binding geometry and free energy.

PubMed

Rahaman, Obaidur; Estrada, Trilce P; Doren, Douglas J; Taufer, Michela; Brooks, Charles L; Armen, Roger S

2011-09-26

The performances of several two-step scoring approaches for molecular docking were assessed for their ability to predict binding geometries and free energies. Two new scoring functions designed for "step 2 discrimination" were proposed and compared to our CHARMM implementation of the linear interaction energy (LIE) approach using the Generalized-Born with Molecular Volume (GBMV) implicit solvation model. A scoring function S1 was proposed by considering only "interacting" ligand atoms as the "effective size" of the ligand and extended to an empirical regression-based pair potential S2. The S1 and S2 scoring schemes were trained and 5-fold cross-validated on a diverse set of 259 protein-ligand complexes from the Ligand Protein Database (LPDB). The regression-based parameters for S1 and S2 also demonstrated reasonable transferability in the CSARdock 2010 benchmark using a new data set (NRC HiQ) of diverse protein-ligand complexes. The ability of the scoring functions to accurately predict ligand geometry was evaluated by calculating the discriminative power (DP) of the scoring functions to identify native poses. The parameters for the LIE scoring function with the optimal discriminative power (DP) for geometry (step 1 discrimination) were found to be very similar to the best-fit parameters for binding free energy over a large number of protein-ligand complexes (step 2 discrimination). Reasonable performance of the scoring functions in enrichment of active compounds in four different protein target classes established that the parameters for S1 and S2 provided reasonable accuracy and transferability. Additional analysis was performed to definitively separate scoring function performance from molecular weight effects. This analysis included the prediction of ligand binding efficiencies for a subset of the CSARdock NRC HiQ data set where the number of ligand heavy atoms ranged from 17 to 35. This range of ligand heavy atoms is where improved accuracy of predicted ligand efficiencies is most relevant to real-world drug design efforts.

Mutant phenotypes for thousands of bacterial genes of unknown function

DOE PAGES

Price, Morgan N.; Wetmore, Kelly M.; Waters, R. Jordan; ...

2018-05-16

One-third of all protein-coding genes from bacterial genomes cannot be annotated with a function. Here, to investigate the functions of these genes, we present genome-wide mutant fitness data from 32 diverse bacteria across dozens of growth conditions. We identified mutant phenotypes for 11,779 protein-coding genes that had not been annotated with a specific function. Many genes could be associated with a specific condition because the gene affected fitness only in that condition, or with another gene in the same bacterium because they had similar mutant phenotypes. Of the poorly annotated genes, 2,316 had associations that have high confidence because theymore » are conserved in other bacteria. By combining these conserved associations with comparative genomics, we identified putative DNA repair proteins; in addition, we propose specific functions for poorly annotated enzymes and transporters and for uncharacterized protein families. Lastly, our study demonstrates the scalability of microbial genetics and its utility for improving gene annotations.« less

Mutant phenotypes for thousands of bacterial genes of unknown function

DOE Office of Scientific and Technical Information (OSTI.GOV)

Price, Morgan N.; Wetmore, Kelly M.; Waters, R. Jordan

One-third of all protein-coding genes from bacterial genomes cannot be annotated with a function. Here, to investigate the functions of these genes, we present genome-wide mutant fitness data from 32 diverse bacteria across dozens of growth conditions. We identified mutant phenotypes for 11,779 protein-coding genes that had not been annotated with a specific function. Many genes could be associated with a specific condition because the gene affected fitness only in that condition, or with another gene in the same bacterium because they had similar mutant phenotypes. Of the poorly annotated genes, 2,316 had associations that have high confidence because theymore » are conserved in other bacteria. By combining these conserved associations with comparative genomics, we identified putative DNA repair proteins; in addition, we propose specific functions for poorly annotated enzymes and transporters and for uncharacterized protein families. Lastly, our study demonstrates the scalability of microbial genetics and its utility for improving gene annotations.« less

Emergence of Complexity in Protein Functions and Metabolic Networks

NASA Technical Reports Server (NTRS)

Pohorille, Andzej

2009-01-01

In modern organisms proteins perform a majority of cellular functions, such as chemical catalysis, energy transduction and transport of material across cell walls. Although great strides have been made towards understanding protein evolution, a meaningful extrapolation from contemporary proteins to their earliest ancestors is virtually impossible. In an alternative approach, the origin of water-soluble proteins was probed through the synthesis of very large libraries of random amino acid sequences and subsequently subjecting them to in vitro evolution. In combination with computer modeling and simulations, these experiments allow us to address a number of fundamental questions about the origins of proteins. Can functionality emerge from random sequences of proteins? How did the initial repertoire of functional proteins diversify to facilitate new functions? Did this diversification proceed primarily through drawing novel functionalities from random sequences or through evolution of already existing proto-enzymes? Did protein evolution start from a pool of proteins defined by a frozen accident and other collections of proteins could start a different evolutionary pathway? Although we do not have definitive answers to these questions, important clues have been uncovered. Considerable progress has been also achieved in understanding the origins of membrane proteins. We will address this issue in the example of ion channels - proteins that mediate transport of ions across cell walls. Remarkably, despite overall complexity of these proteins in contemporary cells, their structural motifs are quite simple, with -helices being most common. By combining results of experimental and computer simulation studies on synthetic models and simple, natural channels, I will show that, even though architectures of membrane proteins are not nearly as diverse as those of water-soluble proteins, they are sufficiently flexible to adapt readily to the functional demands arising during evolution.

Bioinformatic and Comparative Localization of Rab Proteins Reveals Functional Insights into the Uncharacterized GTPases Ypt10p and Ypt11p†

PubMed Central

Buvelot Frei, Stéphanie; Rahl, Peter B.; Nussbaum, Maria; Briggs, Benjamin J.; Calero, Monica; Janeczko, Stephanie; Regan, Andrew D.; Chen, Catherine Z.; Barral, Yves; Whittaker, Gary R.; Collins, Ruth N.

2006-01-01

A striking characteristic of a Rab protein is its steady-state localization to the cytosolic surface of a particular subcellular membrane. In this study, we have undertaken a combined bioinformatic and experimental approach to examine the evolutionary conservation of Rab protein localization. A comprehensive primary sequence classification shows that 10 out of the 11 Rab proteins identified in the yeast (Saccharomyces cerevisiae) genome can be grouped within a major subclass, each comprising multiple Rab orthologs from diverse species. We compared the locations of individual yeast Rab proteins with their localizations following ectopic expression in mammalian cells. Our results suggest that green fluorescent protein-tagged Rab proteins maintain localizations across large evolutionary distances and that the major known player in the Rab localization pathway, mammalian Rab-GDI, is able to function in yeast. These findings enable us to provide insight into novel gene functions and classify the uncharacterized Rab proteins Ypt10p (YBR264C) as being involved in endocytic function and Ypt11p (YNL304W) as being localized to the endoplasmic reticulum, where we demonstrate it is required for organelle inheritance. PMID:16980630

The Diverse Structures and Functions of Surfactant Proteins.

PubMed

Schor, Marieke; Reid, Jack L; MacPhee, Cait E; Stanley-Wall, Nicola R

2016-07-01

Surface tension at liquid-air interfaces is a major barrier that needs to be surmounted by a wide range of organisms; surfactant and interfacially active proteins have evolved for this purpose. Although these proteins are essential for a variety of biological processes, our understanding of how they elicit their function has been limited. However, with the recent determination of high-resolution 3D structures of several examples, we have gained insight into the distinct shapes and mechanisms that have evolved to confer interfacial activity. It is now a matter of harnessing this information, and these systems, for biotechnological purposes. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Ice-Binding Proteins and Their Function.

PubMed

Bar Dolev, Maya; Braslavsky, Ido; Davies, Peter L

2016-06-02

Ice-binding proteins (IBPs) are a diverse class of proteins that assist organism survival in the presence of ice in cold climates. They have different origins in many organisms, including bacteria, fungi, algae, diatoms, plants, insects, and fish. This review covers the gamut of IBP structures and functions and the common features they use to bind ice. We discuss mechanisms by which IBPs adsorb to ice and interfere with its growth, evidence for their irreversible association with ice, and methods for enhancing the activity of IBPs. The applications of IBPs in the food industry, in cryopreservation, and in other technologies are vast, and we chart out some possibilities.

Applications of Functional Amyloids from Fungi: Surface Modification by Class I Hydrophobins.

PubMed

Piscitelli, Alessandra; Cicatiello, Paola; Gravagnuolo, Alfredo Maria; Sorrentino, Ilaria; Pezzella, Cinzia; Giardina, Paola

2017-06-26

Class I hydrophobins produced from fungi are amongst the first proteins recognized as functional amyloids. They are amphiphilic proteins involved in the formation of aerial structures such as spores or fruiting bodies. They form chemically robust layers which can only be dissolved in strong acids. These layers adhere to different surfaces, changing their wettability, and allow the binding of other proteins. Herein, the modification of diverse types of surfaces with Class I hydrophobins is reported, highlighting the applications of the coated surfaces. Indeed, these coatings can be exploited in several fields, spanning from biomedical to industrial applications, which include biosensing and textile manufacturing.

Cloning and analysis of 16 Rab genes from macronuclear DNA of Euplotes octocarinatus.

PubMed

Zhi, Hui; Wang, Wei; Li, Lingyan; Chai, Baofeng; Sun, Yonghua; Liang, Aihua

2005-08-01

Rab proteins belong to the largest family of the Ras superfamily of small GTPase that play an important role in intracellular vesicular traffic. So far, almost 60 members of Rab family have been identified in mammalian cells. To further study the diversity and function of Rab protein in evolution, unicellular protozoa ciliates, Euplotes octocarinatus, were used in this study, Rab genes were screened by PCR method from macronuclear DNA of E. octocarinatus. Sixteen Rab genes were obtained. They share 87.6-99.5% identities. Highly conserved GTP-binding domains were found. There are some hot regions that diverse sharply in these genes as well.

Empirical scoring functions for advanced protein-ligand docking with PLANTS.

PubMed

Korb, Oliver; Stützle, Thomas; Exner, Thomas E

2009-01-01

In this paper we present two empirical scoring functions, PLANTS(CHEMPLP) and PLANTS(PLP), designed for our docking algorithm PLANTS (Protein-Ligand ANT System), which is based on ant colony optimization (ACO). They are related, regarding their functional form, to parts of already published scoring functions and force fields. The parametrization procedure described here was able to identify several parameter settings showing an excellent performance for the task of pose prediction on two test sets comprising 298 complexes in total. Up to 87% of the complexes of the Astex diverse set and 77% of the CCDC/Astex clean listnc (noncovalently bound complexes of the clean list) could be reproduced with root-mean-square deviations of less than 2 A with respect to the experimentally determined structures. A comparison with the state-of-the-art docking tool GOLD clearly shows that this is, especially for the druglike Astex diverse set, an improvement in pose prediction performance. Additionally, optimized parameter settings for the search algorithm were identified, which can be used to balance pose prediction reliability and search speed.

GIANT 2.0: genome-scale integrated analysis of gene networks in tissues.

PubMed

Wong, Aaron K; Krishnan, Arjun; Troyanskaya, Olga G

2018-05-25

GIANT2 (Genome-wide Integrated Analysis of gene Networks in Tissues) is an interactive web server that enables biomedical researchers to analyze their proteins and pathways of interest and generate hypotheses in the context of genome-scale functional maps of human tissues. The precise actions of genes are frequently dependent on their tissue context, yet direct assay of tissue-specific protein function and interactions remains infeasible in many normal human tissues and cell-types. With GIANT2, researchers can explore predicted tissue-specific functional roles of genes and reveal changes in those roles across tissues, all through interactive multi-network visualizations and analyses. Additionally, the NetWAS approach available through the server uses tissue-specific/cell-type networks predicted by GIANT2 to re-prioritize statistical associations from GWAS studies and identify disease-associated genes. GIANT2 predicts tissue-specific interactions by integrating diverse functional genomics data from now over 61 400 experiments for 283 diverse tissues and cell-types. GIANT2 does not require any registration or installation and is freely available for use at http://giant-v2.princeton.edu.

Protein-Protein Interactions in Clathrin Vesicular Assembly: Radial Distribution of Evolutionary Constraints in Interfaces

PubMed Central

Gadkari, Rupali A.; Srinivasan, Narayanaswamy

2012-01-01

In eukaryotic organisms clathrin-coated vesicles are instrumental in the processes of endocytosis as well as intracellular protein trafficking. Hence, it is important to understand how these vesicles have evolved across eukaryotes, to carry cargo molecules of varied shapes and sizes. The intricate nature and functional diversity of the vesicles are maintained by numerous interacting protein partners of the vesicle system. However, to delineate functionally important residues participating in protein-protein interactions of the assembly is a daunting task as there are no high-resolution structures of the intact assembly available. The two cryoEM structures closely representing intact assembly were determined at very low resolution and provide positions of Cα atoms alone. In the present study, using the method developed by us earlier, we predict the protein-protein interface residues in clathrin assembly, taking guidance from the available low-resolution structures. The conservation status of these interfaces when investigated across eukaryotes, revealed a radial distribution of evolutionary constraints, i.e., if the members of the clathrin vesicular assembly can be imagined to be arranged in spherical manner, the cargo being at the center and clathrins being at the periphery, the detailed phylogenetic analysis of these members of the assembly indicated high-residue variation in the members of the assembly closer to the cargo while high conservation was noted in clathrins and in other proteins at the periphery of the vesicle. This points to the strategy adopted by the nature to package diverse proteins but transport them through a highly conserved mechanism. PMID:22384024

Identifying functionally informative evolutionary sequence profiles.

PubMed

Gil, Nelson; Fiser, Andras

2018-04-15

Multiple sequence alignments (MSAs) can provide essential input to many bioinformatics applications, including protein structure prediction and functional annotation. However, the optimal selection of sequences to obtain biologically informative MSAs for such purposes is poorly explored, and has traditionally been performed manually. We present Selection of Alignment by Maximal Mutual Information (SAMMI), an automated, sequence-based approach to objectively select an optimal MSA from a large set of alternatives sampled from a general sequence database search. The hypothesis of this approach is that the mutual information among MSA columns will be maximal for those MSAs that contain the most diverse set possible of the most structurally and functionally homogeneous protein sequences. SAMMI was tested to select MSAs for functional site residue prediction by analysis of conservation patterns on a set of 435 proteins obtained from protein-ligand (peptides, nucleic acids and small substrates) and protein-protein interaction databases. Availability and implementation: A freely accessible program, including source code, implementing SAMMI is available at https://github.com/nelsongil92/SAMMI.git. andras.fiser@einstein.yu.edu. Supplementary data are available at Bioinformatics online.

A structural analysis of the AAA+ domains in Saccharomyces cerevisiae cytoplasmic dynein

PubMed Central

Gleave, Emma S.; Schmidt, Helgo; Carter, Andrew P.

2014-01-01

Dyneins are large protein complexes that act as microtubule based molecular motors. The dynein heavy chain contains a motor domain which is a member of the AAA+ protein family (ATPases Associated with diverse cellular Activities). Proteins of the AAA+ family show a diverse range of functionalities, but share a related core AAA+ domain, which often assembles into hexameric rings. Dynein is unusual because it has all six AAA+ domains linked together, in one long polypeptide. The dynein motor domain generates movement by coupling ATP driven conformational changes in the AAA+ ring to the swing of a motile element called the linker. Dynein binds to its microtubule track via a long antiparallel coiled-coil stalk that emanates from the AAA+ ring. Recently the first high resolution structures of the dynein motor domain were published. Here we provide a detailed structural analysis of the six AAA+ domains using our Saccharomycescerevisiae crystal structure. We describe how structural similarities in the dynein AAA+ domains suggest they share a common evolutionary origin. We analyse how the different AAA+ domains have diverged from each other. We discuss how this is related to the function of dynein as a motor protein and how the AAA+ domains of dynein compare to those of other AAA+ proteins. PMID:24680784

The Importance of Being Tyrosine: Lessons in Molecular Recognition from Minimalist Synthetic Binding Proteins

PubMed Central

Koide, Shohei; Sidhu, Sachdev S.

2010-01-01

Summary Combinatorial libraries built with severely restricted chemical diversity have yielded highly functional synthetic binding proteins. Structural analyses of these minimalist binding sites have revealed the dominant role of large tyrosine residues for mediating molecular contacts and of small serine/glycine residues for providing space and flexibility. The concept of using limited residue types to construct optimized binding proteins mirrors findings in the field of small molecule drug development, where it has been proposed that most drugs are built from a limited set of side chains presented by diverse frameworks. The physicochemical properties of tyrosine make it the amino acid that is most effective for mediating molecular recognition, and protein engineers have taken advantage of these characteristics to build tyrosine-rich protein binding sites that outperform natural proteins in terms of affinity and specificity. Knowledge from preceding studies can be used to improve current designs, and thus, synthetic protein libraries will continue to evolve and improve. In the near future, it seems likely that synthetic binding proteins will supersede natural antibodies for most purposes, and moreover, synthetic proteins will enable many new applications beyond the scope of natural proteins. PMID:19298050

A quantitative chaperone interaction network reveals the architecture of cellular protein homeostasis pathways

PubMed Central

Taipale, Mikko; Tucker, George; Peng, Jian; Krykbaeva, Irina; Lin, Zhen-Yuan; Larsen, Brett; Choi, Hyungwon; Berger, Bonnie; Gingras, Anne-Claude; Lindquist, Susan

2014-01-01

Chaperones are abundant cellular proteins that promote the folding and function of their substrate proteins (clients). In vivo, chaperones also associate with a large and diverse set of co-factors (co-chaperones) that regulate their specificity and function. However, how these co-chaperones regulate protein folding and whether they have chaperone-independent biological functions is largely unknown. We have combined mass spectrometry and quantitative high-throughput LUMIER assays to systematically characterize the chaperone/co-chaperone/client interaction network in human cells. We uncover hundreds of novel chaperone clients, delineate their participation in specific co-chaperone complexes, and establish a surprisingly distinct network of protein/protein interactions for co-chaperones. As a salient example of the power of such analysis, we establish that NUDC family co-chaperones specifically associate with structurally related but evolutionarily distinct β-propeller folds. We provide a framework for deciphering the proteostasis network, its regulation in development and disease, and expand the use of chaperones as sensors for drug/target engagement. PMID:25036637

ORF phage display to identify cellular proteins with different functions.

PubMed

Li, Wei

2012-09-01

Open reading frame (ORF) phage display is a new branch of phage display aimed at improving its efficiency to identify cellular proteins with specific binding or functional activities. Despite the success of phage display with antibody libraries and random peptide libraries, phage display with cDNA libraries of cellular proteins identifies a high percentage of non-ORF clones encoding unnatural short peptides with minimal biological implications. This is mainly because of the uncontrollable reading frames of cellular proteins in conventional cDNA libraries. ORF phage display solves this problem by eliminating non-ORF clones to generate ORF cDNA libraries. Here I summarize the procedures of ORF phage display, discuss the factors influencing its efficiency, present examples of its versatile applications, and highlight evidence of its capability of identifying biologically relevant cellular proteins. ORF phage display coupled with different selection strategies is capable of delineating diverse functions of cellular proteins with unique advantages. Copyright © 2012 Elsevier Inc. All rights reserved.

Evolution of an ancient protein function involved in organized multicellularity in animals

PubMed Central

Anderson, Douglas P; Whitney, Dustin S; Hanson-Smith, Victor; Woznica, Arielle; Campodonico-Burnett, William; Volkman, Brian F; King, Nicole; Thornton, Joseph W; Prehoda, Kenneth E

2016-01-01

To form and maintain organized tissues, multicellular organisms orient their mitotic spindles relative to neighboring cells. A molecular complex scaffolded by the GK protein-interaction domain (GKPID) mediates spindle orientation in diverse animal taxa by linking microtubule motor proteins to a marker protein on the cell cortex localized by external cues. Here we illuminate how this complex evolved and commandeered control of spindle orientation from a more ancient mechanism. The complex was assembled through a series of molecular exploitation events, one of which – the evolution of GKPID’s capacity to bind the cortical marker protein – can be recapitulated by reintroducing a single historical substitution into the reconstructed ancestral GKPID. This change revealed and repurposed an ancient molecular surface that previously had a radically different function. We show how the physical simplicity of this binding interface enabled the evolution of a new protein function now essential to the biological complexity of many animals. DOI: http://dx.doi.org/10.7554/eLife.10147.001 PMID:26740169

Improving protein delivery of fibroblast growth factor-2 from bacterial inclusion bodies used as cell culture substrates.

PubMed

Seras-Franzoso, Joaquin; Peebo, Karl; García-Fruitós, Elena; Vázquez, Esther; Rinas, Ursula; Villaverde, Antonio

2014-03-01

Bacterial inclusion bodies (IBs) have recently been used to generate biocompatible cell culture interfaces, with diverse effects on cultured cells such as cell adhesion enhancement, stimulation of cell growth or induction of mesenchymal stem cell differentiation. Additionally, novel applications of IBs as sustained protein delivery systems with potential applications in regenerative medicine have been successfully explored. In this scenario, with IBs gaining significance in the biomedical field, the fine tuning of this functional biomaterial is crucial. In this work, the effect of temperature on fibroblast growth factor-2 (FGF-2) IB production and performance has been evaluated. FGF-2 was overexpressed in Escherichia coli at 25 and 37 °C, producing IBs with differences in size, particle structure and biological activity. Cell culture topographies made with FGF-2 IBs biofabricated at 25 °C showed higher levels of biological activity as well as a looser supramolecular structure, enabling a higher protein release from the particles. In addition, the controlled use of FGF-2 protein particles enabled the generation of functional topographies with multiple biological activities being effective on diverse cell types. Copyright © 2013 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

A High Throughput Protein Microarray Approach to Classify HIV Monoclonal Antibodies and Variant Antigens

PubMed Central

Dotsey, Emmanuel Y.; Gorlani, Andrea; Ingale, Sampat; Achenbach, Chad J.; Forthal, Donald N.; Felgner, Philip L.; Gach, Johannes S.

2015-01-01

In recent years, high throughput discovery of human recombinant monoclonal antibodies (mAbs) has been applied to greatly advance our understanding of the specificity, and functional activity of antibodies against HIV. Thousands of antibodies have been generated and screened in functional neutralization assays, and antibodies associated with cross-strain neutralization and passive protection in primates, have been identified. To facilitate this type of discovery, a high throughput-screening tool is needed to accurately classify mAbs, and their antigen targets. In this study, we analyzed and evaluated a prototype microarray chip comprised of the HIV-1 recombinant proteins gp140, gp120, gp41, and several membrane proximal external region peptides. The protein microarray analysis of 11 HIV-1 envelope-specific mAbs revealed diverse binding affinities and specificities across clades. Half maximal effective concentrations, generated by our chip analysis, correlated significantly (P<0.0001) with concentrations from ELISA binding measurements. Polyclonal immune responses in plasma samples from HIV-1 infected subjects exhibited different binding patterns, and reactivity against printed proteins. Examining the totality of the specificity of the humoral response in this way reveals the exquisite diversity, and specificity of the humoral response to HIV. PMID:25938510

Functional diversification and specialization of cytosolic 70-kDa heat shock proteins.

PubMed

McCallister, Chelsea; Siracusa, Matthew C; Shirazi, Farzaneh; Chalkia, Dimitra; Nikolaidis, Nikolas

2015-03-20

A fundamental question in molecular evolution is how protein functional differentiation alters the ability of cells and organisms to cope with stress and survive. To answer this question we used two paralogous Hsp70s from mouse and explored whether these highly similar cytosolic molecular chaperones, which apart their temporal expression have been considered functionally interchangeable, are differentiated with respect to their lipid-binding function. We demonstrate that the two proteins bind to diverse lipids with different affinities and therefore are functionally specialized. The observed lipid-binding patterns may be related with the ability of both Hsp70s to induce cell death by binding to a particular plasma-membrane lipid, and the potential of only one of them to promote cell survival by binding to a specific lysosomal-membrane lipid. These observations reveal that two seemingly identical proteins differentially modulate cellular adaptation and survival by having acquired specialized functions via sequence divergence. Therefore, this study provides an evolutionary paradigm, where promiscuity, specificity, sub- and neo-functionalization orchestrate one of the most conserved systems in nature, the cellular stress-response.

Resolving protein structure-function-binding site relationships from a binding site similarity network perspective.

PubMed

Mudgal, Richa; Srinivasan, Narayanaswamy; Chandra, Nagasuma

2017-07-01

Functional annotation is seldom straightforward with complexities arising due to functional divergence in protein families or functional convergence between non-homologous protein families, leading to mis-annotations. An enzyme may contain multiple domains and not all domains may be involved in a given function, adding to the complexity in function annotation. To address this, we use binding site information from bound cognate ligands and catalytic residues, since it can help in resolving fold-function relationships at a finer level and with higher confidence. A comprehensive database of 2,020 fold-function-binding site relationships has been systematically generated. A network-based approach is employed to capture the complexity in these relationships, from which different types of associations are deciphered, that identify versatile protein folds performing diverse functions, same function associated with multiple folds and one-to-one relationships. Binding site similarity networks integrated with fold, function, and ligand similarity information are generated to understand the depth of these relationships. Apart from the observed continuity in the functional site space, network properties of these revealed versatile families with topologically different or dissimilar binding sites and structural families that perform very similar functions. As a case study, subtle changes in the active site of a set of evolutionarily related superfamilies are studied using these networks. Tracing of such similarities in evolutionarily related proteins provide clues into the transition and evolution of protein functions. Insights from this study will be helpful in accurate and reliable functional annotations of uncharacterized proteins, poly-pharmacology, and designing enzymes with new functional capabilities. Proteins 2017; 85:1319-1335. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Evolutionary tradeoffs in cellular composition across diverse bacteria

PubMed Central

Kempes, Christopher P; Wang, Lawrence; Amend, Jan P; Doyle, John; Hoehler, Tori

2016-01-01

One of the most important classic and contemporary interests in biology is the connection between cellular composition and physiological function. Decades of research have allowed us to understand the detailed relationship between various cellular components and processes for individual species, and have uncovered common functionality across diverse species. However, there still remains the need for frameworks that can mechanistically predict the tradeoffs between cellular functions and elucidate and interpret average trends across species. Here we provide a comprehensive analysis of how cellular composition changes across the diversity of bacteria as connected with physiological function and metabolism, spanning five orders of magnitude in body size. We present an analysis of the trends with cell volume that covers shifts in genomic, protein, cellular envelope, RNA and ribosomal content. We show that trends in protein content are more complex than a simple proportionality with the overall genome size, and that the number of ribosomes is simply explained by cross-species shifts in biosynthesis requirements. Furthermore, we show that the largest and smallest bacteria are limited by physical space requirements. At the lower end of size, cell volume is dominated by DNA and protein content—the requirement for which predicts a lower limit on cell size that is in good agreement with the smallest observed bacteria. At the upper end of bacterial size, we have identified a point at which the number of ribosomes required for biosynthesis exceeds available cell volume. Between these limits we are able to discuss systematic and dramatic shifts in cellular composition. Much of our analysis is connected with the basic energetics of cells where we show that the scaling of metabolic rate is surprisingly superlinear with all cellular components. PMID:27046336

Biofilm Matrix Proteins.

PubMed

Fong, Jiunn N C; Yildiz, Fitnat H

2015-04-01

Proteinaceous components of the biofilm matrix include secreted extracellular proteins, cell surface adhesins, and protein subunits of cell appendages such as flagella and pili. Biofilm matrix proteins play diverse roles in biofilm formation and dissolution. They are involved in attaching cells to surfaces, stabilizing the biofilm matrix via interactions with exopolysaccharide and nucleic acid components, developing three-dimensional biofilm architectures, and dissolving biofilm matrix via enzymatic degradation of polysaccharides, proteins, and nucleic acids. In this article, we will review functions of matrix proteins in a selected set of microorganisms, studies of the matrix proteomes of Vibrio cholerae and Pseudomonas aeruginosa, and roles of outer membrane vesicles and of nucleoid-binding proteins in biofilm formation.

Deviation of the typical AAA substrate-threading pore prevents fatal protein degradation in yeast Cdc48.

PubMed

Esaki, Masatoshi; Islam, Md Tanvir; Tani, Naoki; Ogura, Teru

2017-07-14

Yeast Cdc48 is a well-conserved, essential chaperone of ATPases associated with diverse cellular activity (AAA) proteins, which recognizes substrate proteins and modulates their conformations to carry out many cellular processes. However, the fundamental mechanisms underlying the diverse pivotal roles of Cdc48 remain unknown. Almost all AAA proteins form a ring-shaped structure with a conserved aromatic amino acid residue that is essential for proper function. The threading mechanism hypothesis suggests that this residue guides the intrusion of substrate proteins into a narrow pore of the AAA ring, thereby becoming unfolded. By contrast, the aromatic residue in one of the two AAA rings of Cdc48 has been eliminated through evolution. Here, we show that artificial retrieval of this aromatic residue in Cdc48 is lethal, and essential features to support the threading mechanism are required to exhibit the lethal phenotype. In particular, genetic and biochemical analyses of the Cdc48 lethal mutant strongly suggested that when in complex with the 20S proteasome, essential proteins are abnormally forced to thread through the Cdc48 pore to become degraded, which was not detected in wild-type Cdc48. Thus, the widely applicable threading model is less effective for wild-type Cdc48; rather, Cdc48 might function predominantly through an as-yet-undetermined mechanism.

An expanding universe of small proteins.

PubMed

Hobbs, Errett C; Fontaine, Fanette; Yin, Xuefeng; Storz, Gisela

2011-04-01

Historically, small proteins (sproteins) of less than 50 amino acids, in their final processed forms or genetically encoded as such, have been understudied. However, both serendipity and more recent focused efforts have led to the identification of a number of new sproteins in both Gram-negative and Gram-positive bacteria. Increasing evidence demonstrates that sproteins participate in a wide array of cellular processes and exhibit great diversity in their mechanisms of action, yet general principles of sprotein function are emerging. This review highlights examples of sproteins that participate in cell signaling, act as antibiotics and toxins, and serve as structural proteins. We also describe roles for sproteins in detecting and altering membrane features, acting as chaperones, and regulating the functions of larger proteins. Published by Elsevier Ltd.

Extending CATH: increasing coverage of the protein structure universe and linking structure with function

PubMed Central

Cuff, Alison L.; Sillitoe, Ian; Lewis, Tony; Clegg, Andrew B.; Rentzsch, Robert; Furnham, Nicholas; Pellegrini-Calace, Marialuisa; Jones, David; Thornton, Janet; Orengo, Christine A.

2011-01-01

CATH version 3.3 (class, architecture, topology, homology) contains 128 688 domains, 2386 homologous superfamilies and 1233 fold groups, and reflects a major focus on classifying structural genomics (SG) structures and transmembrane proteins, both of which are likely to add structural novelty to the database and therefore increase the coverage of protein fold space within CATH. For CATH version 3.4 we have significantly improved the presentation of sequence information and associated functional information for CATH superfamilies. The CATH superfamily pages now reflect both the functional and structural diversity within the superfamily and include structural alignments of close and distant relatives within the superfamily, annotated with functional information and details of conserved residues. A significantly more efficient search function for CATH has been established by implementing the search server Solr (http://lucene.apache.org/solr/). The CATH v3.4 webpages have been built using the Catalyst web framework. PMID:21097779

Apoferritin fibers: a new template for 1D fluorescent hybrid nanostructures

NASA Astrophysics Data System (ADS)

Jurado, Rocío; Castello, Fabio; Bondia, Patricia; Casado, Santiago; Flors, Cristina; Cuesta, Rafael; Domínguez-Vera, José M.; Orte, Angel; Gálvez, Natividad

2016-05-01

Recently, research in the field of protein amyloid fibers has gained great attention due to the use of these materials as nanoscale templates for the construction of functional hybrid materials. The formation of apoferritin amyloid-like protein fibers is demonstrated herein for the first time. The morphology, size and stiffness of these one-dimensional structures are comparable to the fibers formed by β-lactoglobulin, a protein frequently used as a model in the study of amyloid-like fibrillar proteins. Nanometer-sized globular apoferritin is capable of self-assembling to form 1D micrometer-sized structures after being subjected to a heating process. Depending on the experimental conditions, fibers with different morphologies and sizes are obtained. The wire-like protein structure is rich in functional groups and allows chemical functionalization with diverse quantum dots (QD), as well as with different Alexa Fluor (AF) dyes, leading to hybrid fluorescent fibers with variable emission wavelengths, from green to near infrared, depending on the QD and AFs coupled. For fibers containing the pair AF488 and AF647, efficient fluorescence energy transfer from the covalently coupled donor (AF488) to acceptor tags (AF647) takes place. Apoferritin fibers are proposed here as a new promising template for obtaining hybrid functional materials.Recently, research in the field of protein amyloid fibers has gained great attention due to the use of these materials as nanoscale templates for the construction of functional hybrid materials. The formation of apoferritin amyloid-like protein fibers is demonstrated herein for the first time. The morphology, size and stiffness of these one-dimensional structures are comparable to the fibers formed by β-lactoglobulin, a protein frequently used as a model in the study of amyloid-like fibrillar proteins. Nanometer-sized globular apoferritin is capable of self-assembling to form 1D micrometer-sized structures after being subjected to a heating process. Depending on the experimental conditions, fibers with different morphologies and sizes are obtained. The wire-like protein structure is rich in functional groups and allows chemical functionalization with diverse quantum dots (QD), as well as with different Alexa Fluor (AF) dyes, leading to hybrid fluorescent fibers with variable emission wavelengths, from green to near infrared, depending on the QD and AFs coupled. For fibers containing the pair AF488 and AF647, efficient fluorescence energy transfer from the covalently coupled donor (AF488) to acceptor tags (AF647) takes place. Apoferritin fibers are proposed here as a new promising template for obtaining hybrid functional materials. Electronic supplementary information (ESI) available: TEM images of ferritin protein fiber formation, and apoferritin after 18 days of heat treatment; FLIM-PIE technique details; fluorescence emission spectra of apoferritin and β-lactoglobulin fibers functionalized with different QDs. See DOI: 10.1039/c6nr01044j

The organization and contribution of helicases to RNA splicing.

PubMed

De, Inessa; Schmitzová, Jana; Pena, Vladimir

2016-01-01

Splicing is an essential step of gene expression. It occurs in two consecutive chemical reactions catalyzed by a large protein-RNA complex named the spliceosome. Assembled on the pre-mRNA substrate from five small nuclear proteins, the spliceosome acts as a protein-controlled ribozyme to catalyze the two reactions and finally dissociates into its components, which are re-used for a new round of splicing. Upon following this cyclic pathway, the spliceosome undergoes numerous intermediate stages that differ in composition as well as in their internal RNA-RNA and RNA-protein contacts. The driving forces and control mechanisms of these remodeling processes are provided by specific molecular motors called RNA helicases. While eight spliceosomal helicases are present in all organisms, higher eukaryotes contain five additional ones potentially required to drive a more intricate splicing pathway and link it to an RNA metabolism of increasing complexity. Spliceosomal helicases exhibit a notable structural diversity in their accessory domains and overall architecture, in accordance with the diversity of their task-specific functions. This review summarizes structure-function knowledge about all spliceosomal helicases, including the latter five, which traditionally are treated separately from the conserved ones. The implications of the structural characteristics of helicases for their functions, as well as for their structural communication within the multi-subunits environment of the spliceosome, are pointed out. © 2016 Wiley Periodicals, Inc.

Proteome labelling and protein identification in specific tissues and at specific developmental stages in an animal

PubMed Central

Elliott, Thomas S.; Townsley, Fiona M.; Bianco, Ambra; Ernst, Russell J.; Sachdeva, Amit; Elsässer, Simon J.; Davis, Lloyd; Lang, Kathrin; Pisa, Rudolf; Greiss, Sebastian.; Lilley, Kathryn S.; Chin, Jason W.

2014-01-01

Identifying the proteins synthesized in defined cells at specific times in an animal will facilitate the study of cellular functions and dynamic processes. Here we introduce stochastic orthogonal recoding of translation with chemoselective modification (SORT-M) to address this challenge. SORT-M involves modifying cells to express an orthogonal aminoacyl-tRNA synthetase/tRNA pair to enable the incorporation of chemically modifiable analogs of amino acids at diverse sense codons in cells in rich media. We apply SORT-M to Drosophila melanogaster fed standard food to label and image proteins in specific tissues at precise developmental stages with diverse chemistries, including cyclopropene-tetrazine inverse electron demand Diels-Alder cycloaddition reactions. We also use SORT-M to identify proteins synthesized in germ cells of the fly ovary without dissection. SORT-M will facilitate the definition of proteins synthesized in specific sets of cells to study development, and learning and memory in flies, and may be extended to other animals. PMID:24727715

GEneSTATION 1.0: a synthetic resource of diverse evolutionary and functional genomic data for studying the evolution of pregnancy-associated tissues and phenotypes

PubMed Central

Kim, Mara; Cooper, Brian A.; Venkat, Rohit; Phillips, Julie B.; Eidem, Haley R.; Hirbo, Jibril; Nutakki, Sashank; Williams, Scott M.; Muglia, Louis J.; Capra, J. Anthony; Petren, Kenneth; Abbot, Patrick; Rokas, Antonis; McGary, Kriston L.

2016-01-01

Mammalian gestation and pregnancy are fast evolving processes that involve the interaction of the fetal, maternal and paternal genomes. Version 1.0 of the GEneSTATION database (http://genestation.org) integrates diverse types of omics data across mammals to advance understanding of the genetic basis of gestation and pregnancy-associated phenotypes and to accelerate the translation of discoveries from model organisms to humans. GEneSTATION is built using tools from the Generic Model Organism Database project, including the biology-aware database CHADO, new tools for rapid data integration, and algorithms that streamline synthesis and user access. GEneSTATION contains curated life history information on pregnancy and reproduction from 23 high-quality mammalian genomes. For every human gene, GEneSTATION contains diverse evolutionary (e.g. gene age, population genetic and molecular evolutionary statistics), organismal (e.g. tissue-specific gene and protein expression, differential gene expression, disease phenotype), and molecular data types (e.g. Gene Ontology Annotation, protein interactions), as well as links to many general (e.g. Entrez, PubMed) and pregnancy disease-specific (e.g. PTBgene, dbPTB) databases. By facilitating the synthesis of diverse functional and evolutionary data in pregnancy-associated tissues and phenotypes and enabling their quick, intuitive, accurate and customized meta-analysis, GEneSTATION provides a novel platform for comprehensive investigation of the function and evolution of mammalian pregnancy. PMID:26567549

Discovering Conformational Sub-States Relevant to Protein Function

PubMed Central

Ramanathan, Arvind; Savol, Andrej J.; Langmead, Christopher J.; Agarwal, Pratul K.; Chennubhotla, Chakra S.

2011-01-01

Background Internal motions enable proteins to explore a range of conformations, even in the vicinity of native state. The role of conformational fluctuations in the designated function of a protein is widely debated. Emerging evidence suggests that sub-groups within the range of conformations (or sub-states) contain properties that may be functionally relevant. However, low populations in these sub-states and the transient nature of conformational transitions between these sub-states present significant challenges for their identification and characterization. Methods and Findings To overcome these challenges we have developed a new computational technique, quasi-anharmonic analysis (QAA). QAA utilizes higher-order statistics of protein motions to identify sub-states in the conformational landscape. Further, the focus on anharmonicity allows identification of conformational fluctuations that enable transitions between sub-states. QAA applied to equilibrium simulations of human ubiquitin and T4 lysozyme reveals functionally relevant sub-states and protein motions involved in molecular recognition. In combination with a reaction pathway sampling method, QAA characterizes conformational sub-states associated with cis/trans peptidyl-prolyl isomerization catalyzed by the enzyme cyclophilin A. In these three proteins, QAA allows identification of conformational sub-states, with critical structural and dynamical features relevant to protein function. Conclusions Overall, QAA provides a novel framework to intuitively understand the biophysical basis of conformational diversity and its relevance to protein function. PMID:21297978

Fe-S Cluster Hsp70 Chaperones: The ATPase Cycle and Protein Interactions.

PubMed

Dutkiewicz, Rafal; Nowak, Malgorzata; Craig, Elizabeth A; Marszalek, Jaroslaw

2017-01-01

Hsp70 chaperones and their obligatory J-protein cochaperones function together in many cellular processes. Via cycles of binding to short stretches of exposed amino acids on substrate proteins, Hsp70/J-protein chaperones not only facilitate protein folding but also drive intracellular protein transport, biogenesis of cellular structures, and disassembly of protein complexes. The biogenesis of iron-sulfur (Fe-S) clusters is one of the critical cellular processes that require Hsp70/J-protein action. Fe-S clusters are ubiquitous cofactors critical for activity of proteins performing diverse functions in, for example, metabolism, RNA/DNA transactions, and environmental sensing. This biogenesis process can be divided into two sequential steps: first, the assembly of an Fe-S cluster on a conserved scaffold protein, and second, the transfer of the cluster from the scaffold to a recipient protein. The second step involves Hsp70/J-protein chaperones. Via binding to the scaffold, chaperones enable cluster transfer to recipient proteins. In eukaryotic cells mitochondria have a key role in Fe-S cluster biogenesis. In this review, we focus on methods that enabled us to dissect protein interactions critical for the function of Hsp70/J-protein chaperones in the mitochondrial process of Fe-S cluster biogenesis in the yeast Saccharomyces cerevisiae. © 2017 Elsevier Inc. All rights reserved.

Fungal hemolysins

PubMed Central

Nayak, Ajay P.; Green, Brett J.; Beezhold, Donald H.

2015-01-01

Hemolysins are a class of proteins defined by their ability to lyse red cells but have been described to exhibit pleiotropic functions. These proteins have been extensively studied in bacteria and more recently in fungi. Within the last decade, a number of studies have characterized fungal hemolysins and revealed a fascinating yet diverse group of proteins. The purpose of this review is to provide a synopsis of the known fungal hemolysins with an emphasis on those belonging to the aegerolysin protein family. New insight and perspective into fungal hemolysins in biotechnology and health are additionally presented. PMID:22769586

Tandem-repeat protein domains across the tree of life

PubMed Central

Jernigan, Kristin K.

2015-01-01

Tandem-repeat protein domains, composed of repeated units of conserved stretches of 20–40 amino acids, are required for a wide array of biological functions. Despite their diverse and fundamental functions, there has been no comprehensive assessment of their taxonomic distribution, incidence, and associations with organismal lifestyle and phylogeny. In this study, we assess for the first time the abundance of armadillo (ARM) and tetratricopeptide (TPR) repeat domains across all three domains in the tree of life and compare the results to our previous analysis on ankyrin (ANK) repeat domains in this journal. All eukaryotes and a majority of the bacterial and archaeal genomes analyzed have a minimum of one TPR and ARM repeat. In eukaryotes, the fraction of ARM-containing proteins is approximately double that of TPR and ANK-containing proteins, whereas bacteria and archaea are enriched in TPR-containing proteins relative to ARM- and ANK-containing proteins. We show in bacteria that phylogenetic history, rather than lifestyle or pathogenicity, is a predictor of TPR repeat domain abundance, while neither phylogenetic history nor lifestyle predicts ARM repeat domain abundance. Surprisingly, pathogenic bacteria were not enriched in TPR-containing proteins, which have been associated within virulence factors in certain species. Taken together, this comparative analysis provides a newly appreciated view of the prevalence and diversity of multiple types of tandem-repeat protein domains across the tree of life. A central finding of this analysis is that tandem repeat domain-containing proteins are prevalent not just in eukaryotes, but also in bacterial and archaeal species. PMID:25653910

Mutations in NONO lead to syndromic intellectual disability and inhibitory synaptic defects.

PubMed

Mircsof, Dennis; Langouët, Maéva; Rio, Marlène; Moutton, Sébastien; Siquier-Pernet, Karine; Bole-Feysot, Christine; Cagnard, Nicolas; Nitschke, Patrick; Gaspar, Ludmila; Žnidarič, Matej; Alibeu, Olivier; Fritz, Ann-Kristina; Wolfer, David P; Schröter, Aileen; Bosshard, Giovanna; Rudin, Markus; Koester, Christina; Crestani, Florence; Seebeck, Petra; Boddaert, Nathalie; Prescott, Katrina; Hines, Rochelle; Moss, Steven J; Fritschy, Jean-Marc; Munnich, Arnold; Amiel, Jeanne; Brown, Steven A; Tyagarajan, Shiva K; Colleaux, Laurence

2015-12-01

The NONO protein has been characterized as an important transcriptional regulator in diverse cellular contexts. Here we show that loss of NONO function is a likely cause of human intellectual disability and that NONO-deficient mice have cognitive and affective deficits. Correspondingly, we find specific defects at inhibitory synapses, where NONO regulates synaptic transcription and gephyrin scaffold structure. Our data identify NONO as a possible neurodevelopmental disease gene and highlight the key role of the DBHS protein family in functional organization of GABAergic synapses.

Functional diversity of the superfamily of K⁺ transporters to meet various requirements.

PubMed

Diskowski, Marina; Mikusevic, Vedrana; Stock, Charlott; Hänelt, Inga

2015-09-01

The superfamily of K+ transporters unites proteins from plants, fungi, bacteria, and archaea that translocate K+ and/or Na+ across membranes. These proteins are key components in osmotic regulation, pH homeostasis, and resistance to high salinity and dryness. The members of the superfamily are closely related to K+ channels such as KcsA but also show several striking differences that are attributed to their altered functions. This review highlights these functional differences, focusing on the bacterial superfamily members KtrB, TrkH, and KdpA. The functional variations within the family and comparison to MPM-type K+ channels are discussed in light of the recently solved structures of the Ktr and Trk systems.

A-kinase anchoring proteins that regulate cardiac remodeling.

PubMed

Carnegie, Graeme K; Burmeister, Brian T

2011-11-01

In response to injury or stress, the adult heart undergoes maladaptive changes, collectively defined as pathological cardiac remodeling. Here, we focus on the role of A-kinase anchoring proteins (AKAPs) in 3 main areas associated with cardiac remodeling and the progression of heart failure: excitation-contraction coupling, sarcomeric regulation, and induction of pathological hypertrophy. AKAPs are a diverse family of scaffold proteins that form multiprotein complexes, integrating cAMP signaling with protein kinases, phosphatases, and other effector proteins. Many AKAPs have been characterized in the heart, where they play a critical role in modulating cardiac function.

Evidence for nuclear interaction of a cytoskeleton protein (OsIFL) with metallothionein and its role in salinity stress tolerance

PubMed Central

Soda, Neelam; Sharan, Ashutosh; Gupta, Brijesh K.; Singla-Pareek, Sneh L.; Pareek, Ashwani

2016-01-01

Soil salinity is being perceived as a major threat to agriculture. Plant breeders and molecular biologist are putting their best efforts to raise salt-tolerant crops. The discovery of the Saltol QTL, a major QTL localized on chromosome I, responsible for salt tolerance at seedling stage in rice has given new hopes for raising salinity tolerant rice genotypes. In the present study, we have functionally characterized a Saltol QTL localized cytoskeletal protein, intermediate filament like protein (OsIFL), of rice. Studies related to intermediate filaments are emerging in plants, especially with respect to their involvement in abiotic stress response. Our investigations clearly establish that the heterologous expression of OsIFL in three diverse organisms (bacteria, yeast and tobacco) provides survival advantage towards diverse abiotic stresses. Screening of rice cDNA library revealed OsIFL to be strongly interacting with metallothionein protein. Bimolecular fluorescence complementation assay further confirmed this interaction to be occurring inside the nucleus. Overexpression of OsIFL in transgenic tobacco plants conferred salinity stress tolerance by maintaining favourable K+/Na+ ratio and thus showed protection from salinity stress induced ion toxicity. This study provides the first evidence for the involvement of a cytoskeletal protein in salinity stress tolerance in diverse organisms. PMID:27708383

Applications of cell-free protein synthesis in synthetic biology: Interfacing bio-machinery with synthetic environments.

PubMed

Lee, Kyung-Ho; Kim, Dong-Myung

2013-11-01

Synthetic biology is built on the synthesis, engineering, and assembly of biological parts. Proteins are the first components considered for the construction of systems with designed biological functions because proteins carry out most of the biological functions and chemical reactions inside cells. Protein synthesis is considered to comprise the most basic levels of the hierarchical structure of synthetic biology. Cell-free protein synthesis has emerged as a powerful technology that can potentially transform the concept of bioprocesses. With the ability to harness the synthetic power of biology without many of the constraints of cell-based systems, cell-free protein synthesis enables the rapid creation of protein molecules from diverse sources of genetic information. Cell-free protein synthesis is virtually free from the intrinsic constraints of cell-based methods and offers greater flexibility in system design and manipulability of biological synthetic machinery. Among its potential applications, cell-free protein synthesis can be combined with various man-made devices for rapid functional analysis of genomic sequences. This review covers recent efforts to integrate cell-free protein synthesis with various reaction devices and analytical platforms. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Expression, stabilization and purification of membrane proteins via diverse protein synthesis systems and detergents involving cell-free associated with self-assembly peptide surfactants.

PubMed

Zheng, Xuan; Dong, Shuangshuang; Zheng, Jie; Li, Duanhua; Li, Feng; Luo, Zhongli

2014-01-01

G-protein coupled receptors (GPCRs) are involved in regulating most of physiological actions and metabolism in the bodies, which have become most frequently addressed therapeutic targets for various disorders and diseases. Purified GPCR-based drug discoveries have become routine that approaches to structural study, novel biophysical and biochemical function analyses. However, several bottlenecks that GPCR-directed drugs need to conquer the problems including overexpression, solubilization, and purification as well as stabilization. The breakthroughs are to obtain efficient protein yield and stabilize their functional conformation which are both urgently requiring of effective protein synthesis system methods and optimal surfactants. Cell-free protein synthesis system is superior to the high yields and post-translation modifications, and early signs of self-assembly peptide detergents also emerged to superiority in purification of membrane proteins. We herein focus several predominant protein synthesis systems and surfactants involving the novel peptide detergents, and uncover the advantages of cell-free protein synthesis system with self-assembling peptide detergents in purification of functional GPCRs. This review is useful to further study in membrane proteins as well as the new drug exploration. Copyright © 2014 Elsevier Inc. All rights reserved.

The Physiology of Protein S-acylation

PubMed Central

Chamberlain, Luke H.; Shipston, Michael J.

2015-01-01

Protein S-acylation, the only fully reversible posttranslational lipid modification of proteins, is emerging as a ubiquitous mechanism to control the properties and function of a diverse array of proteins and consequently physiological processes. S-acylation results from the enzymatic addition of long-chain lipids, most typically palmitate, onto intracellular cysteine residues of soluble and transmembrane proteins via a labile thioester linkage. Addition of lipid results in increases in protein hydrophobicity that can impact on protein structure, assembly, maturation, trafficking, and function. The recent explosion in global S-acylation (palmitoyl) proteomic profiling as a result of improved biochemical tools to assay S-acylation, in conjunction with the recent identification of enzymes that control protein S-acylation and de-acylation, has opened a new vista into the physiological function of S-acylation. This review introduces key features of S-acylation and tools to interrogate this process, and highlights the eclectic array of proteins regulated including membrane receptors, ion channels and transporters, enzymes and kinases, signaling adapters and chaperones, cell adhesion, and structural proteins. We highlight recent findings correlating disruption of S-acylation to pathophysiology and disease and discuss some of the major challenges and opportunities in this rapidly expanding field. PMID:25834228

Spontaneous formation of structurally diverse membrane channel architectures from a single antimicrobial peptide

NASA Astrophysics Data System (ADS)

Wang, Yukun; Chen, Charles H.; Hu, Dan; Ulmschneider, Martin B.; Ulmschneider, Jakob P.

2016-11-01

Many antimicrobial peptides (AMPs) selectively target and form pores in microbial membranes. However, the mechanisms of membrane targeting, pore formation and function remain elusive. Here we report an experimentally guided unbiased simulation methodology that yields the mechanism of spontaneous pore assembly for the AMP maculatin at atomic resolution. Rather than a single pore, maculatin forms an ensemble of structurally diverse temporarily functional low-oligomeric pores, which mimic integral membrane protein channels in structure. These pores continuously form and dissociate in the membrane. Membrane permeabilization is dominated by hexa-, hepta- and octamers, which conduct water, ions and small dyes. Pores form by consecutive addition of individual helices to a transmembrane helix or helix bundle, in contrast to current poration models. The diversity of the pore architectures--formed by a single sequence--may be a key feature in preventing bacterial resistance and could explain why sequence-function relationships in AMPs remain elusive.

Searching for Potential Silicon-associated Genes in Cyanobacteria

NASA Astrophysics Data System (ADS)

Collier, J.; Brzezinski, M. A.; Baines, S. B.; Krause, J. W.; Ohnemus, D.; Twining, B. S.

2016-02-01

Recent studies have demonstrated the accumulation of Si in both wild cells and laboratory cultures of marine Synechococcus. Because of their abundance, the cellular Si quotas measured are sufficient to suggest a substantial, unrecognized role for these organisms in the marine Si cycle. Since there is no known role for Si in cyanobacteria, we are using sequenced cyanobacterial genomes to search for pathways of Si metabolism known from other organisms. Si transporters belonging to four different protein superfamilies have been identified in diverse Si-metabolizing organisms, including diatoms and other protists, plants, bacteria, and sponges. A homolog of ArsB/Lsi2, the arsenite-antimonite efflux porter that can also transport silicate in plants, can be found in many cyanobacteria. However, we have been unable to identify likely influx porter homologs in cyanobacteria, except for predicted proteins with similarity to diatom SIT but only half the length, as well as a few atypical members of the Major Intrinsic Protein (aquaporin) superfamily. Proteins catalyzing and/or controlling the polymerization of silica have been identified in diatoms and sponges. We have been unable to identify clear homologs of these proteins in cyanobacteria, although cathepsins (belonging to the same protein superfamily as silicateins) are broadly present in cyanobacteria. Proteins that may bind silica in other bacteria (CotB in Bacillus) also lack clear homologs in cyanobacteria. However, since the function of these proteins may depend largely on charge and protein folding characteristics, proteins involved in Si deposition may not be readily identifiable by primary sequence similarity. The broad diversity of proteins involved in Si metabolism in diverse organisms suggests that each had an independent evolutionary origin. Our results suggest that if Si-associated proteins exist in Synechococcus, they also may have a distinct evolutionary origin unrelated to known Si metabolic pathways.

Interactions of signaling proteins, growth factors and other proteins with heparan sulfate: mechanisms and mysteries.

PubMed

Billings, Paul C; Pacifici, Maurizio

2015-01-01

Heparan sulfate (HS) is a component of cell surface and matrix-associated proteoglycans (HSPGs) that, collectively, play crucial roles in many physiologic processes including cell differentiation, organ morphogenesis and cancer. A key function of HS is to bind and interact with signaling proteins, growth factors, plasma proteins, immune-modulators and other factors. In doing so, the HS chains and HSPGs are able to regulate protein distribution, bio-availability and action on target cells and can also serve as cell surface co-receptors, facilitating ligand-receptor interactions. These proteins contain an HS/heparin-binding domain (HBD) that mediates their association and contacts with HS. HBDs are highly diverse in sequence and predicted structure, contain clusters of basic amino acids (Lys and Arg) and possess an overall net positive charge, most often within a consensus Cardin-Weintraub (CW) motif. Interestingly, other domains and residues are now known to influence protein-HS interactions, as well as interactions with other glycosaminoglycans, such as chondroitin sulfate. In this review, we provide a description and analysis of HBDs in proteins including amphiregulin, fibroblast growth factor family members, heparanase, sclerostin and hedgehog protein family members. We discuss HBD structural and functional features and important roles carried out by other protein domains, and also provide novel conformational insights into the diversity of CW motifs present in Sonic, Indian and Desert hedgehogs. Finally, we review progress in understanding the pathogenesis of a rare pediatric skeletal disorder, Hereditary Multiple Exostoses (HME), characterized by HS deficiency and cartilage tumor formation. Advances in understanding protein-HS interactions will have broad implications for basic biology and translational medicine as well as for the development of HS-based therapeutics.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mei, Yang; Glover, Karen; Su, Minfei

BECN1 (Beclin 1), a highly conserved eukaryotic protein, is a key regulator of autophagy, a cellular homeostasis pathway, and also participates in vacuolar protein sorting, endocytic trafficking, and apoptosis. BECN1 is important for embryonic development, the innate immune response, tumor suppression, and protection against neurodegenerative disorders, diabetes, and heart disease. BECN1 mediates autophagy as a core component of the class III phosphatidylinositol 3-kinase complexes. However, the exact mechanism by which it regulates the activity of these complexes, or mediates its other diverse functions is unclear. BECN1 interacts with several diverse protein partners, perhaps serving as a scaffold or interaction hubmore » for autophagy. Based on extensive structural, biophysical and bioinformatics analyses, BECN1 consists of an intrinsically disordered region (IDR), which includes a BH3 homology domain (BH3D); a flexible helical domain (FHD); a coiled-coil domain (CCD); and a β-α-repeated autophagy-specific domain (BARAD). Each of these BECN1 domains mediates multiple diverse interactions that involve concomitant conformational changes. Thus, BECN1 conformational flexibility likely plays a key role in facilitating diverse protein interactions. Further, BECN1 conformation and interactions are also modulated by numerous post-translational modifications. A better structure-based understanding of the interplay between different BECN1 conformational and binding states, and the impact of post-translational modifications will be essential to elucidating the mechanism of its multiple biological roles.« less

Global profiling of lysine reactivity and ligandability in the human proteome

NASA Astrophysics Data System (ADS)

Hacker, Stephan M.; Backus, Keriann M.; Lazear, Michael R.; Forli, Stefano; Correia, Bruno E.; Cravatt, Benjamin F.

2017-12-01

Nucleophilic amino acids make important contributions to protein function, including performing key roles in catalysis and serving as sites for post-translational modification. Electrophilic groups that target amino-acid nucleophiles have been used to create covalent ligands and drugs, but have, so far, been mainly limited to cysteine and serine. Here, we report a chemical proteomic platform for the global and quantitative analysis of lysine residues in native biological systems. We have quantified, in total, more than 9,000 lysines in human cell proteomes and have identified several hundred residues with heightened reactivity that are enriched at protein functional sites and can frequently be targeted by electrophilic small molecules. We have also discovered lysine-reactive fragment electrophiles that inhibit enzymes by active site and allosteric mechanisms, as well as disrupt protein-protein interactions in transcriptional regulatory complexes, emphasizing the broad potential and diverse functional consequences of liganding lysine residues throughout the human proteome.

Global profiling of lysine reactivity and ligandability in the human proteome.

PubMed

Hacker, Stephan M; Backus, Keriann M; Lazear, Michael R; Forli, Stefano; Correia, Bruno E; Cravatt, Benjamin F

2017-12-01

Nucleophilic amino acids make important contributions to protein function, including performing key roles in catalysis and serving as sites for post-translational modification. Electrophilic groups that target amino-acid nucleophiles have been used to create covalent ligands and drugs, but have, so far, been mainly limited to cysteine and serine. Here, we report a chemical proteomic platform for the global and quantitative analysis of lysine residues in native biological systems. We have quantified, in total, more than 9,000 lysines in human cell proteomes and have identified several hundred residues with heightened reactivity that are enriched at protein functional sites and can frequently be targeted by electrophilic small molecules. We have also discovered lysine-reactive fragment electrophiles that inhibit enzymes by active site and allosteric mechanisms, as well as disrupt protein-protein interactions in transcriptional regulatory complexes, emphasizing the broad potential and diverse functional consequences of liganding lysine residues throughout the human proteome.

Prediction of rat protein subcellular localization with pseudo amino acid composition based on multiple sequential features.

PubMed

Shi, Ruijia; Xu, Cunshuan

2011-06-01

The study of rat proteins is an indispensable task in experimental medicine and drug development. The function of a rat protein is closely related to its subcellular location. Based on the above concept, we construct the benchmark rat proteins dataset and develop a combined approach for predicting the subcellular localization of rat proteins. From protein primary sequence, the multiple sequential features are obtained by using of discrete Fourier analysis, position conservation scoring function and increment of diversity, and these sequential features are selected as input parameters of the support vector machine. By the jackknife test, the overall success rate of prediction is 95.6% on the rat proteins dataset. Our method are performed on the apoptosis proteins dataset and the Gram-negative bacterial proteins dataset with the jackknife test, the overall success rates are 89.9% and 96.4%, respectively. The above results indicate that our proposed method is quite promising and may play a complementary role to the existing predictors in this area.

Diversity, classification and function of the plant protein kinase superfamily

PubMed Central

Lehti-Shiu, Melissa D.; Shiu, Shin-Han

2012-01-01

Eukaryotic protein kinases belong to a large superfamily with hundreds to thousands of copies and are components of essentially all cellular functions. The goals of this study are to classify protein kinases from 25 plant species and to assess their evolutionary history in conjunction with consideration of their molecular functions. The protein kinase superfamily has expanded in the flowering plant lineage, in part through recent duplications. As a result, the flowering plant protein kinase repertoire, or kinome, is in general significantly larger than other eukaryotes, ranging in size from 600 to 2500 members. This large variation in kinome size is mainly due to the expansion and contraction of a few families, particularly the receptor-like kinase/Pelle family. A number of protein kinases reside in highly conserved, low copy number families and often play broadly conserved regulatory roles in metabolism and cell division, although functions of plant homologues have often diverged from their metazoan counterparts. Members of expanded plant kinase families often have roles in plant-specific processes and some may have contributed to adaptive evolution. Nonetheless, non-adaptive explanations, such as kinase duplicate subfunctionalization and insufficient time for pseudogenization, may also contribute to the large number of seemingly functional protein kinases in plants. PMID:22889912

A transposase strategy for creating libraries of circularly permuted proteins.

PubMed

Mehta, Manan M; Liu, Shirley; Silberg, Jonathan J

2012-05-01

A simple approach for creating libraries of circularly permuted proteins is described that is called PERMutation Using Transposase Engineering (PERMUTE). In PERMUTE, the transposase MuA is used to randomly insert a minitransposon that can function as a protein expression vector into a plasmid that contains the open reading frame (ORF) being permuted. A library of vectors that express different permuted variants of the ORF-encoded protein is created by: (i) using bacteria to select for target vectors that acquire an integrated minitransposon; (ii) excising the ensemble of ORFs that contain an integrated minitransposon from the selected vectors; and (iii) circularizing the ensemble of ORFs containing integrated minitransposons using intramolecular ligation. Construction of a Thermotoga neapolitana adenylate kinase (AK) library using PERMUTE revealed that this approach produces vectors that express circularly permuted proteins with distinct sequence diversity from existing methods. In addition, selection of this library for variants that complement the growth of Escherichia coli with a temperature-sensitive AK identified functional proteins with novel architectures, suggesting that PERMUTE will be useful for the directed evolution of proteins with new functions.

A transposase strategy for creating libraries of circularly permuted proteins

PubMed Central

Mehta, Manan M.; Liu, Shirley; Silberg, Jonathan J.

2012-01-01

A simple approach for creating libraries of circularly permuted proteins is described that is called PERMutation Using Transposase Engineering (PERMUTE). In PERMUTE, the transposase MuA is used to randomly insert a minitransposon that can function as a protein expression vector into a plasmid that contains the open reading frame (ORF) being permuted. A library of vectors that express different permuted variants of the ORF-encoded protein is created by: (i) using bacteria to select for target vectors that acquire an integrated minitransposon; (ii) excising the ensemble of ORFs that contain an integrated minitransposon from the selected vectors; and (iii) circularizing the ensemble of ORFs containing integrated minitransposons using intramolecular ligation. Construction of a Thermotoga neapolitana adenylate kinase (AK) library using PERMUTE revealed that this approach produces vectors that express circularly permuted proteins with distinct sequence diversity from existing methods. In addition, selection of this library for variants that complement the growth of Escherichia coli with a temperature-sensitive AK identified functional proteins with novel architectures, suggesting that PERMUTE will be useful for the directed evolution of proteins with new functions. PMID:22319214

EB-Family Proteins: Functions and Microtubule Interaction Mechanisms.

PubMed

Mustyatsa, V V; Boyakhchyan, A V; Ataullakhanov, F I; Gudimchuk, N B

2017-07-01

Microtubules are polymers of tubulin protein, one of the key components of cytoskeleton. They are polar filaments whose plus-ends usually oriented toward the cell periphery are more dynamic than their minus-ends, which face the center of the cell. In cells, microtubules are organized into a network that is being constantly rebuilt and renovated due to stochastic switching of its individual filaments from growth to shrinkage and back. Because of these dynamics and their mechanical properties, microtubules take part in various essential processes, from intracellular transport to search and capture of chromosomes during mitosis. Microtubule dynamics are regulated by many proteins that are located on the plus-ends of these filaments. One of the most important and abundant groups of plus-end-interacting proteins are EB-family proteins, which autonomously recognize structures of the microtubule growing plus-ends, modulate their dynamics, and recruit multiple partner proteins with diverse functions onto the microtubule plus-ends. In this review, we summarize the published data about the properties and functions of EB-proteins, focusing on analysis of their mechanism of interaction with the microtubule growing ends.

Sirtuins of parasitic protozoa: In search of function(s)

PubMed Central

Religa, Agnieszka A.; Waters, Andrew P.

2012-01-01

The SIR2 family of NAD+-dependent protein deacetylases, collectively called sirtuins, has been of central interest due to their proposed roles in life-span regulation and ageing. Sirtuins are one group of environment sensors of a cell interpreting external information and orchestrating internal responses at the sub-cellular level, through participation in gene regulation mechanisms. Remarkably conserved across all kingdoms of life SIR2 proteins in several protozoan parasites appear to have both conserved and intriguing unique functions. This review summarises our current knowledge of the members of the sirtuin families in Apicomplexa, including Plasmodium, and other protozoan parasites such as Trypanosoma and Leishmania. The wide diversity of processes regulated by SIR2 proteins makes them targets worthy of exploitation in anti-parasitic therapies. PMID:22906508

[The effect of substance P on functional proteins in human neutrophil].

PubMed

Yang, Lin; Fa, Xiang-guang

2002-02-01

To explore the effect of substance P (SP) on the functional proteins on plasma membrane of neutrophil (Np). The response of Np to SP was examined by measuring the level of respiratory burst, the activities of ACP and ALP, the fluoroscopy intensity of CR3, CD45 and FM-LP. It was found that SP could increase respiratory burst of Np, decrease the activity of acid phosphatase (ACP), but had no effect on alkaline phosphatase (ALP). SP could also promote the amount of CD45, complement receptor type 3 (CR3) and N-Formyl-Met-Leu-Phe (FMLP) receptors. The results showed that the effects of SP on functional proteins in human Np membrane were universality and diversity. It implied that SP could affect various inflammation responses in Np.

Systematic Analysis of Arabidopsis Organelles and a Protein Localization Database for Facilitating Fluorescent Tagging of Full-Length Arabidopsis Proteins1[W

PubMed Central

Li, Shijun; Ehrhardt, David W.; Rhee, Seung Y.

2006-01-01

Cells are organized into a complex network of subcellular compartments that are specialized for various biological functions. Subcellular location is an important attribute of protein function. To facilitate systematic elucidation of protein subcellular location, we analyzed experimentally verified protein localization data of 1,300 Arabidopsis (Arabidopsis thaliana) proteins. The 1,300 experimentally verified proteins are distributed among 40 different compartments, with most of the proteins localized to four compartments: mitochondria (36%), nucleus (28%), plastid (17%), and cytosol (13.3%). About 19% of the proteins are found in multiple compartments, in which a high proportion (36.4%) is localized to both cytosol and nucleus. Characterization of the overrepresented Gene Ontology molecular functions and biological processes suggests that the Golgi apparatus and peroxisome may play more diverse functions but are involved in more specialized processes than other compartments. To support systematic empirical determination of protein subcellular localization using a technology called fluorescent tagging of full-length proteins, we developed a database and Web application to provide preselected green fluorescent protein insertion position and primer sequences for all Arabidopsis proteins to study their subcellular localization and to store experimentally verified protein localization images, videos, and their annotations of proteins generated using the fluorescent tagging of full-length proteins technology. The database can be searched, browsed, and downloaded using a Web browser at http://aztec.stanford.edu/gfp/. The software can also be downloaded from the same Web site for local installation. PMID:16617091

Structural Analysis of PTM Hotspots (SAPH-ire) – A Quantitative Informatics Method Enabling the Discovery of Novel Regulatory Elements in Protein Families*

PubMed Central

Dewhurst, Henry M.; Choudhury, Shilpa; Torres, Matthew P.

2015-01-01

Predicting the biological function potential of post-translational modifications (PTMs) is becoming increasingly important in light of the exponential increase in available PTM data from high-throughput proteomics. We developed structural analysis of PTM hotspots (SAPH-ire)—a quantitative PTM ranking method that integrates experimental PTM observations, sequence conservation, protein structure, and interaction data to allow rank order comparisons within or between protein families. Here, we applied SAPH-ire to the study of PTMs in diverse G protein families, a conserved and ubiquitous class of proteins essential for maintenance of intracellular structure (tubulins) and signal transduction (large and small Ras-like G proteins). A total of 1728 experimentally verified PTMs from eight unique G protein families were clustered into 451 unique hotspots, 51 of which have a known and cited biological function or response. Using customized software, the hotspots were analyzed in the context of 598 unique protein structures. By comparing distributions of hotspots with known versus unknown function, we show that SAPH-ire analysis is predictive for PTM biological function. Notably, SAPH-ire revealed high-ranking hotspots for which a functional impact has not yet been determined, including phosphorylation hotspots in the N-terminal tails of G protein gamma subunits—conserved protein structures never before reported as regulators of G protein coupled receptor signaling. To validate this prediction we used the yeast model system for G protein coupled receptor signaling, revealing that gamma subunit–N-terminal tail phosphorylation is activated in response to G protein coupled receptor stimulation and regulates protein stability in vivo. These results demonstrate the utility of integrating protein structural and sequence features into PTM prioritization schemes that can improve the analysis and functional power of modification-specific proteomics data. PMID:26070665

Small Cofactors May Assist Protein Emergence from RNA World: Clues from RNA-Protein Complexes

PubMed Central

Shen, Liang; Ji, Hong-Fang

2011-01-01

It is now widely accepted that at an early stage in the evolution of life an RNA world arose, in which RNAs both served as the genetic material and catalyzed diverse biochemical reactions. Then, proteins have gradually replaced RNAs because of their superior catalytic properties in catalysis over time. Therefore, it is important to investigate how primitive functional proteins emerged from RNA world, which can shed light on the evolutionary pathway of life from RNA world to the modern world. In this work, we proposed that the emergence of most primitive functional proteins are assisted by the early primitive nucleotide cofactors, while only a minority are induced directly by RNAs based on the analysis of RNA-protein complexes. Furthermore, the present findings have significant implication for exploring the composition of primitive RNA, i.e., adenine base as principal building blocks. PMID:21789260

Structural and Functional Assessment of APOBEC3G Macromolecular Complexes

PubMed Central

Polevoda, Bogdan; McDougall, William M.; Bennett, Ryan P.; Salter, Jason D.; Smith, Harold C.

2016-01-01

There are eleven members in the human APOBEC family of proteins that are evolutionarily related through their zinc-dependent cytidine deaminase domains. The human APOBEC gene clusters arose on chromosome 6 and 22 through gene duplication and divergence to where current day APOBEC proteins are functionally diverse and broadly expressed in tissues. APOBEC serve enzymatic and non enzymatic functions in cells. In both cases, formation of higher-order structures driven by APOBEC protein-protein interactions and binding to RNA and/or single stranded DNA are integral to their function. In some circumstances, these interactions are regulatory and modulate APOBEC activities. We are just beginning to understand how macromolecular interactions drive processes such as APOBEC subcellular compartmentalization, formation of holoenzyme complexes, gene targeting, foreign DNA restriction, anti-retroviral activity, formation of ribonucleoprotein particles and APOBEC degradation. Protein-protein and protein-nucleic acid cross-linking methods coupled with mass spectrometry, electrophoretic mobility shift assays, glycerol gradient sedimentation, fluorescence anisotropy and APOBEC deaminase assays are enabling mapping of interacting surfaces that are essential for these functions. The goal of this methods review is through example of our research on APOBEC3G, describe the application of cross-linking methods to characterize and quantify macromolecular interactions and their functional implications. Given the homology in structure and function, it is proposed that these methods will be generally applicable to the discovery process for other APOBEC and RNA and DNA editing and modifying proteins. PMID:26988126

An emerging link between LIM domain proteins and nuclear receptors.

PubMed

Sala, Stefano; Ampe, Christophe

2018-06-01

Nuclear receptors are ligand-activated transcription factors that partake in several biological processes including development, reproduction and metabolism. Over the last decade, evidence has accumulated that group 2, 3 and 4 LIM domain proteins, primarily known for their roles in actin cytoskeleton organization, also partake in gene transcription regulation. They shuttle between the cytoplasm and the nucleus, amongst other as a consequence of triggering cells with ligands of nuclear receptors. LIM domain proteins act as important coregulators of nuclear receptor-mediated gene transcription, in which they can either function as coactivators or corepressors. In establishing interactions with nuclear receptors, the LIM domains are important, yet pleiotropy of LIM domain proteins and nuclear receptors frequently occurs. LIM domain protein-nuclear receptor complexes function in diverse physiological processes. Their association is, however, often linked to diseases including cancer.

Structure, Biology, and Therapeutic Application of Toxin-Antitoxin Systems in Pathogenic Bacteria.

PubMed

Lee, Ki-Young; Lee, Bong-Jin

2016-10-22

Bacterial toxin-antitoxin (TA) systems have received increasing attention for their diverse identities, structures, and functional implications in cell cycle arrest and survival against environmental stresses such as nutrient deficiency, antibiotic treatments, and immune system attacks. In this review, we describe the biological functions and the auto-regulatory mechanisms of six different types of TA systems, among which the type II TA system has been most extensively studied. The functions of type II toxins include mRNA/tRNA cleavage, gyrase/ribosome poison, and protein phosphorylation, which can be neutralized by their cognate antitoxins. We mainly explore the similar but divergent structures of type II TA proteins from 12 important pathogenic bacteria, including various aspects of protein-protein interactions. Accumulating knowledge about the structure-function correlation of TA systems from pathogenic bacteria has facilitated a novel strategy to develop antibiotic drugs that target specific pathogens. These molecules could increase the intrinsic activity of the toxin by artificially interfering with the intermolecular network of the TA systems.

Regulation of T-cell receptor signalling by membrane microdomains

PubMed Central

Razzaq, Tahir M; Ozegbe, Patricia; Jury, Elizabeth C; Sembi, Phupinder; Blackwell, Nathan M; Kabouridis, Panagiotis S

2004-01-01

There is now considerable evidence suggesting that the plasma membrane of mammalian cells is compartmentalized by functional lipid raft microdomains. These structures are assemblies of specialized lipids and proteins and have been implicated in diverse biological functions. Analysis of their protein content using proteomics and other methods revealed enrichment of signalling proteins, suggesting a role for these domains in intracellular signalling. In T lymphocytes, structure/function experiments and complementary pharmacological studies have shown that raft microdomains control the localization and function of proteins which are components of signalling pathways regulated by the T-cell antigen receptor (TCR). Based on these studies, a model for TCR phosphorylation in lipid rafts is presented. However, despite substantial progress in the field, critical questions remain. For example, it is unclear if membrane rafts represent a homogeneous population and if their structure is modified upon TCR stimulation. In the future, proteomics and the parallel development of complementary analytical methods will undoubtedly contribute in further delineating the role of lipid rafts in signal transduction mechanisms. PMID:15554919

E3Net: a system for exploring E3-mediated regulatory networks of cellular functions.

PubMed

Han, Youngwoong; Lee, Hodong; Park, Jong C; Yi, Gwan-Su

2012-04-01

Ubiquitin-protein ligase (E3) is a key enzyme targeting specific substrates in diverse cellular processes for ubiquitination and degradation. The existing findings of substrate specificity of E3 are, however, scattered over a number of resources, making it difficult to study them together with an integrative view. Here we present E3Net, a web-based system that provides a comprehensive collection of available E3-substrate specificities and a systematic framework for the analysis of E3-mediated regulatory networks of diverse cellular functions. Currently, E3Net contains 2201 E3s and 4896 substrates in 427 organisms and 1671 E3-substrate specific relations between 493 E3s and 1277 substrates in 42 organisms, extracted mainly from MEDLINE abstracts and UniProt comments with an automatic text mining method and additional manual inspection and partly from high throughput experiment data and public ubiquitination databases. The significant functions and pathways of the extracted E3-specific substrate groups were identified from a functional enrichment analysis with 12 functional category resources for molecular functions, protein families, protein complexes, pathways, cellular processes, cellular localization, and diseases. E3Net includes interactive analysis and navigation tools that make it possible to build an integrative view of E3-substrate networks and their correlated functions with graphical illustrations and summarized descriptions. As a result, E3Net provides a comprehensive resource of E3s, substrates, and their functional implications summarized from the regulatory network structures of E3-specific substrate groups and their correlated functions. This resource will facilitate further in-depth investigation of ubiquitination-dependent regulatory mechanisms. E3Net is freely available online at http://pnet.kaist.ac.kr/e3net.

Mining secreted proteins that function in pepper fruit development and ripening using a yeast secretion trap (YST)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Je Min, E-mail: jemin@knu.ac.kr; Department of Horticultural Science, Kyungpook National University, Daegu; Lee, Sang-Jik

Highlights: • Yeast secretion trap (YST) is a valuable tool for mining secretome. • A total of 80 secreted proteins are newly identified via YST in pepper fruits. • The secreted proteins are differentially regulated during pepper development and ripening. • Transient GFP-fusion assay and in planta secretion trap can effectively validate the secretion of proteins. - Abstract: Plant cells secrete diverse sets of constitutively- and conditionally-expressed proteins under various environmental and developmental states. Secreted protein populations, or secretomes have multiple functions, including defense responses, signaling, metabolic processes, and developmental regulation. To identify genes encoding secreted proteins that function inmore » fruit development and ripening, a yeast secretion trap (YST) screen was employed using pepper (Capsicum annuum) fruit cDNAs. The YST screen revealed 80 pepper fruit-related genes (CaPFRs) encoding secreted proteins including cell wall proteins, several of which have not been previously described. Transient GFP-fusion assay and an in planta secretion trap were used to validate the secretion of proteins encoded by selected YST clones. In addition, RNA gel blot analyses provided further insights into their expression and regulation during fruit development and ripening. Integrating our data, we conclude that the YST provides a valuable functional genomics tool for the identification of substantial numbers of novel secreted plant proteins that are associated with biological processes, including fruit development and ripening.« less

Looking at the Disordered Proteins through the Computational Microscope.

PubMed

Das, Payel; Matysiak, Silvina; Mittal, Jeetain

2018-05-23

Intrinsically disordered proteins (IDPs) have attracted wide interest over the past decade due to their surprising prevalence in the proteome and versatile roles in cell physiology and pathology. A large selection of IDPs has been identified as potential targets for therapeutic intervention. Characterizing the structure-function relationship of disordered proteins is therefore an essential but daunting task, as these proteins can adapt transient structure, necessitating a new paradigm for connecting structural disorder to function. Molecular simulation has emerged as a natural complement to experiments for atomic-level characterizations and mechanistic investigations of this intriguing class of proteins. The diverse range of length and time scales involved in IDP function requires performing simulations at multiple levels of resolution. In this Outlook, we focus on summarizing available simulation methods, along with a few interesting example applications. We also provide an outlook on how these simulation methods can be further improved in order to provide a more accurate description of IDP structure, binding, and assembly.

Rap G protein signal in normal and disordered lymphohematopoiesis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Minato, Nagahiro, E-mail: minato@imm.med.kyoto-u.ac.jp

2013-09-10

Rap proteins (Rap1, Rap2a, b, c) are small molecular weight GTPases of the Ras family. Rap G proteins mediate diverse cellular events such as cell adhesion, proliferation, and gene activation through various signaling pathways. Activation of Rap signal is regulated tightly by several specific regulatory proteins including guanine nucleotide exchange factors and GTPase-activating proteins. Beyond cell biological studies, increasing attempts have been made in the past decade to define the roles of Rap signal in specific functions of normal tissue systems as well as in cancer. In the immune and hematopoietic systems, Rap signal plays crucial roles in the developmentmore » and function of essentially all lineages of lymphocytes and hematopoietic cells, and importantly, deregulated Rap signal may lead to unique pathological conditions depending on the affected cell types, including various types of leukemia and autoimmunity. The phenotypical studies have unveiled novel, even unexpected functional aspects of Rap signal in cells from a variety of tissues, providing potentially important clues for controlling human diseases, including malignancy.« less

Looking at the Disordered Proteins through the Computational Microscope

PubMed Central

2018-01-01

Intrinsically disordered proteins (IDPs) have attracted wide interest over the past decade due to their surprising prevalence in the proteome and versatile roles in cell physiology and pathology. A large selection of IDPs has been identified as potential targets for therapeutic intervention. Characterizing the structure–function relationship of disordered proteins is therefore an essential but daunting task, as these proteins can adapt transient structure, necessitating a new paradigm for connecting structural disorder to function. Molecular simulation has emerged as a natural complement to experiments for atomic-level characterizations and mechanistic investigations of this intriguing class of proteins. The diverse range of length and time scales involved in IDP function requires performing simulations at multiple levels of resolution. In this Outlook, we focus on summarizing available simulation methods, along with a few interesting example applications. We also provide an outlook on how these simulation methods can be further improved in order to provide a more accurate description of IDP structure, binding, and assembly.

Enzymes in human milk

PubMed Central

Dallas, David C.; German, J. Bruce

2017-01-01

Milk proteins are a complex and diverse source of biological activities. Beyond their function intact, milk proteins also act as carriers of encrypted functional sequences that when released as peptides exert biological functions, including antimicrobial and immunomodulatory, which could contribute to the infant’s competitive success. Research has now revealed that the release of these functional peptides begins within the mammary gland itself. A complex array of proteases produced in mother’s milk have been shown to be active in the milk, releasing these peptides. Moreover, our recent research demonstrates that these milk proteases continue to digest milk proteins within the infant’s stomach, possibly even to a larger extent than the infant’s own proteases. As the neonate has relatively low digestive capacity, the activity of milk proteases in the infant may provide important assistance to digesting milk proteins. The coordinated release of these encrypted sequences is accomplished by selective proteolytic action provided by an array of native milk proteases and infant-produced enzymes. The task for scientists is now to discover the selective advantages of this protein-protease based peptide release system. PMID:28346930

Enzymes in Human Milk.

PubMed

Dallas, David C; German, J Bruce

2017-01-01

Milk proteins are a complex and diverse source of biological activities. Beyond their function, intact milk proteins also act as carriers of encrypted functional sequences that, when released as peptides, exert biological functions, including antimicrobial and immunomodulatory activity, which could contribute to the infant's competitive success. Research has now revealed that the release of these functional peptides begins within the mammary gland itself. A complex array of proteases produced in mother's milk has been shown to be active in the milk, releasing these peptides. Moreover, our recent research demonstrates that these milk proteases continue to digest milk proteins within the infant's stomach, possibly even to a larger extent than the infant's own proteases. As the neonate has relatively low digestive capacity, the activity of milk proteases in the infant may provide important assistance to digesting milk proteins. The coordinated release of these encrypted sequences is accomplished by selective proteolytic action provided by an array of native milk proteases and infant-produced enzymes. The task for scientists is now to discover the selective advantages of this protein-protease-based peptide release system. © 2017 Nestec Ltd., Vevey/S. Karger AG, Basel.

LenVarDB: database of length-variant protein domains.

PubMed

Mutt, Eshita; Mathew, Oommen K; Sowdhamini, Ramanathan

2014-01-01

Protein domains are functionally and structurally independent modules, which add to the functional variety of proteins. This array of functional diversity has been enabled by evolutionary changes, such as amino acid substitutions or insertions or deletions, occurring in these protein domains. Length variations (indels) can introduce changes at structural, functional and interaction levels. LenVarDB (freely available at http://caps.ncbs.res.in/lenvardb/) traces these length variations, starting from structure-based sequence alignments in our Protein Alignments organized as Structural Superfamilies (PASS2) database, across 731 structural classification of proteins (SCOP)-based protein domain superfamilies connected to 2 730 625 sequence homologues. Alignment of sequence homologues corresponding to a structural domain is available, starting from a structure-based sequence alignment of the superfamily. Orientation of the length-variant (indel) regions in protein domains can be visualized by mapping them on the structure and on the alignment. Knowledge about location of length variations within protein domains and their visual representation will be useful in predicting changes within structurally or functionally relevant sites, which may ultimately regulate protein function. Non-technical summary: Evolutionary changes bring about natural changes to proteins that may be found in many organisms. Such changes could be reflected as amino acid substitutions or insertions-deletions (indels) in protein sequences. LenVarDB is a database that provides an early overview of observed length variations that were set among 731 protein families and after examining >2 million sequences. Indels are followed up to observe if they are close to the active site such that they can affect the activity of proteins. Inclusion of such information can aid the design of bioengineering experiments.

Plant Abiotic Stress Proteomics: The Major Factors Determining Alterations in Cellular Proteome

PubMed Central

Kosová, Klára; Vítámvás, Pavel; Urban, Milan O.; Prášil, Ilja T.; Renaut, Jenny

2018-01-01

HIGHLIGHTS: Major environmental and genetic factors determining stress-related protein abundance are discussed.Major aspects of protein biological function including protein isoforms and PTMs, cellular localization and protein interactions are discussed.Functional diversity of protein isoforms and PTMs is discussed. Abiotic stresses reveal profound impacts on plant proteomes including alterations in protein relative abundance, cellular localization, post-transcriptional and post-translational modifications (PTMs), protein interactions with other protein partners, and, finally, protein biological functions. The main aim of the present review is to discuss the major factors determining stress-related protein accumulation and their final biological functions. A dynamics of stress response including stress acclimation to altered ambient conditions and recovery after the stress treatment is discussed. The results of proteomic studies aimed at a comparison of stress response in plant genotypes differing in stress adaptability reveal constitutively enhanced levels of several stress-related proteins (protective proteins, chaperones, ROS scavenging- and detoxification-related enzymes) in the tolerant genotypes with respect to the susceptible ones. Tolerant genotypes can efficiently adjust energy metabolism to enhanced needs during stress acclimation. Stress tolerance vs. stress susceptibility are relative terms which can reflect different stress-coping strategies depending on the given stress treatment. The role of differential protein isoforms and PTMs with respect to their biological functions in different physiological constraints (cellular compartments and interacting partners) is discussed. The importance of protein functional studies following high-throughput proteome analyses is presented in a broader context of plant biology. In summary, the manuscript tries to provide an overview of the major factors which have to be considered when interpreting data from proteomic studies on stress-treated plants. PMID:29472941

Fascin- and α-Actinin-Bundled Networks Contain Intrinsic Structural Features that Drive Protein Sorting.

PubMed

Winkelman, Jonathan D; Suarez, Cristian; Hocky, Glen M; Harker, Alyssa J; Morganthaler, Alisha N; Christensen, Jenna R; Voth, Gregory A; Bartles, James R; Kovar, David R

2016-10-24

Cells assemble and maintain functionally distinct actin cytoskeleton networks with various actin filament organizations and dynamics through the coordinated action of different sets of actin-binding proteins. The biochemical and functional properties of diverse actin-binding proteins, both alone and in combination, have been increasingly well studied. Conversely, how different sets of actin-binding proteins properly sort to distinct actin filament networks in the first place is not nearly as well understood. Actin-binding protein sorting is critical for the self-organization of diverse dynamic actin cytoskeleton networks within a common cytoplasm. Using in vitro reconstitution techniques including biomimetic assays and single-molecule multi-color total internal reflection fluorescence microscopy, we discovered that sorting of the prominent actin-bundling proteins fascin and α-actinin to distinct networks is an intrinsic behavior, free of complicated cellular signaling cascades. When mixed, fascin and α-actinin mutually exclude each other by promoting their own recruitment and inhibiting recruitment of the other, resulting in the formation of distinct fascin- or α-actinin-bundled domains. Subdiffraction-resolution light microscopy and negative-staining electron microscopy revealed that fascin domains are densely packed, whereas α-actinin domains consist of widely spaced parallel actin filaments. Importantly, other actin-binding proteins such as fimbrin and espin show high specificity between these two bundle types within the same reaction. Here we directly observe that fascin and α-actinin intrinsically segregate to discrete bundled domains that are specifically recognized by other actin-binding proteins. Copyright © 2016 Elsevier Ltd. All rights reserved.

Dissecting the protein architecture of DNA-binding transcription factors in bacteria and archaea.

PubMed

Rivera-Gómez, Nancy; Martínez-Núñez, Mario Alberto; Pastor, Nina; Rodriguez-Vazquez, Katya; Perez-Rueda, Ernesto

2017-08-01

Gene regulation at the transcriptional level is a central process in all organisms where DNA-binding transcription factors play a fundamental role. This class of proteins binds specifically at DNA sequences, activating or repressing gene expression as a function of the cell's metabolic status, operator context and ligand-binding status, among other factors, through the DNA-binding domain (DBD). In addition, TFs may contain partner domains (PaDos), which are involved in ligand binding and protein-protein interactions. In this work, we systematically evaluated the distribution, abundance and domain organization of DNA-binding TFs in 799 non-redundant bacterial and archaeal genomes. We found that the distributions of the DBDs and their corresponding PaDos correlated with the size of the genome. We also identified specific combinations between the DBDs and their corresponding PaDos. Within each class of DBDs there are differences in the actual angle formed at the dimerization interface, responding to the presence/absence of ligands and/or crystallization conditions, setting the orientation of the resulting helices and wings facing the DNA. Our results highlight the importance of PaDos as central elements that enhance the diversity of regulatory functions in all bacterial and archaeal organisms, and our results also demonstrate the role of PaDos in sensing diverse signal compounds. The highly specific interactions between DBDs and PaDos observed in this work, together with our structural analysis highlighting the difficulty in predicting both inter-domain geometry and quaternary structure, suggest that these systems appeared once and evolved with diverse duplication events in all the analysed organisms.

Identification and expression analysis of the apple (Malus × domestica) basic helix-loop-helix transcription factor family.

PubMed

Yang, Jinhua; Gao, Min; Huang, Li; Wang, Yaqiong; van Nocker, Steve; Wan, Ran; Guo, Chunlei; Wang, Xiping; Gao, Hua

2017-02-09

Basic helix-loop-helix (bHLH) proteins, which are characterized by a conserved bHLH domain, comprise one of the largest families of transcription factors in both plants and animals, and have been shown to have a wide range of biological functions. However, there have been very few studies of bHLH proteins from perennial tree species. We describe here the identification and characterization of 175 bHLH transcription factors from apple (Malus × domestica). Phylogenetic analysis of apple bHLH (MdbHLH) genes and their Arabidopsis thaliana (Arabidopsis) orthologs indicated that they can be classified into 23 subgroups. Moreover, integrated synteny analysis suggested that the large-scale expansion of the bHLH transcription factor family occurred before the divergence of apple and Arabidopsis. An analysis of the exon/intron structure and protein domains was conducted to suggest their functional roles. Finally, we observed that MdbHLH subgroup III and IV genes displayed diverse expression profiles in various organs, as well as in response to abiotic stresses and various hormone treatments. Taken together, these data provide new information regarding the composition and diversity of the apple bHLH transcription factor family that will provide a platform for future targeted functional characterization.

Tropomodulins: pointed-end capping proteins that regulate actin filament architecture in diverse cell types

PubMed Central

Yamashiro, Sawako; Gokhin, David S.; Kimura, Sumiko; Nowak, Roberta B.; Fowler, Velia M.

2012-01-01

Tropomodulins are a family of four proteins (Tmods 1–4) that cap the pointed ends of actin filaments in actin cytoskeletal structures in a developmentally regulated and tissue-specific manner. Unique among capping proteins, Tmods also bind tropomyosins (TMs), which greatly enhance the actin filament pointed-end capping activity of Tmods. Tmods are defined by a tropomyosin (TM)-regulated/Pointed-End Actin Capping (TM-Cap) domain in their unstructured N-terminal portion, followed by a compact, folded Leucine-Rich Repeat/Pointed-End Actin Capping (LRR-Cap) domain. By inhibiting actin monomer association and dissociation from pointed ends, Tmods regulate regulate actin dynamics and turnover, stabilizing actin filament lengths and cytoskeletal architecture. In this review, we summarize the genes, structural features, molecular and biochemical properties, actin regulatory mechanisms, expression patterns, and cell and tissue functions of Tmods. By understanding Tmods’ functions in the context of their molecular structure, actin regulation, binding partners, and related variants (leiomodins 1–3), we can draw broad conclusions that can explain the diverse morphological and functional phenotypes that arise from Tmod perturbation experiments in vitro and in vivo. Tmod-based stabilization and organization of intracellular actin filament networks provide key insights into how the emergent properties of the actin cytoskeleton drive tissue morphogenesis and physiology. PMID:22488942

Hsp40 function in yeast prion propagation: Amyloid diversity necessitates chaperone functional complexity.

PubMed

Sporn, Zachary A; Hines, Justin K

2015-01-01

Yeast prions are heritable protein-based elements, most of which are formed of amyloid aggregates that rely on the action of molecular chaperones for transmission to progeny. Prions can form distinct amyloid structures, known as 'strains' in mammalian systems, that dictate both pathological progression and cross-species infection barriers. In yeast these same amyloid structural polymorphisms, called 'variants', dictate the intensity of prion-associated phenotypes and stability in mitosis. We recently reported that [PSI(+)] prion variants differ in the fundamental domain requirements for one chaperone, the Hsp40/J-protein Sis1, which are mutually exclusive between 2 different yeast prions, demonstrating a functional plurality for Sis1. Here we extend that analysis to incorporate additional data that collectively support the hypothesis that Sis1 has multiple functional roles that can be accomplished by distinct sets of domains. These functions are differentially required by distinct prions and prion variants. We also present new data regarding Hsp104-mediated prion elimination and show that some Sis1 functions, but not all, are conserved in the human homolog Hdj1/DNAJB1. Importantly, of the 10 amyloid-based prions indentified to date in Saccharomyces cerevisiae, the chaperone requirements of only 4 are known, leaving a great diversity of amyloid structures, and likely modes of amyloid-chaperone interaction, largely unexplored.

Diverse Roles for Auxiliary Subunits in Phosphorylation-Dependent Regulation of Mammalian Brain Voltage-Gated Potassium Channels

PubMed Central

Vacher, Helene; Trimmer, James S.

2012-01-01

Voltage-gated ion channels are a diverse family of signaling proteins that mediate rapid electrical signaling events. Among these, voltage-gated potassium or Kv channels are the most diverse, in part due to the large number of principal (or α) subunits and auxiliary subunits that can assemble in different combinations to generate Kv channel complexes with distinct structures and functions. The diversity of Kv channels underlies much of the variability in the active properties between different mammalian central neurons, and the dynamic changes that lead to experience-dependent plasticity in intrinsic excitability. Recent studies have revealed that Kv channel α subunits and auxiliary subunits are extensively phosphorylated, contributing to additional structural and functional diversity. Here we highlight recent studies that show that auxiliary subunits exert some of their profound effects on dendritic Kv4 and axonal Kv1 channels through phosphorylation-dependent mechanisms, either due to phosphorylation on the auxiliary subunit itself, or by influencing the extent and/or impact of α subunit phosphorylation. The complex effects of auxiliary subunits and phosphorylation provide a potent mechanism to generate additional diversity in the structure and function of Kv4 and Kv1 channels, as well as allowing for dynamic reversible regulation of these important ion channels. PMID:21822597

Insights into the Shc Family of Adaptor Proteins

PubMed Central

Prigent, Sally A.

2017-01-01

The Shc family of adaptor proteins is a group of proteins that lacks intrinsic enzymatic activity. Instead, Shc proteins possess various domains that allow them to recruit different signalling molecules. Shc proteins help to transduce an extracellular signal into an intracellular signal, which is then translated into a biological response. The Shc family of adaptor proteins share the same structural topography, CH2-PTB-CH1-SH2, which is more than an isoform of Shc family proteins; this structure, which includes multiple domains, allows for the posttranslational modification of Shc proteins and increases the functional diversity of Shc proteins. The deregulation of Shc proteins has been linked to different disease conditions, including cancer and Alzheimer’s, which indicates their key roles in cellular functions. Accordingly, a question might arise as to whether Shc proteins could be targeted therapeutically to correct their disturbance. To answer this question, thorough knowledge must be acquired; herein, we aim to shed light on the Shc family of adaptor proteins to understand their intracellular role in normal and disease states, which later might be applied to connote mechanisms to reverse the disease state.

The increasing diversity of functions attributed to the SAFB family of RNA-/DNA-binding proteins.

PubMed

Norman, Michael; Rivers, Caroline; Lee, Youn-Bok; Idris, Jalilah; Uney, James

2016-12-01

RNA-binding proteins play a central role in cellular metabolism by orchestrating the complex interactions of coding, structural and regulatory RNA species. The SAFB (scaffold attachment factor B) proteins (SAFB1, SAFB2 and SAFB-like transcriptional modulator, SLTM), which are highly conserved evolutionarily, were first identified on the basis of their ability to bind scaffold attachment region DNA elements, but attention has subsequently shifted to their RNA-binding and protein-protein interactions. Initial studies identified the involvement of these proteins in the cellular stress response and other aspects of gene regulation. More recently, the multifunctional capabilities of SAFB proteins have shown that they play crucial roles in DNA repair, processing of mRNA and regulatory RNA, as well as in interaction with chromatin-modifying complexes. With the advent of new techniques for identifying RNA-binding sites, enumeration of individual RNA targets has now begun. This review aims to summarise what is currently known about the functions of SAFB proteins. © 2016 The Author(s).

A surprising role for conformational entropy in protein function

PubMed Central

Wand, A. Joshua; Moorman, Veronica R.; Harpole, Kyle W.

2014-01-01

Formation of high-affinity complexes is critical for the majority of enzymatic reactions involving proteins. The creation of the family of Michaelis and other intermediate complexes during catalysis clearly involves a complicated manifold of interactions that are diverse and complex. Indeed, computing the energetics of interactions between proteins and small molecule ligands using molecular structure alone remains a grand challenge. One of the most difficult contributions to the free energy of protein-ligand complexes to experimentally access is that due to changes in protein conformational entropy. Fortunately, recent advances in solution nuclear magnetic resonance (NMR) relaxation methods have enabled the use of measures-of-motion between conformational states of a protein as a proxy for conformational entropy. This review briefly summarizes the experimental approaches currently employed to characterize fast internal motion in proteins, how this information is used to gain insight into conformational entropy, what has been learned and what the future may hold for this emerging view of protein function. PMID:23478875

A library of protein cage architectures as nanomaterials.

PubMed

Flenniken, M L; Uchida, M; Liepold, L O; Kang, S; Young, M J; Douglas, T

2009-01-01

Virus capsids and other structurally related cage-like proteins such as ferritins, dps, and heat shock proteins have three distinct surfaces (inside, outside, interface) that can be exploited to generate nanomaterials with multiple functionality by design. Protein cages are biological in origin and each cage exhibits extremely homogeneous size distribution. This homogeneity can be used to attain a high degree of homogeneity of the templated material and its associated property. A series of protein cages exhibiting diversity in size, functionality, and chemical and thermal stabilities can be utilized for materials synthesis under a variety of conditions. Since synthetic approaches to materials science often use harsh temperature and pH, it is an advantage to utilize protein cages from extreme environments. In this chapter, we review recent studies on discovering novel protein cages from harsh natural environments such as the acidic thermal hot springs at Yellowstone National Park (YNP) and on utilizing protein cages as nano-scale platforms for developing nanomaterials with wide range of applications from electronics to biomedicine.

Genomics and structure/function studies of Rhabdoviridae proteins involved in replication and transcription.

PubMed

Assenberg, R; Delmas, O; Morin, B; Graham, S C; De Lamballerie, X; Laubert, C; Coutard, B; Grimes, J M; Neyts, J; Owens, R J; Brandt, B W; Gorbalenya, A; Tucker, P; Stuart, D I; Canard, B; Bourhy, H

2010-08-01

Some mammalian rhabdoviruses may infect humans, and also infect invertebrates, dogs, and bats, which may act as vectors transmitting viruses among different host species. The VIZIER programme, an EU-funded FP6 program, has characterized viruses that belong to the Vesiculovirus, Ephemerovirus and Lyssavirus genera of the Rhabdoviridae family to perform ground-breaking research on the identification of potential new drug targets against these RNA viruses through comprehensive structural characterization of the replicative machinery. The contribution of VIZIER programme was of several orders. First, it contributed substantially to research aimed at understanding the origin, evolution and diversity of rhabdoviruses. This diversity was then used to obtain further structural information on the proteins involved in replication. Two strategies were used to produce recombinant proteins by expression of both full length or domain constructs in either E. coli or insect cells, using the baculovirus system. In both cases, parallel cloning and expression screening at small-scale of multiple constructs based on different viruses including the addition of fusion tags, was key to the rapid generation of expression data. As a result, some progress has been made in the VIZIER programme towards dissecting the multi-functional L protein into components suitable for structural and functional studies. However, the phosphoprotein polymerase co-factor and the structural matrix protein, which play a number of roles during viral replication and drives viral assembly, have both proved much more amenable to structural biology. Applying the multi-construct/multi-virus approach central to protein production processes in VIZIER has yielded new structural information which may ultimately be exploitable in the derivation of novel ways of intervening in viral replication. Copyright 2010 Elsevier B.V. All rights reserved.

PREFACE: Protein protein interactions: principles and predictions

NASA Astrophysics Data System (ADS)

Nussinov, Ruth; Tsai, Chung-Jung

2005-06-01

Proteins are the `workhorses' of the cell. Their roles span functions as diverse as being molecular machines and signalling. They carry out catalytic reactions, transport, form viral capsids, traverse membranes and form regulated channels, transmit information from DNA to RNA, making possible the synthesis of new proteins, and they are responsible for the degradation of unnecessary proteins and nucleic acids. They are the vehicles of the immune response and are responsible for viral entry into the cell. Given their importance, considerable effort has been centered on the prediction of protein function. A prime way to do this is through identification of binding partners. If the function of at least one of the components with which the protein interacts is known, that should let us assign its function(s) and the pathway(s) in which it plays a role. This holds since the vast majority of their chores in the living cell involve protein-protein interactions. Hence, through the intricate network of these interactions we can map cellular pathways, their interconnectivities and their dynamic regulation. Their identification is at the heart of functional genomics; their prediction is crucial for drug discovery. Knowledge of the pathway, its topology, length, and dynamics may provide useful information for forecasting side effects. The goal of predicting protein-protein interactions is daunting. Some associations are obligatory, others are continuously forming and dissociating. In principle, from the physical standpoint, any two proteins can interact, but under what conditions and at which strength? The principles of protein-protein interactions are general: the non-covalent interactions of two proteins are largely the outcome of the hydrophobic effect, which drives the interactions. In addition, hydrogen bonds and electrostatic interactions play important roles. Thus, many of the interactions observed in vitro are the outcome of experimental overexpression. Protein disorder is important in protein-protein association. It has been estimated that a large fraction of cellular proteins are `natively disordered', i.e., unstable in solution. The disordered state has a significant residual structure. In this state, a protein exists in an ensemble of rapidly interconverting conformers. They play roles in cell-cycle control, signal transduction, transcriptional and translational regulation, and in large macromolecular complexes. It has been suggested that natively disordered proteins are more `adaptive', and thus advantageous in regulation and in binding diverse ligands. Alternatively, since the native conformation is still likely to be the most abundant within the ensemble, disordered proteins, which typically have larger interface to size ratios, lead to smaller protein, genome and cell sizes, and thus are functionally advantageous. To be able to predict protein-protein interactions, we need to discern various aspects of their associations: from their shape complementarity to the organization and relative contributions of the different physical components to their stability. They involve the static and the dynamic. Proteins interact through their surfaces. Thus, to analyze their interactions, we typically study residues (or atoms) which are in contact across the two-chain interface. In addition, we often inspect the residues in their vicinity, exploring their supporting matrix. The hope is that through the understanding of the principles and mechanisms of the interactions, we shall eventually be able to solve the protein-protein interaction puzzle.

De Novo Evolutionary Emergence of a Symmetrical Protein Is Shaped by Folding Constraints

PubMed Central

Smock, Robert G.; Yadid, Itamar; Dym, Orly; Clarke, Jane; Tawfik, Dan S.

2016-01-01

Summary Molecular evolution has focused on the divergence of molecular functions, yet we know little about how structurally distinct protein folds emerge de novo. We characterized the evolutionary trajectories and selection forces underlying emergence of β-propeller proteins, a globular and symmetric fold group with diverse functions. The identification of short propeller-like motifs (<50 amino acids) in natural genomes indicated that they expanded via tandem duplications to form extant propellers. We phylogenetically reconstructed 47-residue ancestral motifs that form five-bladed lectin propellers via oligomeric assembly. We demonstrate a functional trajectory of tandem duplications of these motifs leading to monomeric lectins. Foldability, i.e., higher efficiency of folding, was the main parameter leading to improved functionality along the entire evolutionary trajectory. However, folding constraints changed along the trajectory: initially, conflicts between monomer folding and oligomer assembly dominated, whereas subsequently, upon tandem duplication, tradeoffs between monomer stability and foldability took precedence. PMID:26806127

Programmable biofilm-based materials from engineered curli nanofibres.

PubMed

Nguyen, Peter Q; Botyanszki, Zsofia; Tay, Pei Kun R; Joshi, Neel S

2014-09-17

The significant role of biofilms in pathogenicity has spurred research into preventing their formation and promoting their disruption, resulting in overlooked opportunities to develop biofilms as a synthetic biological platform for self-assembling functional materials. Here we present Biofilm-Integrated Nanofiber Display (BIND) as a strategy for the molecular programming of the bacterial extracellular matrix material by genetically appending peptide domains to the amyloid protein CsgA, the dominant proteinaceous component in Escherichia coli biofilms. These engineered CsgA fusion proteins are successfully secreted and extracellularly self-assemble into amyloid nanofibre networks that retain the functions of the displayed peptide domains. We show the use of BIND to confer diverse artificial functions to the biofilm matrix, such as nanoparticle biotemplating, substrate adhesion, covalent immobilization of proteins or a combination thereof. BIND is a versatile nanobiotechnological platform for developing robust materials with programmable functions, demonstrating the potential of utilizing biofilms as large-scale designable biomaterials.

Biomaterials Made from Coiled-Coil Peptides.

PubMed

Conticello, Vincent; Hughes, Spencer; Modlin, Charles

The development of biomaterials designed for specific applications is an important objective in personalized medicine. While the breadth and prominence of biomaterials have increased exponentially over the past decades, critical challenges remain to be addressed, particularly in the development of biomaterials that exhibit highly specific functions. These functional properties are often encoded within the molecular structure of the component molecules. Proteins, as a consequence of their structural specificity, represent useful substrates for the construction of functional biomaterials through rational design. This chapter provides an in-depth survey of biomaterials constructed from coiled-coils, one of the best-understood protein structural motifs. We discuss the utility of this structurally diverse and functionally tunable class of proteins for the creation of novel biomaterials. This discussion illustrates the progress that has been made in the development of coiled-coil biomaterials by showcasing studies that bridge the gap between the academic science and potential technological impact.

Continually emerging mechanistic complexity of the multi-enzyme cellulosome complex.

PubMed

Smith, Steven P; Bayer, Edward A; Czjzek, Mirjam

2017-06-01

The robust plant cell wall polysaccharide-degrading properties of anaerobic bacteria are harnessed within elegant, marcomolecular assemblages called cellulosomes, in which proteins of complementary activities amass on scaffold protein networks. Research efforts have focused and continue to focus on providing detailed mechanistic insights into cellulosomal complex assembly, topology, and function. The accumulated information is expanding our fundamental understanding of the lignocellulosic biomass decomposition process and enhancing the potential of engineered cellulosomal systems for biotechnological purposes. Ongoing biochemical studies continue to reveal unexpected functional diversity within traditional cellulase families. Genomic, proteomic, and functional analyses have uncovered unanticipated cellulosomal proteins that augment the function of the native and designer cellulosomes. In addition, complementary structural and computational methods are continuing to provide much needed insights on the influence of cellulosomal interdomain linker regions on cellulosomal assembly and activity. Copyright © 2017 Elsevier Ltd. All rights reserved.

Predicting permanent and transient protein-protein interfaces.

PubMed

La, David; Kong, Misun; Hoffman, William; Choi, Youn Im; Kihara, Daisuke

2013-05-01

Protein-protein interactions (PPIs) are involved in diverse functions in a cell. To optimize functional roles of interactions, proteins interact with a spectrum of binding affinities. Interactions are conventionally classified into permanent and transient, where the former denotes tight binding between proteins that result in strong complexes, whereas the latter compose of relatively weak interactions that can dissociate after binding to regulate functional activity at specific time point. Knowing the type of interactions has significant implications for understanding the nature and function of PPIs. In this study, we constructed amino acid substitution models that capture mutation patterns at permanent and transient type of protein interfaces, which were found to be different with statistical significance. Using the substitution models, we developed a novel computational method that predicts permanent and transient protein binding interfaces (PBIs) in protein surfaces. Without knowledge of the interacting partner, the method uses a single query protein structure and a multiple sequence alignment of the sequence family. Using a large dataset of permanent and transient proteins, we show that our method, BindML+, performs very well in protein interface classification. A very high area under the curve (AUC) value of 0.957 was observed when predicted protein binding sites were classified. Remarkably, near prefect accuracy was achieved with an AUC of 0.991 when actual binding sites were classified. The developed method will be also useful for protein design of permanent and transient PBIs. Copyright © 2013 Wiley Periodicals, Inc.

Plant Cation-Chloride Cotransporters (CCC): Evolutionary Origins and Functional Insights.

PubMed

Henderson, Sam W; Wege, Stefanie; Gilliham, Matthew

2018-02-06

Genomes of unicellular and multicellular green algae, mosses, grasses and dicots harbor genes encoding cation-chloride cotransporters (CCC). CCC proteins from the plant kingdom have been comparatively less well investigated than their animal counterparts, but proteins from both plants and animals have been shown to mediate ion fluxes, and are involved in regulation of osmotic processes. In this review, we show that CCC proteins from plants form two distinct phylogenetic clades (CCC1 and CCC2). Some lycophytes and bryophytes possess members from each clade, most land plants only have members of the CCC1 clade, and green algae possess only the CCC2 clade. It is currently unknown whether CCC1 and CCC2 proteins have similar or distinct functions, however they are both more closely related to animal KCC proteins compared to NKCCs. Existing heterologous expression systems that have been used to functionally characterize plant CCC proteins, namely yeast and Xenopus laevis oocytes, have limitations that are discussed. Studies from plants exposed to chemical inhibitors of animal CCC protein function are reviewed for their potential to discern CCC function in planta. Thus far, mutations in plant CCC genes have been evaluated only in two species of angiosperms, and such mutations cause a diverse array of phenotypes-seemingly more than could simply be explained by localized disruption of ion transport alone. We evaluate the putative roles of plant CCC proteins and suggest areas for future investigation.

New Dimensions in Microbial Ecology-Functional Genes in Studies to Unravel the Biodiversity and Role of Functional Microbial Groups in the Environment.

PubMed

Imhoff, Johannes F

2016-05-24

During the past decades, tremendous advances have been made in the possibilities to study the diversity of microbial communities in the environment. The development of methods to study these communities on the basis of 16S rRNA gene sequences analysis was a first step into the molecular analysis of environmental communities and the study of biodiversity in natural habitats. A new dimension in this field was reached with the introduction of functional genes of ecological importance and the establishment of genetic tools to study the diversity of functional microbial groups and their responses to environmental factors. Functional gene approaches are excellent tools to study the diversity of a particular function and to demonstrate changes in the composition of prokaryote communities contributing to this function. The phylogeny of many functional genes largely correlates with that of the 16S rRNA gene, and microbial species may be identified on the basis of functional gene sequences. Functional genes are perfectly suited to link culture-based microbiological work with environmental molecular genetic studies. In this review, the development of functional gene studies in environmental microbiology is highlighted with examples of genes relevant for important ecophysiological functions. Examples are presented for bacterial photosynthesis and two types of anoxygenic phototrophic bacteria, with genes of the Fenna-Matthews-Olson-protein (fmoA) as target for the green sulfur bacteria and of two reaction center proteins (pufLM) for the phototrophic purple bacteria, with genes of adenosine-5'phosphosulfate (APS) reductase (aprA), sulfate thioesterase (soxB) and dissimilatory sulfite reductase (dsrAB) for sulfur oxidizing and sulfate reducing bacteria, with genes of ammonia monooxygenase (amoA) for nitrifying/ammonia-oxidizing bacteria, with genes of particulate nitrate reductase and nitrite reductases (narH/G, nirS, nirK) for denitrifying bacteria and with genes of methane monooxygenase (pmoA) for methane oxidizing bacteria.

Spatially restricted G protein-coupled receptor activity via divergent endocytic compartments.

PubMed

Jean-Alphonse, Frederic; Bowersox, Shanna; Chen, Stanford; Beard, Gemma; Puthenveedu, Manojkumar A; Hanyaloglu, Aylin C

2014-02-14

Postendocytic sorting of G protein-coupled receptors (GPCRs) is driven by their interactions between highly diverse receptor sequence motifs with their interacting proteins, such as postsynaptic density protein (PSD95), Drosophila disc large tumor suppressor (Dlg1), zonula occludens-1 protein (zo-1) (PDZ) domain proteins. However, whether these diverse interactions provide an underlying functional specificity, in addition to driving sorting, is unknown. Here we identify GPCRs that recycle via distinct PDZ ligand/PDZ protein pairs that exploit their recycling machinery primarily for targeted endosomal localization and signaling specificity. The luteinizing hormone receptor (LHR) and β2-adrenergic receptor (B2AR), two GPCRs sorted to the regulated recycling pathway, underwent divergent trafficking to distinct endosomal compartments. Unlike B2AR, which traffics to early endosomes (EE), LHR internalizes to distinct pre-early endosomes (pre-EEs) for its recycling. Pre-EE localization required interactions of the LHR C-terminal tail with the PDZ protein GAIP-interacting protein C terminus, inhibiting its traffic to EEs. Rerouting the LHR to EEs, or EE-localized GPCRs to pre-EEs, spatially reprograms MAPK signaling. Furthermore, LHR-mediated activation of MAPK signaling requires internalization and is maintained upon loss of the EE compartment. We propose that combinatorial specificity between GPCR sorting sequences and interacting proteins dictates an unprecedented spatiotemporal control in GPCR signal activity.

Protein Secretion Systems in Pseudomonas aeruginosa: An Essay on Diversity, Evolution, and Function

PubMed Central

Filloux, Alain

2011-01-01

Protein secretion systems are molecular nanomachines used by Gram-negative bacteria to thrive within their environment. They are used to release enzymes that hydrolyze complex carbon sources into usable compounds, or to release proteins that capture essential ions such as iron. They are also used to colonize and survive within eukaryotic hosts, causing acute or chronic infections, subverting the host cell response and escaping the immune system. In this article, the opportunistic human pathogen Pseudomonas aeruginosa is used as a model to review the diversity of secretion systems that bacteria have evolved to achieve these goals. This diversity may result from a progressive transformation of cell envelope complexes that initially may not have been dedicated to secretion. The striking similarities between secretion systems and type IV pili, flagella, bacteriophage tail, or efflux pumps is a nice illustration of this evolution. Differences are also needed since various secretion configurations call for diversity. For example, some proteins are released in the extracellular medium while others are directly injected into the cytosol of eukaryotic cells. Some proteins are folded before being released and transit into the periplasm. Other proteins cross the whole cell envelope at once in an unfolded state. However, the secretion system requires conserved basic elements or features. For example, there is a need for an energy source or for an outer membrane channel. The structure of this review is thus quite unconventional. Instead of listing secretion types one after each other, it presents a melting pot of concepts indicating that secretion types are in constant evolution and use basic principles. In other words, emergence of new secretion systems could be predicted the way Mendeleïev had anticipated characteristics of yet unknown elements. PMID:21811488

Prediction of scaffold proteins based on protein interaction and domain architectures.

PubMed

Oh, Kimin; Yi, Gwan-Su

2016-07-28

Scaffold proteins are known for being crucial regulators of various cellular functions by assembling multiple proteins involved in signaling and metabolic pathways. Identification of scaffold proteins and the study of their molecular mechanisms can open a new aspect of cellular systemic regulation and the results can be applied in the field of medicine and engineering. Despite being highlighted as the regulatory roles of dozens of scaffold proteins, there was only one known computational approach carried out so far to find scaffold proteins from interactomes. However, there were limitations in finding diverse types of scaffold proteins because their criteria were restricted to the classical scaffold proteins. In this paper, we will suggest a systematic approach to predict massive scaffold proteins from interactomes and to characterize the roles of scaffold proteins comprehensively. From a total of 10,419 basic scaffold protein candidates in protein interactomes, we classified them into three classes according to the structural evidences for scaffolding, such as domain architectures, domain interactions and protein complexes. Finally, we could define 2716 highly reliable scaffold protein candidates and their characterized functional features. To assess the accuracy of our prediction, the gold standard positive and negative data sets were constructed. We prepared 158 gold standard positive data and 844 gold standard negative data based on the functional information from Gene Ontology consortium. The precision, sensitivity and specificity of our testing was 80.3, 51.0, and 98.5 % respectively. Through the function enrichment analysis of highly reliable scaffold proteins, we could confirm the significantly enriched functions that are related to scaffold protein binding. We also identified functional association between scaffold proteins and their recruited proteins. Furthermore, we checked that the disease association of scaffold proteins is higher than kinases. In conclusion, we could predict larger volume of scaffold proteins and analyzed their functional characteristics. Deeper understandings about the roles of scaffold proteins from this study will provide a higher opportunity to find therapeutic or engineering applications of scaffold proteins using their functional characteristics.

Ligand-specific regulation of the extracellular surface of a G-protein-coupled receptor

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bokoch, Michael P.; Zou, Yaozhong; Rasmussen, Søren G.F.

G-protein-coupled receptors (GPCRs) are seven-transmembrane proteins that mediate most cellular responses to hormones and neurotransmitters. They are the largest group of therapeutic targets for a broad spectrum of diseases. Recent crystal structures of GPCRs have revealed structural conservation extending from the orthosteric ligand-binding site in the transmembrane core to the cytoplasmic G-protein-coupling domains. In contrast, the extracellular surface (ECS) of GPCRs is remarkably diverse and is therefore an ideal target for the discovery of subtype-selective drugs. However, little is known about the functional role of the ECS in receptor activation, or about conformational coupling of this surface to the nativemore » ligand-binding pocket. Here we use NMR spectroscopy to investigate ligand-specific conformational changes around a central structural feature in the ECS of the {beta}{sub 2} adrenergic receptor: a salt bridge linking extracellular loops 2 and 3. Small-molecule drugs that bind within the transmembrane core and exhibit different efficacies towards G-protein activation (agonist, neutral antagonist and inverse agonist) also stabilize distinct conformations of the ECS. We thereby demonstrate conformational coupling between the ECS and the orthosteric binding site, showing that drugs targeting this diverse surface could function as allosteric modulators with high subtype selectivity. Moreover, these studies provide a new insight into the dynamic behaviour of GPCRs not addressable by static, inactive-state crystal structures.« less

Interplay between intrinsic noise and the stochasticity of the cell cycle in bacterial colonies.

PubMed

Canela-Xandri, Oriol; Sagués, Francesc; Buceta, Javier

2010-06-02

Herein we report on the effects that different stochastic contributions induce in bacterial colonies in terms of protein concentration and production. In particular, we consider for what we believe to be the first time cell-to-cell diversity due to the unavoidable randomness of the cell-cycle duration and its interplay with other noise sources. To that end, we model a recent experimental setup that implements a protein dilution protocol by means of division events to characterize the gene regulatory function at the single cell level. This approach allows us to investigate the effect of different stochastic terms upon the total randomness experimentally reported for the gene regulatory function. In addition, we show that the interplay between intrinsic fluctuations and the stochasticity of the cell-cycle duration leads to different constructive roles. On the one hand, we show that there is an optimal value of protein concentration (alternatively an optimal value of the cell cycle phase) such that the noise in protein concentration attains a minimum. On the other hand, we reveal that there is an optimal value of the stochasticity of the cell cycle duration such that the coherence of the protein production with respect to the colony average production is maximized. The latter can be considered as a novel example of the recently reported phenomenon of diversity induced resonance. Copyright (c) 2010 Biophysical Society. Published by Elsevier Inc. All rights reserved.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lammert, Heiko; Noel, Jeffrey K.; Haglund, Ellinor

The diversity in a set of protein nuclear magnetic resonance (NMR) structures provides an estimate of native state fluctuations that can be used to refine and enrich structure-based protein models (SBMs). Dynamics are an essential part of a protein’s functional native state. The dynamics in the native state are controlled by the same funneled energy landscape that guides the entire folding process. SBMs apply the principle of minimal frustration, drawn from energy landscape theory, to construct a funneled folding landscape for a given protein using only information from the native structure. On an energy landscape smoothed by evolution towards minimalmore » frustration, geometrical constraints, imposed by the native structure, control the folding mechanism and shape the native dynamics revealed by the model. Native-state fluctuations can alternatively be estimated directly from the diversity in the set of NMR structures for a protein. Based on this information, we identify a highly flexible loop in the ribosomal protein S6 and modify the contact map in a SBM to accommodate the inferred dynamics. By taking into account the probable native state dynamics, the experimental transition state is recovered in the model, and the correct order of folding events is restored. Our study highlights how the shared energy landscape connects folding and function by showing that a better description of the native basin improves the prediction of the folding mechanism.« less

A structural analysis of the AAA+ domains in Saccharomyces cerevisiae cytoplasmic dynein.

PubMed

Gleave, Emma S; Schmidt, Helgo; Carter, Andrew P

2014-06-01

Dyneins are large protein complexes that act as microtubule based molecular motors. The dynein heavy chain contains a motor domain which is a member of the AAA+ protein family (ATPases Associated with diverse cellular Activities). Proteins of the AAA+ family show a diverse range of functionalities, but share a related core AAA+ domain, which often assembles into hexameric rings. Dynein is unusual because it has all six AAA+ domains linked together, in one long polypeptide. The dynein motor domain generates movement by coupling ATP driven conformational changes in the AAA+ ring to the swing of a motile element called the linker. Dynein binds to its microtubule track via a long antiparallel coiled-coil stalk that emanates from the AAA+ ring. Recently the first high resolution structures of the dynein motor domain were published. Here we provide a detailed structural analysis of the six AAA+ domains using our Saccharomycescerevisiae crystal structure. We describe how structural similarities in the dynein AAA+ domains suggest they share a common evolutionary origin. We analyse how the different AAA+ domains have diverged from each other. We discuss how this is related to the function of dynein as a motor protein and how the AAA+ domains of dynein compare to those of other AAA+ proteins. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

Frameshifting in alphaviruses: a diversity of 3' stimulatory structures.

PubMed

Chung, Betty Y-W; Firth, Andrew E; Atkins, John F

2010-03-26

Programmed ribosomal frameshifting allows the synthesis of alternative, N-terminally coincident, C-terminally distinct proteins from the same RNA. Many viruses utilize frameshifting to optimize the coding potential of compact genomes, to circumvent the host cell's canonical rule of one functional protein per mRNA, or to express alternative proteins in a fixed ratio. Programmed frameshifting is also used in the decoding of a small number of cellular genes. Recently, specific ribosomal -1 frameshifting was discovered at a conserved U_UUU_UUA motif within the sequence encoding the alphavirus 6K protein. In this case, frameshifting results in the synthesis of an additional protein, termed TF (TransFrame). This new case of frameshifting is unusual in that the -1 frame ORF is very short and completely embedded within the sequence encoding the overlapping polyprotein. The present work shows that there is remarkable diversity in the 3' sequences that are functionally important for efficient frameshifting at the U_UUU_UUA motif. While many alphavirus species utilize a 3' RNA structure such as a hairpin or pseudoknot, some species (such as Semliki Forest virus) apparently lack any intra-mRNA stimulatory structure, yet just 20 nt 3'-adjacent to the shift site stimulates up to 10% frameshifting. The analysis, both experimental and bioinformatic, significantly expands the known repertoire of -1 frameshifting stimulators in mammalian and insect systems.

Phylogeny-Based Systematization of Arabidopsis Proteins with Histone H1 Globular Domain1[OPEN

PubMed Central

Knizewski, Lukasz; Schmidt, Anja; Ginalski, Krzysztof

2017-01-01

H1 (or linker) histones are basic nuclear proteins that possess an evolutionarily conserved nucleosome-binding globular domain, GH1. They perform critical functions in determining the accessibility of chromatin DNA to trans-acting factors. In most metazoan species studied so far, linker histones are highly heterogenous, with numerous nonallelic variants cooccurring in the same cells. The phylogenetic relationships among these variants as well as their structural and functional properties have been relatively well established. This contrasts markedly with the rather limited knowledge concerning the phylogeny and structural and functional roles of an unusually diverse group of GH1-containing proteins in plants. The dearth of information and the lack of a coherent phylogeny-based nomenclature of these proteins can lead to misunderstandings regarding their identity and possible relationships, thereby hampering plant chromatin research. Based on published data and our in silico and high-throughput analyses, we propose a systematization and coherent nomenclature of GH1-containing proteins of Arabidopsis (Arabidopsis thaliana [L.] Heynh) that will be useful for both the identification and structural and functional characterization of homologous proteins from other plant species. PMID:28298478

Phylogeny-Based Systematization of Arabidopsis Proteins with Histone H1 Globular Domain.

PubMed

Kotliński, Maciej; Knizewski, Lukasz; Muszewska, Anna; Rutowicz, Kinga; Lirski, Maciej; Schmidt, Anja; Baroux, Célia; Ginalski, Krzysztof; Jerzmanowski, Andrzej

2017-05-01

H1 (or linker) histones are basic nuclear proteins that possess an evolutionarily conserved nucleosome-binding globular domain, GH1. They perform critical functions in determining the accessibility of chromatin DNA to trans-acting factors. In most metazoan species studied so far, linker histones are highly heterogenous, with numerous nonallelic variants cooccurring in the same cells. The phylogenetic relationships among these variants as well as their structural and functional properties have been relatively well established. This contrasts markedly with the rather limited knowledge concerning the phylogeny and structural and functional roles of an unusually diverse group of GH1-containing proteins in plants. The dearth of information and the lack of a coherent phylogeny-based nomenclature of these proteins can lead to misunderstandings regarding their identity and possible relationships, thereby hampering plant chromatin research. Based on published data and our in silico and high-throughput analyses, we propose a systematization and coherent nomenclature of GH1-containing proteins of Arabidopsis ( Arabidopsis thaliana [L.] Heynh) that will be useful for both the identification and structural and functional characterization of homologous proteins from other plant species. © 2017 American Society of Plant Biologists. All Rights Reserved.

Proteomic analysis reveals diverse proline hydroxylation-mediated oxygen-sensing cellular pathways in cancer cells

PubMed Central

Liu, Bing; Gao, Yankun; Ruan, Hai-Bin; Chen, Yue

2016-01-01

Proline hydroxylation is a critical cellular mechanism regulating oxygen-response pathways in tumor initiation and progression. Yet, its substrate diversity and functions remain largely unknown. Here, we report a system-wide analysis to characterize proline hydroxylation substrates in cancer cells using an immunoaffinity-purification assisted proteomics strategy. We identified 562 sites from 272 proteins in HeLa cells. Bioinformatic analysis revealed that proline hydroxylation substrates are significantly enriched with mRNA processing and stress-response cellular pathways with canonical and diverse flanking sequence motifs. Structural analysis indicates a significant enrichment of proline hydroxylation participating in the secondary structure of substrate proteins. Our study identified and validated Brd4, a key transcription factor, as a novel proline hydroxylation substrate. Functional analysis showed that the inhibition of proline hydroxylation pathway significantly reduced the proline hydroxylation abundance on Brd4 and affected Brd4-mediated transcriptional activity as well as cell proliferation in AML leukemia cells. Taken together, our study identified a broad regulatory role of proline hydroxylation in cellular oxygen-sensing pathways and revealed potentially new targets that dynamically respond to hypoxia microenvironment in tumor cells. PMID:27764789

Chlorella vulgaris: A Multifunctional Dietary Supplement with Diverse Medicinal Properties.

PubMed

Panahi, Yunes; Darvishi, Behrad; Jowzi, Narges; Beiraghdar, Fatemeh; Sahebkar, Amirhossein

2016-01-01

Chlorella vulgaris is a green unicellular microalgae with biological and pharmacological properties important for human health. C. vulgaris has a long history of use as a food source and contains a unique and diverse composition of functional macro- and micro-nutrients including proteinsChlorella vulgaris is a green unicellular microalgae with biological and pharmacological properties important for human health. C. vulgaris has a long history of use as a food source and contains a unique and diverse composition of functional macro- and micro-nutrients including proteins, omega-3 polyunsaturated fatty acids, polysaccharides, vitamins and minerals. Clinical trials have suggested that supplementation with C. vulgaris can ameliorate amelioration hyperlipidemia and hyperglycemia, and protect against oxidative stress, cancer and chronic obstructive pulmonary disease. In this review, we summarize the findings on the health benefits of Chlorella supplementation and the molecular mechanisms underlying these effects., omega-3 polyunsaturated fatty acids, polysaccharides, vitamins and minerals. Clinical trials have suggested that supplementation with C. vulgaris can ameliorate amelioration hyperlipidemia and hyperglycemia, and protect against oxidative stress, cancer and chronic obstructive pulmonary disease. In this review, we summarize the findings on the health benefits of Chlorella supplementation and the molecular mechanisms underlying these effects.

RAID: a comprehensive resource for human RNA-associated (RNA–RNA/RNA–protein) interaction

PubMed Central

Zhang, Xiaomeng; Wu, Deng; Chen, Liqun; Li, Xiang; Yang, Jinxurong; Fan, Dandan; Dong, Tingting; Liu, Mingyue; Tan, Puwen; Xu, Jintian; Yi, Ying; Wang, Yuting; Zou, Hua; Hu, Yongfei; Fan, Kaili; Kang, Juanjuan; Huang, Yan; Miao, Zhengqiang; Bi, Miaoman; Jin, Nana; Li, Kongning; Li, Xia; Xu, Jianzhen; Wang, Dong

2014-01-01

Transcriptomic analyses have revealed an unexpected complexity in the eukaryote transcriptome, which includes not only protein-coding transcripts but also an expanding catalog of noncoding RNAs (ncRNAs). Diverse coding and noncoding RNAs (ncRNAs) perform functions through interaction with each other in various cellular processes. In this project, we have developed RAID (http://www.rna-society.org/raid), an RNA-associated (RNA–RNA/RNA–protein) interaction database. RAID intends to provide the scientific community with all-in-one resources for efficient browsing and extraction of the RNA-associated interactions in human. This version of RAID contains more than 6100 RNA-associated interactions obtained by manually reviewing more than 2100 published papers, including 4493 RNA–RNA interactions and 1619 RNA–protein interactions. Each entry contains detailed information on an RNA-associated interaction, including RAID ID, RNA/protein symbol, RNA/protein categories, validated method, expressing tissue, literature references (Pubmed IDs), and detailed functional description. Users can query, browse, analyze, and manipulate RNA-associated (RNA–RNA/RNA–protein) interaction. RAID provides a comprehensive resource of human RNA-associated (RNA–RNA/RNA–protein) interaction network. Furthermore, this resource will help in uncovering the generic organizing principles of cellular function network. PMID:24803509

Chromatin-Bound Cullin-Ring Ligases: Regulatory Roles in DNA Replication and Potential Targeting for Cancer Therapy

PubMed Central

Jang, Sang-Min; Redon, Christophe E.; Aladjem, Mirit I.

2018-01-01

Cullin-RING (Really Interesting New Gene) E3 ubiquitin ligases (CRLs), the largest family of E3 ubiquitin ligases, are functional multi-subunit complexes including substrate receptors, adaptors, cullin scaffolds, and RING-box proteins. CRLs are responsible for ubiquitination of ~20% of cellular proteins and are involved in diverse biological processes including cell cycle progression, genome stability, and oncogenesis. Not surprisingly, cullins are deregulated in many diseases and instances of cancer. Recent studies have highlighted the importance of CRL-mediated ubiquitination in the regulation of DNA replication/repair, including specific roles in chromatin assembly and disassembly of the replication machinery. The development of novel therapeutics targeting the CRLs that regulate the replication machinery and chromatin in cancer is now an attractive therapeutic strategy. In this review, we summarize the structure and assembly of CRLs and outline their cellular functions and their diverse roles in cancer, emphasizing the regulatory functions of nuclear CRLs in modulating the DNA replication machinery. Finally, we discuss the current strategies for targeting CRLs against cancer in the clinic. PMID:29594129

Genetics of Mitochondrial Disease.

PubMed

Saneto, Russell P

2017-01-01

Mitochondria are intracellular organelles responsible for adenosine triphosphate production. The strict control of intracellular energy needs require proper mitochondrial functioning. The mitochondria are under dual controls of mitochondrial DNA (mtDNA) and nuclear DNA (nDNA). Mitochondrial dysfunction can arise from changes in either mtDNA or nDNA genes regulating function. There are an estimated ∼1500 proteins in the mitoproteome, whereas the mtDNA genome has 37 proteins. There are, to date, ∼275 genes shown to give rise to disease. The unique physiology of mitochondrial functioning contributes to diverse gene expression. The onset and range of phenotypic expression of disease is diverse, with onset from neonatal to seventh decade of life. The range of dysfunction is heterogeneous, ranging from single organ to multisystem involvement. The complexity of disease expression has severely limited gene discovery. Combining phenotypes with improvements in gene sequencing strategies are improving the diagnosis process. This chapter focuses on the interplay of the unique physiology and gene discovery in the current knowledge of genetically derived mitochondrial disease. Copyright © 2017 Elsevier Inc. All rights reserved.

Diversity in recognition of glycans by F-type lectins and galectins: molecular, structural, and biophysical aspects

PubMed Central

Vasta, Gerardo R.; Ahmed, Hafiz; Bianchet, Mario A.; Fernández-Robledo, José A.; Amzel, L. Mario

2013-01-01

Although lectins are “hard-wired” in the germline, the presence of tandemly arrayed carbohydrate recognition domains (CRDs), of chimeric structures displaying distinct CRDs, of polymorphic genes resulting in multiple isoforms, and in some cases, of a considerable recognition plasticity of their carbohydrate binding sites, significantly expand the lectin ligand-recognition spectrum and lectin functional diversification. Analysis of structural/functional aspects of galectins and F-lectins—the most recently identified lectin family characterized by a unique CRD sequence motif (a distinctive structural fold) and nominal specificity for l-Fuc—has led to a greater understanding of self/nonself recognition by proteins with tandemly arrayed CRDs. For lectins with a single CRD, however, recognition of self and nonself glycans can only be rationalized in terms of protein oligomerization and ligand clustering and presentation. Spatial and temporal changes in lectin expression, secretion, and local concentrations in extracellular microenvironments, as well as structural diversity and spatial display of their carbohydrate ligands on the host or microbial cell surface, are suggestive of a dynamic interplay of their recognition and effector functions in development and immunity. PMID:22973821

Molecular chaperones and photoreceptor function

PubMed Central

Kosmaoglou, Maria; Schwarz, Nele; Bett, John S.; Cheetham, Michael E.

2008-01-01

Molecular chaperones facilitate and regulate protein conformational change within cells. This encompasses many fundamental cellular processes: including the correct folding of nascent chains; protein transport and translocation; signal transduction and protein quality control. Chaperones are, therefore, important in several forms of human disease, including neurodegeneration. Within the retina, the highly specialized photoreceptor cell presents a fascinating paradigm to investigate the specialization of molecular chaperone function and reveals unique chaperone requirements essential to photoreceptor function. Mutations in several photoreceptor proteins lead to protein misfolding mediated neurodegeneration. The best characterized of these are mutations in the molecular light sensor, rhodopsin, which cause autosomal dominant retinitis pigmentosa. Rhodopsin biogenesis is likely to require chaperones, while rhodopsin misfolding involves molecular chaperones in quality control and the cellular response to protein aggregation. Furthermore, the specialization of components of the chaperone machinery to photoreceptor specific roles has been revealed by the identification of mutations in molecular chaperones that cause inherited retinal dysfunction and degeneration. These chaperones are involved in several important cellular pathways and further illuminate the essential and diverse roles of molecular chaperones. PMID:18490186

Prohibitin( PHB) roles in granulosa cell physiology.

PubMed

Chowdhury, Indrajit; Thomas, Kelwyn; Thompson, Winston E

2016-01-01

Ovarian granulosa cells (GC) play an important role in the growth and development of the follicle in the process known as folliculogenesis. In the present review, we focus on recent developments in prohibitin (PHB) research in relation to GC physiological functions. PHB is a member of a highly conserved eukaryotic protein family containing the repressor of estrogen activity (REA)/stomatin/PHB/flotillin/HflK/C (SPFH) domain (also known as the PHB domain) found in diverse species from prokaryotes to eukaryotes. PHB is ubiquitously expressed in a circulating free form or is present in multiple cellular compartments including mitochondria, nucleus and plasma membrane. In mitochondria, PHB is anchored to the mitochondrial inner membrane and forms complexes with the ATPases associated with proteases having diverse cellular activities. PHB continuously shuttles between the mitochondria, cytosol and nucleus. In the nucleus, PHB interacts with various transcription factors and modulates transcriptional activity directly or through interactions with chromatin remodeling proteins. Many functions have been attributed to the mitochondrial and nuclear PHB complexes such as cellular differentiation, anti-proliferation, morphogenesis and maintenance of the functional integrity of the mitochondria. However, to date, the regulation of PHB expression patterns and GC physiological functions are not completely understood.

Functional requirements for bacteriophage growth: gene essentiality and expression in mycobacteriophage Giles.

PubMed

Dedrick, Rebekah M; Marinelli, Laura J; Newton, Gerald L; Pogliano, Kit; Pogliano, Joseph; Hatfull, Graham F

2013-05-01

Bacteriophages represent a majority of all life forms, and the vast, dynamic population with early origins is reflected in their enormous genetic diversity. A large number of bacteriophage genomes have been sequenced. They are replete with novel genes without known relatives. We know little about their functions, which genes are required for lytic growth, and how they are expressed. Furthermore, the diversity is such that even genes with required functions - such as virion proteins and repressors - cannot always be recognized. Here we describe a functional genomic dissection of mycobacteriophage Giles, in which the virion proteins are identified, genes required for lytic growth are determined, the repressor is identified, and the transcription patterns determined. We find that although all of the predicted phage genes are expressed either in lysogeny or in lytic growth, 45% of the predicted genes are non-essential for lytic growth. We also describe genes required for DNA replication, show that recombination is required for lytic growth, and that Giles encodes a novel repressor. RNAseq analysis reveals abundant expression of a small non-coding RNA in a lysogen and in late lytic growth, although it is non-essential for lytic growth and does not alter lysogeny. © 2013 Blackwell Publishing Ltd.

The evolution of compositionally and functionally distinct actin filaments.

PubMed

Gunning, Peter W; Ghoshdastider, Umesh; Whitaker, Shane; Popp, David; Robinson, Robert C

2015-06-01

The actin filament is astonishingly well conserved across a diverse set of eukaryotic species. It has essentially remained unchanged in the billion years that separate yeast, Arabidopsis and man. In contrast, bacterial actin-like proteins have diverged to the extreme, and many of them are not readily identified from sequence-based homology searches. Here, we present phylogenetic analyses that point to an evolutionary drive to diversify actin filament composition across kingdoms. Bacteria use a one-filament-one-function system to create distinct filament systems within a single cell. In contrast, eukaryotic actin is a universal force provider in a wide range of processes. In plants, there has been an expansion of the number of closely related actin genes, whereas in fungi and metazoa diversification in tropomyosins has increased the compositional variety in actin filament systems. Both mechanisms dictate the subset of actin-binding proteins that interact with each filament type, leading to specialization in function. In this Hypothesis, we thus propose that different mechanisms were selected in bacteria, plants and metazoa, which achieved actin filament compositional variation leading to the expansion of their functional diversity. © 2015. Published by The Company of Biologists Ltd.

Proteomic Stable Isotope Probing Reveals Biosynthesis Dynamics of Slow Growing Methane Based Microbial Communities

DOE PAGES

Marlow, Jeffery; Skennerton, Connor T.; Li, Zhou; ...

2016-04-29

Marine methane seep habitats represent an important control on the global flux of methane between the subsurface and water column reservoirs. Meta-omics studies have begun to outline community-wide metabolic potential, but expression patterns of proteins that enact sulfate-mediated anaerobic methane oxidation in seeps are poorly characterized. Proteomic stable isotope probing (proteomic SIP) offers an additional layer of information for characterizing phylogenetically specific, functionally relevant activity in mixed microbial communities. Here we applied proteomic SIP to 15NH4+ and CH4 amended seep sediment microcosms in an attempt to track the protein synthesis of slow-growing, low-energy microbial systems. Across all samples, 3495 proteinsmore » were identified, 21% of which were 15N-labeled. We observed active synthesis (15N enrichment) of all proteins believed to be involved in sulfate reduction and reverse methanogenesis including methylenetetrahydromethanopterin reductase (Mer). The abundance and phylogenetic range of methyl-coenzyme M reductase (Mcr) orthologs produced during incubation experiments suggests that seeps provide sufficient niches for multiple organisms performing analogous metabolisms. Twenty-eight previously unreported post-translational modifications of McrA were measured, indicating dynamic enzymatic machinery and offering a dimension of functional diversity beyond gene-dictated sequence. RNA polymerase associated with putative sulfur-oxidizing Epsilonproteobacteria and aerobic Gammaproteobacteria were more abundant among pre-incubation proteins, suggesting diminished metabolic activity in long-term anoxic, sulfidic experimental incubations. Twenty-six proteins of unknown function were detected in all proteomic experiments and actively expressed in labeled experiments, suggesting that they play important roles in methane seep ecosystems. The addition of stable isotope probing to environmental proteomics experiments provides a mechanism to begin to assess the degree to which diagnostic meatbolic proteins are long-lived or acively synthesized in complex, slow-growing microbial communities. Our work here demonstrates that sediment-hosted microbial assemblages in marine methane seeps are dynamic, heterogeneous systems with broad functional diversity.« less

Proteomic Stable Isotope Probing Reveals Biosynthesis Dynamics of Slow Growing Methane Based Microbial Communities

DOE Office of Scientific and Technical Information (OSTI.GOV)

Marlow, Jeffery; Skennerton, Connor T.; Li, Zhou

Marine methane seep habitats represent an important control on the global flux of methane between the subsurface and water column reservoirs. Meta-omics studies have begun to outline community-wide metabolic potential, but expression patterns of proteins that enact sulfate-mediated anaerobic methane oxidation in seeps are poorly characterized. Proteomic stable isotope probing (proteomic SIP) offers an additional layer of information for characterizing phylogenetically specific, functionally relevant activity in mixed microbial communities. Here we applied proteomic SIP to 15NH4+ and CH4 amended seep sediment microcosms in an attempt to track the protein synthesis of slow-growing, low-energy microbial systems. Across all samples, 3495 proteinsmore » were identified, 21% of which were 15N-labeled. We observed active synthesis (15N enrichment) of all proteins believed to be involved in sulfate reduction and reverse methanogenesis including methylenetetrahydromethanopterin reductase (Mer). The abundance and phylogenetic range of methyl-coenzyme M reductase (Mcr) orthologs produced during incubation experiments suggests that seeps provide sufficient niches for multiple organisms performing analogous metabolisms. Twenty-eight previously unreported post-translational modifications of McrA were measured, indicating dynamic enzymatic machinery and offering a dimension of functional diversity beyond gene-dictated sequence. RNA polymerase associated with putative sulfur-oxidizing Epsilonproteobacteria and aerobic Gammaproteobacteria were more abundant among pre-incubation proteins, suggesting diminished metabolic activity in long-term anoxic, sulfidic experimental incubations. Twenty-six proteins of unknown function were detected in all proteomic experiments and actively expressed in labeled experiments, suggesting that they play important roles in methane seep ecosystems. The addition of stable isotope probing to environmental proteomics experiments provides a mechanism to begin to assess the degree to which diagnostic meatbolic proteins are long-lived or acively synthesized in complex, slow-growing microbial communities. Our work here demonstrates that sediment-hosted microbial assemblages in marine methane seeps are dynamic, heterogeneous systems with broad functional diversity.« less

Designing ECM-mimetic Materials Using Protein Engineering

PubMed Central

Cai, Lei; Heilshorn, Sarah C.

2014-01-01

The natural extracellular matrix (ECM), with its multitude of evolved cell-instructive and cell-responsive properties, provides inspiration and guidelines for the design of engineered biomaterials. One strategy to create ECM-mimetic materials is the modular design of protein-based engineered ECM (eECM) scaffolds. This modular design strategy involves combining multiple protein domains with different functionalities into a single, modular polymer sequence, resulting in a multifunctional matrix with independent tunability of the individual domain functions. These eECMs often enable decoupled control over multiple material properties for fundamental studies of cell-matrix interactions. In addition, since the eECMs are frequently composed entirely of bioresorbable amino acids, these matrices have immense clinical potential for a variety of regenerative medicine applications. This brief review demonstrates how fundamental knowledge gained from structure-function studies of native proteins can be exploited in the design of novel protein-engineered biomaterials. While the field of protein-engineered biomaterials has existed for over 20 years, the community is only now beginning to fully explore the diversity of functional peptide modules that can be incorporated into these materials. We have chosen to highlight recent examples that either (1) demonstrate exemplary use as matrices with cell-instructive and cell-responsive properties or (2) demonstrate outstanding creativity in terms of novel molecular-level design and macro-level functionality. PMID:24365704

Diverse mechanisms of metaeffector activity in an intracellular bacterial pathogen, Legionella pneumophila

DOE PAGES

Urbanus, Malene L.; Quaile, Andrew T.; Stogios, Peter J.; ...

2016-12-16

Pathogens deliver complex arsenals of translocated effector proteins to host cells during infection, but the extent to which these proteins are regulated once inside the eukaryotic cell remains poorly defined. Among all bacterial pathogens, Legionella pneumophila maintains the largest known set of translocated substrates, delivering over 300 proteins to the host cell via its Type IVB, Icm/Dot translocation system. Backed by a few notable examples of effector–effector regulation in L. pneumophila, we sought to define the extent of this phenomenon through a systematic analysis of effector–effector functional interaction. We used Saccharomyces cerevisiae, an established proxy for the eukaryotic host, tomore » query > 108,000 pairwise genetic interactions between two compatible expression libraries of ~330 L. pneumophila–translocated substrates. While capturing all known examples of effector–effector suppression, we identify fourteen novel translocated substrates that suppress the activity of other bacterial effectors and one pair with synergistic activities. In at least nine instances, this regulation is direct—a hallmark of an emerging class of proteins called metaeffectors, or “effectors of effectors”. Through detailed structural and functional analysis, we show that metaeffector activity derives from a diverse range of mechanisms, shapes evolution, and can be used to reveal important aspects of each cognate effector's function. Here, metaeffectors, along with other, indirect, forms of effector–effector modulation, may be a common feature of many intracellular pathogens—with unrealized potential to inform our understanding of how pathogens regulate their interactions with the host cell.« less

Discovering Protein-Coding Genes from the Environment: Time for the Eukaryotes?

PubMed

Marmeisse, Roland; Kellner, Harald; Fraissinet-Tachet, Laurence; Luis, Patricia

2017-09-01

Eukaryotic microorganisms from diverse environments encompass a large number of taxa, many of them still unknown to science. One strategy to mine these organisms for genes of biotechnological relevance is to use a pool of eukaryotic mRNA directly extracted from environmental samples. Recent reports demonstrate that the resulting metatranscriptomic cDNA libraries can be screened by expression in yeast for a wide range of genes and functions from many of the different eukaryotic taxa. In combination with novel emerging high-throughput technologies, we anticipate that this approach should contribute to exploring the functional diversity of the eukaryotic microbiota. Copyright © 2017 Elsevier Ltd. All rights reserved.

Navigating ligand protein binding free energy landscapes: universality and diversity of protein folding and molecular recognition mechanisms

NASA Astrophysics Data System (ADS)

Verkhivker, Gennady M.; Rejto, Paul A.; Bouzida, Djamal; Arthurs, Sandra; Colson, Anthony B.; Freer, Stephan T.; Gehlhaar, Daniel K.; Larson, Veda; Luty, Brock A.; Marrone, Tami; Rose, Peter W.

2001-03-01

Thermodynamic and kinetic aspects of ligand-protein binding are studied for the methotrexate-dihydrofolate reductase system from the binding free energy profile constructed as a function of the order parameter. Thermodynamic stability of the native complex and a cooperative transition to the unique native structure suggest the nucleation kinetic mechanism at the equilibrium transition temperature. Structural properties of the transition state ensemble and the ensemble of nucleation conformations are determined by kinetic simulations of the transmission coefficient and ligand-protein association pathways. Structural analysis of the transition states and the nucleation conformations reconciles different views on the nucleation mechanism in protein folding.

The Coat Protein and NIa Protease of Two Potyviridae Family Members Independently Confer Superinfection Exclusion

PubMed Central

French, Roy

2016-01-01

ABSTRACT Superinfection exclusion (SIE) is an antagonistic virus-virus interaction whereby initial infection by one virus prevents subsequent infection by closely related viruses. Although SIE has been described in diverse viruses infecting plants, humans, and animals, its mechanisms, including involvement of specific viral determinants, are just beginning to be elucidated. In this study, SIE determinants encoded by two economically important wheat viruses, Wheat streak mosaic virus (WSMV; genus Tritimovirus, family Potyviridae) and Triticum mosaic virus (TriMV; genus Poacevirus, family Potyviridae), were identified in gain-of-function experiments that used heterologous viruses to express individual virus-encoded proteins in wheat. Wheat plants infected with TriMV expressing WSMV P1, HC-Pro, P3, 6K1, CI, 6K2, NIa-VPg, or NIb cistrons permitted efficient superinfection by WSMV expressing green fluorescent protein (WSMV-GFP). In contrast, wheat infected with TriMV expressing WSMV NIa-Pro or coat protein (CP) substantially excluded superinfection by WSMV-GFP, suggesting that both of these cistrons are SIE effectors encoded by WSMV. Importantly, SIE is due to functional WSMV NIa-Pro or CP rather than their encoding RNAs, as altering the coded protein products by minimally changing RNA sequences led to abolishment of SIE. Deletion mutagenesis further revealed that elicitation of SIE by NIa-Pro requires the entire protein while CP requires only a 200-amino-acid (aa) middle fragment (aa 101 to 300) of the 349 aa. Strikingly, reciprocal experiments with WSMV-mediated expression of TriMV proteins showed that TriMV CP, and TriMV NIa-Pro to a lesser extent, likewise excluded superinfection by TriMV-GFP. Collectively, these data demonstrate that WSMV- and TriMV-encoded CP and NIa-Pro proteins are effectors of SIE and that these two proteins trigger SIE independently of each other. IMPORTANCE Superinfection exclusion (SIE) is an antagonistic virus-virus interaction that prevents secondary invasions by identical or closely related viruses in the same host cells. Although known to occur in diverse viruses, SIE remains an enigma in terms of key molecular determinants and action mechanisms. In this study, we found that Wheat streak mosaic virus (WSMV) and Triticum mosaic virus (TriMV) encode two independently functioning cistrons that serve as effectors of SIE at the protein but not the RNA level. The coat protein and NIa-Pro encoded by these two viruses, when expressed from a heterologous virus, exerted SIE to the cognate viruses. The identification of virus-encoded effectors of SIE and their transgenic expression could potentially facilitate the development of virus-resistant crop plants. Additionally, functional conservation of SIE in diverse virus groups suggests that a better understanding of the underlying mechanisms of SIE could facilitate the development of novel antiviral therapies against viral diseases. PMID:27681136

Zinc finger proteins in cancer progression.

PubMed

Jen, Jayu; Wang, Yi-Ching

2016-07-13

Zinc finger proteins are the largest transcription factor family in human genome. The diverse combinations and functions of zinc finger motifs make zinc finger proteins versatile in biological processes, including development, differentiation, metabolism and autophagy. Over the last few decades, increasing evidence reveals the potential roles of zinc finger proteins in cancer progression. However, the underlying mechanisms of zinc finger proteins in cancer progression vary in different cancer types and even in the same cancer type under different types of stress. Here, we discuss general mechanisms of zinc finger proteins in transcription regulation and summarize recent studies on zinc finger proteins in cancer progression. In this review, we also emphasize the importance of further investigations in elucidating the underlying mechanisms of zinc finger proteins in cancer progression.

Analysis of protein-coding genetic variation in 60,706 humans.

PubMed

Lek, Monkol; Karczewski, Konrad J; Minikel, Eric V; Samocha, Kaitlin E; Banks, Eric; Fennell, Timothy; O'Donnell-Luria, Anne H; Ware, James S; Hill, Andrew J; Cummings, Beryl B; Tukiainen, Taru; Birnbaum, Daniel P; Kosmicki, Jack A; Duncan, Laramie E; Estrada, Karol; Zhao, Fengmei; Zou, James; Pierce-Hoffman, Emma; Berghout, Joanne; Cooper, David N; Deflaux, Nicole; DePristo, Mark; Do, Ron; Flannick, Jason; Fromer, Menachem; Gauthier, Laura; Goldstein, Jackie; Gupta, Namrata; Howrigan, Daniel; Kiezun, Adam; Kurki, Mitja I; Moonshine, Ami Levy; Natarajan, Pradeep; Orozco, Lorena; Peloso, Gina M; Poplin, Ryan; Rivas, Manuel A; Ruano-Rubio, Valentin; Rose, Samuel A; Ruderfer, Douglas M; Shakir, Khalid; Stenson, Peter D; Stevens, Christine; Thomas, Brett P; Tiao, Grace; Tusie-Luna, Maria T; Weisburd, Ben; Won, Hong-Hee; Yu, Dongmei; Altshuler, David M; Ardissino, Diego; Boehnke, Michael; Danesh, John; Donnelly, Stacey; Elosua, Roberto; Florez, Jose C; Gabriel, Stacey B; Getz, Gad; Glatt, Stephen J; Hultman, Christina M; Kathiresan, Sekar; Laakso, Markku; McCarroll, Steven; McCarthy, Mark I; McGovern, Dermot; McPherson, Ruth; Neale, Benjamin M; Palotie, Aarno; Purcell, Shaun M; Saleheen, Danish; Scharf, Jeremiah M; Sklar, Pamela; Sullivan, Patrick F; Tuomilehto, Jaakko; Tsuang, Ming T; Watkins, Hugh C; Wilson, James G; Daly, Mark J; MacArthur, Daniel G

2016-08-18

Large-scale reference data sets of human genetic variation are critical for the medical and functional interpretation of DNA sequence changes. Here we describe the aggregation and analysis of high-quality exome (protein-coding region) DNA sequence data for 60,706 individuals of diverse ancestries generated as part of the Exome Aggregation Consortium (ExAC). This catalogue of human genetic diversity contains an average of one variant every eight bases of the exome, and provides direct evidence for the presence of widespread mutational recurrence. We have used this catalogue to calculate objective metrics of pathogenicity for sequence variants, and to identify genes subject to strong selection against various classes of mutation; identifying 3,230 genes with near-complete depletion of predicted protein-truncating variants, with 72% of these genes having no currently established human disease phenotype. Finally, we demonstrate that these data can be used for the efficient filtering of candidate disease-causing variants, and for the discovery of human 'knockout' variants in protein-coding genes.

Protein Profiles Reveal Diverse Responsive Signaling Pathways in Kernels of Two Maize Inbred Lines with Contrasting Drought Sensitivity

PubMed Central

Yang, Liming; Jiang, Tingbo; Fountain, Jake C.; Scully, Brian T.; Lee, Robert D.; Kemerait, Robert C.; Chen, Sixue; Guo, Baozhu

2014-01-01

Drought stress is a major factor that contributes to disease susceptibility and yield loss in agricultural crops. To identify drought responsive proteins and explore metabolic pathways involved in maize tolerance to drought stress, two maize lines (B73 and Lo964) with contrasting drought sensitivity were examined. The treatments of drought and well water were applied at 14 days after pollination (DAP), and protein profiles were investigated in developing kernels (35 DAP) using iTRAQ (isobaric tags for relative and absolute quantitation). Proteomic analysis showed that 70 and 36 proteins were significantly altered in their expression under drought treatments in B73 and Lo964, respectively. The numbers and levels of differentially expressed proteins were generally higher in the sensitive genotype, B73, implying an increased sensitivity to drought given the function of the observed differentially expressed proteins, such as redox homeostasis, cell rescue/defense, hormone regulation and protein biosynthesis and degradation. Lo964 possessed a more stable status with fewer differentially expressed proteins. However, B73 seems to rapidly initiate signaling pathways in response to drought through adjusting diverse defense pathways. These changes in protein expression allow for the production of a drought stress-responsive network in maize kernels. PMID:25334062

Highly branched penta-saccharide-bearing amphiphiles for membrane protein studies

PubMed Central

Ehsan, Muhammad; Du, Yang; Scull, Nicola J.; Tikhonova, Elena; Tarrasch, Jeffrey; Mortensen, Jonas S.; Loland, Claus J.; Skiniotis, Georgios; Guan, Lan; Byrne, Bernadette; Kobilka, Brian K.; Chae, Pil Seok

2016-01-01

Detergents are essential tools for membrane protein manipulation. Micelles formed by detergent molecules have the ability to encapsulate the hydrophobic domains of membrane proteins. The resulting protein-detergent complexes (PDCs) are compatible with the polar environments of aqueous media, making structural and functional analysis feasible. Although a number of novel agents have been developed to overcome the limitations of conventional detergents, most of them have traditional head groups such as glucoside or maltoside. In this study, we introduce a class of amphiphiles, the PSA’Es with a novel highly branched penta-saccharide hydrophilic group. The PSA’Es conferred markedly increased stability to a diverse range of membrane proteins compared to conventional detergents, indicating a positive role for the new hydrophilic group in maintaining the native protein integrity. In addition, PDCs formed by PSA’Es were smaller and more suitable for electron microscopic analysis than those formed by DDM, indicating that the new agents have significant potential for the structure-function studies of membrane proteins. PMID:26966956

Establishment of cell surface engineering and its development.

PubMed

Ueda, Mitsuyoshi

2016-07-01

Cell surface display of proteins/peptides has been established based on mechanisms of localizing proteins to the cell surface. In contrast to conventional intracellular and extracellular (secretion) expression systems, this method, generally called an arming technology, is particularly effective when using yeasts as a host, because the control of protein folding that is often required for the preparation of proteins can be natural. This technology can be employed for basic and applied research purposes. In this review, I describe various strategies for the construction of engineered yeasts and provide an outline of the diverse applications of this technology to industrial processes such as the production of biofuels and chemicals, as well as bioremediation and health-related processes. Furthermore, this technology is suitable for novel protein engineering and directed evolution through high-throughput screening, because proteins/peptides displayed on the cell surface can be directly analyzed using intact cells without concentration and purification. Functional proteins/peptides with improved or novel functions can be created using this beneficial, powerful, and promising technique.

Pannexins and gap junction protein diversity.

PubMed

Shestopalov, V I; Panchin, Y

2008-02-01

Gap junctions (GJs) are composed of proteins that form a channel connecting the cytoplasm of adjacent cells. Connexins were initially considered to be the only proteins capable of GJ formation. Another family of GJ proteins (innexins) were first found in invertebrates and were proposed to be renamed pannexins after their orthologs were discovered in vertebrates. The lack of both connexins and pannexins in the genomes of some metazoans suggests that other, still undiscovered GJ proteins exist. In vertebrates, connexins and pannexins co-exist. Here we discuss whether vertebrate pannexins have a nonredundant role in animal physiology. Pannexin channels appear to be suited for ATP and calcium signaling and play a role in the maintenance of calcium homeostasis by mechanisms implicating both GJ and nonjunctional function. Suggested roles in the ischemic death of neurons, schizophrenia, inflammation and tumor suppression have drawn much attention to exploring the molecular properties and cellular functions of pannexins.

Trafficking and Membrane Organization of GPI-Anchored Proteins in Health and Diseases.

PubMed

Paladino, Simona; Lebreton, Stéphanie; Zurzolo, Chiara

2015-01-01

Glycosylphosphatidylinositol (GPI)-anchored proteins (GPI-APs) are a class of lipid-anchored proteins attached to the membranes by a glycolipid anchor that is added, as posttranslation modification, in the endoplasmic reticulum. GPI-APs are expressed at the cell surface of eukaryotes where they play diverse vital functions. Like all plasma membrane proteins, GPI-APs must be correctly sorted along the different steps of the secretory pathway to their final destination. The presence of both a glycolipid anchor and a protein portion confers special trafficking features to GPI-APs. Here, we discuss the recent advances in the field of GPI-AP trafficking, focusing on the mechanisms regulating their biosynthetic pathway and plasma membrane organization. We also discuss how alterations of these mechanisms can result in different diseases. Finally, we will examine the strict relationship between the trafficking and function of GPI-APs in epithelial cells. Copyright © 2015 Elsevier Inc. All rights reserved.

Splicing factor SFRS1 recognizes a functionally diverse landscape of RNA transcripts.

PubMed

Sanford, Jeremy R; Wang, Xin; Mort, Matthew; Vanduyn, Natalia; Cooper, David N; Mooney, Sean D; Edenberg, Howard J; Liu, Yunlong

2009-03-01

Metazoan genes are encrypted with at least two superimposed codes: the genetic code to specify the primary structure of proteins and the splicing code to expand their proteomic output via alternative splicing. Here, we define the specificity of a central regulator of pre-mRNA splicing, the conserved, essential splicing factor SFRS1. Cross-linking immunoprecipitation and high-throughput sequencing (CLIP-seq) identified 23,632 binding sites for SFRS1 in the transcriptome of cultured human embryonic kidney cells. SFRS1 was found to engage many different classes of functionally distinct transcripts including mRNA, miRNA, snoRNAs, ncRNAs, and conserved intergenic transcripts of unknown function. The majority of these diverse transcripts share a purine-rich consensus motif corresponding to the canonical SFRS1 binding site. The consensus site was not only enriched in exons cross-linked to SFRS1 in vivo, but was also enriched in close proximity to splice sites. mRNAs encoding RNA processing factors were significantly overrepresented, suggesting that SFRS1 may broadly influence the post-transcriptional control of gene expression in vivo. Finally, a search for the SFRS1 consensus motif within the Human Gene Mutation Database identified 181 mutations in 82 different genes that disrupt predicted SFRS1 binding sites. This comprehensive analysis substantially expands the known roles of human SR proteins in the regulation of a diverse array of RNA transcripts.

Targeting Protein Quality Control Mechanisms by Natural Products to Promote Healthy Ageing.

PubMed

Wedel, Sophia; Manola, Maria; Cavinato, Maria; Trougakos, Ioannis P; Jansen-Dürr, Pidder

2018-05-19

Organismal ageing is associated with increased chance of morbidity or mortality and it is driven by diverse molecular pathways that are affected by both environmental and genetic factors. The progression of ageing correlates with the gradual accumulation of stressors and damaged biomolecules due to the time-dependent decline of stress resistance and functional capacity, which eventually compromise cellular homeodynamics. As protein machines carry out the majority of cellular functions, proteome quality control is critical for cellular functionality and is carried out through the curating activity of the proteostasis network (PN). Key components of the PN are the two main degradation machineries, namely the ubiquitin-proteasome and autophagy-lysosome pathways along with several stress-responsive pathways, such as that of nuclear factor erythroid 2-related factor 2 (Nrf2), which mobilises cytoprotective genomic responses against oxidative and/or xenobiotic damage. Reportedly, genetic or dietary interventions that activate components of the PN delay ageing in evolutionarily diverse organisms. Natural products (extracts or pure compounds) represent an extraordinary inventory of highly diverse structural scaffolds that offer promising activities towards meeting the challenge of increasing healthspan and/or delaying ageing (e.g., spermidine, quercetin or sulforaphane). Herein, we review those natural compounds that have been found to activate proteostatic and/or anti-stress cellular responses and hence have the potential to delay cellular senescence and/or in vivo ageing.

Diverse RNA-binding proteins interact with functionally related sets of RNAs, suggesting an extensive regulatory system.

PubMed

Hogan, Daniel J; Riordan, Daniel P; Gerber, André P; Herschlag, Daniel; Brown, Patrick O

2008-10-28

RNA-binding proteins (RBPs) have roles in the regulation of many post-transcriptional steps in gene expression, but relatively few RBPs have been systematically studied. We searched for the RNA targets of 40 proteins in the yeast Saccharomyces cerevisiae: a selective sample of the approximately 600 annotated and predicted RBPs, as well as several proteins not annotated as RBPs. At least 33 of these 40 proteins, including three of the four proteins that were not previously known or predicted to be RBPs, were reproducibly associated with specific sets of a few to several hundred RNAs. Remarkably, many of the RBPs we studied bound mRNAs whose protein products share identifiable functional or cytotopic features. We identified specific sequences or predicted structures significantly enriched in target mRNAs of 16 RBPs. These potential RNA-recognition elements were diverse in sequence, structure, and location: some were found predominantly in 3'-untranslated regions, others in 5'-untranslated regions, some in coding sequences, and many in two or more of these features. Although this study only examined a small fraction of the universe of yeast RBPs, 70% of the mRNA transcriptome had significant associations with at least one of these RBPs, and on average, each distinct yeast mRNA interacted with three of the RBPs, suggesting the potential for a rich, multidimensional network of regulation. These results strongly suggest that combinatorial binding of RBPs to specific recognition elements in mRNAs is a pervasive mechanism for multi-dimensional regulation of their post-transcriptional fate.

The TIM Barrel Architecture Facilitated the Early Evolution of Protein-Mediated Metabolism.

PubMed

Goldman, Aaron David; Beatty, Joshua T; Landweber, Laura F

2016-01-01

The triosephosphate isomerase (TIM) barrel protein fold is a structurally repetitive architecture that is present in approximately 10% of all enzymes. It is generally assumed that this ubiquity in modern proteomes reflects an essential historical role in early protein-mediated metabolism. Here, we provide quantitative and comparative analyses to support several hypotheses about the early importance of the TIM barrel architecture. An information theoretical analysis of protein structures supports the hypothesis that the TIM barrel architecture could arise more easily by duplication and recombination compared to other mixed α/β structures. We show that TIM barrel enzymes corresponding to the most taxonomically broad superfamilies also have the broadest range of functions, often aided by metal and nucleotide-derived cofactors that are thought to reflect an earlier stage of metabolic evolution. By comparison to other putatively ancient protein architectures, we find that the functional diversity of TIM barrel proteins cannot be explained simply by their antiquity. Instead, the breadth of TIM barrel functions can be explained, in part, by the incorporation of a broad range of cofactors, a trend that does not appear to be shared by proteins in general. These results support the hypothesis that the simple and functionally general TIM barrel architecture may have arisen early in the evolution of protein biosynthesis and provided an ideal scaffold to facilitate the metabolic transition from ribozymes, peptides, and geochemical catalysts to modern protein enzymes.

Enzymatically Active Microgels from Self-Assembling Protein Nanofibrils for Microflow Chemistry.

PubMed

Zhou, Xiao-Ming; Shimanovich, Ulyana; Herling, Therese W; Wu, Si; Dobson, Christopher M; Knowles, Tuomas P J; Perrett, Sarah

2015-06-23

Amyloid fibrils represent a generic class of protein structure associated with both pathological states and with naturally occurring functional materials. This class of protein nanostructure has recently also emerged as an excellent foundation for sophisticated functional biocompatible materials including scaffolds and carriers for biologically active molecules. Protein-based materials offer the potential advantage that additional functions can be directly incorporated via gene fusion producing a single chimeric polypeptide that will both self-assemble and display the desired activity. To succeed, a chimeric protein system must self-assemble without the need for harsh triggering conditions which would damage the appended functional protein molecule. However, the micrometer to nanoscale patterning and morphological control of protein-based nanomaterials has remained challenging. This study demonstrates a general approach for overcoming these limitations through the microfluidic generation of enzymatically active microgels that are stabilized by amyloid nanofibrils. The use of scaffolds formed from biomaterials that self-assemble under mild conditions enables the formation of catalytic microgels while maintaining the integrity of the encapsulated enzyme. The enzymatically active microgel particles show robust material properties and their porous architecture allows diffusion in and out of reactants and products. In combination with microfluidic droplet trapping approaches, enzymatically active microgels illustrate the potential of self-assembling materials for enzyme immobilization and recycling, and for biological flow-chemistry. These design principles can be adopted to create countless other bioactive amyloid-based materials with diverse functions.

Transmembrane Mucins: Signaling Receptors at the Intersection of Inflammation and Cancer

PubMed Central

van Putten, Jos P.M.; Strijbis, Karin

2017-01-01

Mucosal surfaces line our body cavities and provide the interaction surface between commensal and pathogenic microbiota and the host. The barrier function of the mucosal layer is largely maintained by gel-forming mucin proteins that are secreted by goblet cells. In addition, mucosal epithelial cells express cell-bound mucins that have both barrier and signaling functions. The family of transmembrane mucins consists of diverse members that share a few characteristics. The highly glycosylated extracellular mucin domains inhibit invasion by pathogenic bacteria and can form a tight mesh structure that protects cells in harmful conditions. The intracellular tails of transmembrane mucins can be phosphorylated and connect to signaling pathways that regulate inflammation, cell-cell interactions, differentiation, and apoptosis. Transmembrane mucins play important roles in preventing infection at mucosal surfaces, but are also renowned for their contributions to the development, progression, and metastasis of adenocarcinomas. In general, transmembrane mucins seem to have evolved to monitor and repair damaged epithelia, but these functions can be highjacked by cancer cells to yield a survival advantage. This review presents an overview of the current knowledge of the functions of transmembrane mucins in inflammatory processes and carcinogenesis in order to better understand the diverse functions of these multifunctional proteins. PMID:28052300

The Double-Edged Sword: Conserved Functions of Extracellular Hsp90 in Wound Healing and Cancer

PubMed Central

Hance, Michael W.; Nolan, Krystal D.; Isaacs, Jennifer S.

2014-01-01

Heat shock proteins (Hsps) represent a diverse group of chaperones that play a vital role in the protection of cells against numerous environmental stresses. Although our understanding of chaperone biology has deepened over the last decade, the “atypical” extracellular functions of Hsps have remained somewhat enigmatic and comparatively understudied. The heat shock protein 90 (Hsp90) chaperone is a prototypic model for an Hsp family member exhibiting a duality of intracellular and extracellular functions. Intracellular Hsp90 is best known as a master regulator of protein folding. Cancers are particularly adept at exploiting this function of Hsp90, providing the impetus for the robust clinical development of small molecule Hsp90 inhibitors. However, in addition to its maintenance of protein homeostasis, Hsp90 has also been identified as an extracellular protein. Although early reports ascribed immunoregulatory functions to extracellular Hsp90 (eHsp90), recent studies have illuminated expanded functions for eHsp90 in wound healing and cancer. While the intended physiological role of eHsp90 remains enigmatic, its evolutionarily conserved functions in wound healing are easily co-opted during malignancy, a pathology sharing many properties of wounded tissue. This review will highlight the emerging functions of eHsp90 and shed light on its seemingly dichotomous roles as a benevolent facilitator of wound healing and as a sinister effector of tumor progression. PMID:24805867

Diversity as Opportunity: Insights from 600 Million Years of AHR Evolution.

PubMed

Hahn, Mark E; Karchner, Sibel I; Merson, Rebeka R

2017-02-01

The aryl hydrocarbon receptor (AHR) was for many years of interest only to pharmacologists and toxicologists. However, this protein has fundamental roles in biology that are being revealed through studies in diverse animal species. The AHR is an ancient protein. AHR homologs exist in most major groups of modern bilaterian animals, including deuterostomes (chordates, hemichordates, echinoderms) and the two major clades of protostome invertebrates [ecdysozoans (e.g. arthropods and nematodes) and lophotrochozoans (e.g. molluscs and annelids)]. AHR homologs also have been identified in cnidarians such as the sea anemone Nematostella and in the genome of Trichoplax , a placozoan. Bilaterians, cnidarians, and placozoans form the clade Eumetazoa , whose last common ancestor lived approximately 600 million years ago (MYA). The presence of AHR homologs in modern representatives of all these groups indicates that the original eumetazoan animal possessed an AHR homolog. Studies in invertebrates and vertebrates reveal parallel functions of AHR in the development and function of sensory neural systems, suggesting that these may be ancestral roles. Vertebrate animals are characterized by the expansion and diversification of AHRs, via gene and genome duplications, from the ancestral protoAHR into at least five classes of AHR-like proteins: AHR, AHR1, AHR2, AHR3, and AHRR. The evolution of multiple AHRs in vertebrates coincided with the acquisition of high-affinity binding of halogenated and polynuclear aromatic hydrocarbons and the emergence of adaptive functions involving regulation of xenobiotic-metabolizing enzymes and roles in adaptive immunity. The existence of multiple AHRs may have facilitated subfunction partitioning and specialization of specific AHR types in some taxa. Additional research in diverse model and non-model species will continue to enrich our understanding of AHR and its pleiotropic roles in biology and toxicology.

Diversity as Opportunity: Insights from 600 Million Years of AHR Evolution

PubMed Central

Hahn, Mark E.; Karchner, Sibel I.; Merson, Rebeka R.

2017-01-01

The aryl hydrocarbon receptor (AHR) was for many years of interest only to pharmacologists and toxicologists. However, this protein has fundamental roles in biology that are being revealed through studies in diverse animal species. The AHR is an ancient protein. AHR homologs exist in most major groups of modern bilaterian animals, including deuterostomes (chordates, hemichordates, echinoderms) and the two major clades of protostome invertebrates [ecdysozoans (e.g. arthropods and nematodes) and lophotrochozoans (e.g. molluscs and annelids)]. AHR homologs also have been identified in cnidarians such as the sea anemone Nematostella and in the genome of Trichoplax, a placozoan. Bilaterians, cnidarians, and placozoans form the clade Eumetazoa, whose last common ancestor lived approximately 600 million years ago (MYA). The presence of AHR homologs in modern representatives of all these groups indicates that the original eumetazoan animal possessed an AHR homolog. Studies in invertebrates and vertebrates reveal parallel functions of AHR in the development and function of sensory neural systems, suggesting that these may be ancestral roles. Vertebrate animals are characterized by the expansion and diversification of AHRs, via gene and genome duplications, from the ancestral protoAHR into at least five classes of AHR-like proteins: AHR, AHR1, AHR2, AHR3, and AHRR. The evolution of multiple AHRs in vertebrates coincided with the acquisition of high-affinity binding of halogenated and polynuclear aromatic hydrocarbons and the emergence of adaptive functions involving regulation of xenobiotic-metabolizing enzymes and roles in adaptive immunity. The existence of multiple AHRs may have facilitated subfunction partitioning and specialization of specific AHR types in some taxa. Additional research in diverse model and non-model species will continue to enrich our understanding of AHR and its pleiotropic roles in biology and toxicology. PMID:28286876

Epstein-Barr Virus Latent Membrane Protein 1 Genetic Variability in Peripheral Blood B Cells and Oropharyngeal Fluids

PubMed Central

Renzette, Nicholas; Somasundaran, Mohan; Brewster, Frank; Coderre, James; Weiss, Eric R.; McManus, Margaret; Greenough, Thomas; Tabak, Barbara; Garber, Manuel; Kowalik, Timothy F.

2014-01-01

ABSTRACT We report the diversity of latent membrane protein 1 (LMP1) gene founder sequences and the level of Epstein-Barr virus (EBV) genome variability over time and across anatomic compartments by using virus genomes amplified directly from oropharyngeal wash specimens and peripheral blood B cells during acute infection and convalescence. The intrahost nucleotide variability of the founder virus was 0.02% across the region sequences, and diversity increased significantly over time in the oropharyngeal compartment (P = 0.004). The LMP1 region showing the greatest level of variability in both compartments, and over time, was concentrated within the functional carboxyl-terminal activating regions 2 and 3 (CTAR2 and CTAR3). Interestingly, a deletion in a proline-rich repeat region (amino acids 274 to 289) of EBV commonly reported in EBV sequenced from cancer specimens was not observed in acute infectious mononucleosis (AIM) patients. Taken together, these data highlight the diversity in circulating EBV genomes and its potential importance in disease pathogenesis and vaccine design. IMPORTANCE This study is among the first to leverage an improved high-throughput deep-sequencing methodology to investigate directly from patient samples the degree of diversity in Epstein-Barr virus (EBV) populations and the extent to which viral genome diversity develops over time in the infected host. Significant variability of circulating EBV latent membrane protein 1 (LMP1) gene sequences was observed between cellular and oral wash samples, and this variability increased over time in oral wash samples. The significance of EBV genetic diversity in transmission and disease pathogenesis are discussed. PMID:24429365

Epstein-Barr virus latent membrane protein 1 genetic variability in peripheral blood B cells and oropharyngeal fluids.

PubMed

Renzette, Nicholas; Somasundaran, Mohan; Brewster, Frank; Coderre, James; Weiss, Eric R; McManus, Margaret; Greenough, Thomas; Tabak, Barbara; Garber, Manuel; Kowalik, Timothy F; Luzuriaga, Katherine

2014-04-01

We report the diversity of latent membrane protein 1 (LMP1) gene founder sequences and the level of Epstein-Barr virus (EBV) genome variability over time and across anatomic compartments by using virus genomes amplified directly from oropharyngeal wash specimens and peripheral blood B cells during acute infection and convalescence. The intrahost nucleotide variability of the founder virus was 0.02% across the region sequences, and diversity increased significantly over time in the oropharyngeal compartment (P = 0.004). The LMP1 region showing the greatest level of variability in both compartments, and over time, was concentrated within the functional carboxyl-terminal activating regions 2 and 3 (CTAR2 and CTAR3). Interestingly, a deletion in a proline-rich repeat region (amino acids 274 to 289) of EBV commonly reported in EBV sequenced from cancer specimens was not observed in acute infectious mononucleosis (AIM) patients. Taken together, these data highlight the diversity in circulating EBV genomes and its potential importance in disease pathogenesis and vaccine design. This study is among the first to leverage an improved high-throughput deep-sequencing methodology to investigate directly from patient samples the degree of diversity in Epstein-Barr virus (EBV) populations and the extent to which viral genome diversity develops over time in the infected host. Significant variability of circulating EBV latent membrane protein 1 (LMP1) gene sequences was observed between cellular and oral wash samples, and this variability increased over time in oral wash samples. The significance of EBV genetic diversity in transmission and disease pathogenesis are discussed.

Global genetic diversity of the Plasmodium vivax transmission-blocking vaccine candidate Pvs48/45.

PubMed

Vallejo, Andres F; Martinez, Nora L; Tobon, Alejandra; Alger, Jackeline; Lacerda, Marcus V; Kajava, Andrey V; Arévalo-Herrera, Myriam; Herrera, Sócrates

2016-04-12

Plasmodium vivax 48/45 protein is expressed on the surface of gametocytes/gametes and plays a key role in gamete fusion during fertilization. This protein was recently expressed in Escherichia coli host as a recombinant product that was highly immunogenic in mice and monkeys and induced antibodies with high transmission-blocking activity, suggesting its potential as a P. vivax transmission-blocking vaccine candidate. To determine sequence polymorphism of natural parasite isolates and its potential influence on the protein structure, all pvs48/45 sequences reported in databases from around the world as well as those from low-transmission settings of Latin America were compared. Plasmodium vivax parasite isolates from malaria-endemic regions of Colombia, Brazil and Honduras (n = 60) were used to sequence the Pvs48/45 gene, and compared to those previously reported to GenBank and PlasmoDB (n = 222). Pvs48/45 gene haplotypes were analysed to determine the functional significance of genetic variation in protein structure and vaccine potential. Nine non-synonymous substitutions (E35K, Y196H, H211N, K250N, D335Y, E353Q, A376T, K390T, K418R) and three synonymous substitutions (I73, T149, C156) that define seven different haplotypes were found among the 282 isolates from nine countries when compared with the Sal I reference sequence. Nucleotide diversity (π) was 0.00173 for worldwide samples (range 0.00033-0.00216), resulting in relatively high diversity in Myanmar and Colombia, and low diversity in Mexico, Peru and South Korea. The two most frequent substitutions (E353Q: 41.9 %, K250N: 39.5 %) were predicted to be located in antigenic regions without affecting putative B cell epitopes or the tertiary protein structure. There is limited sequence polymorphism in pvs48/45 with noted geographical clustering among Asian and American isolates. The low genetic diversity of the protein does not influence the predicted antigenicity or protein structure and, therefore, supports its further development as transmission-blocking vaccine candidate.

A high throughput mutagenic analysis of yeast sumo structure and function

PubMed Central

Newman, Heather A.; Lu, Jian; Carson, Caryn; Boeke, Jef D.

2017-01-01

Sumoylation regulates a wide range of essential cellular functions through diverse mechanisms that remain to be fully understood. Using S. cerevisiae, a model organism with a single essential SUMO gene (SMT3), we developed a library of >250 mutant strains with single or multiple amino acid substitutions of surface or core residues in the Smt3 protein. By screening this library using plate-based assays, we have generated a comprehensive structure-function based map of Smt3, revealing essential amino acid residues and residues critical for function under a variety of genotoxic and proteotoxic stress conditions. Functionally important residues mapped to surfaces affecting Smt3 precursor processing and deconjugation from protein substrates, covalent conjugation to protein substrates, and non-covalent interactions with E3 ligases and downstream effector proteins containing SUMO-interacting motifs. Lysine residues potentially involved in formation of polymeric chains were also investigated, revealing critical roles for polymeric chains, but redundancy in specific chain linkages. Collectively, our findings provide important insights into the molecular basis of signaling through sumoylation. Moreover, the library of Smt3 mutants represents a valuable resource for further exploring the functions of sumoylation in cellular stress response and other SUMO-dependent pathways. PMID:28166236

Arabidopsis thaliana type I and II chaperonins.

PubMed

Hill, J E; Hemmingsen, S M

2001-07-01

An examination of the Arabidopsis thaliana genome sequence led to the identification of 29 predicted genes with the potential to encode members of the chaperonin family of chaperones (CPN60 and CCT), their associated cochaperonins, and the cytoplasmic chaperonin cofactor prefoldin. These comprise the first complete set of plant chaperonin protein sequences and indicate that the CPN family is more diverse than previously described. In addition to surprising sequence diversity within CPN subclasses, the genomic data also suggest the existence of previously undescribed family members, including a 10-kDa chloroplast cochaperonin. Consideration of the sequence data described in this review prompts questions about the complexities of plant CPN systems and the evolutionary relationships and functions of the component proteins, most of which have not been studied experimentally.

Putting a photon to biological work: lessons from novel photoactive yellow protein homologs

NASA Astrophysics Data System (ADS)

Hoff, Wouter; Hara, Miwa; Ren, Jie; Kumauchi, Masato; Xie, Aihua; Larsen, Delmar; Mix, Tyler; Haraguchi, Shojiro; Shingae, Takahito; Unno, Masashi

The photoactive yellow protein (PYP) is a photosensory protein from the bacterium Halorhodospira halophila that has been used extensively as a model system for functional protein dynamics and biological signaling. We have been studying PYP homologs from a diverse set of bacteria. The PYP from Salinibacter ruber (Srub PYP) revealed a novel ``spectral isotope effect'' when the protein is dissolved in D2O. We were able to assign this effect to H/D exchange of an active side COOH group that forms an ionic hydrogen bond to the deprotonated (negatively charged) light-absorbing chromophore in PYP. Srub PYP also allowed us to measure Raman Optical Activity (ROA) spectra under resonance and pre-resonance conditions. This work revealed that pre-resonance (ROA) spectra are informative for analyzing active site distortions in PYP. Finally, ultrafast pump-probe experiments on three different PYPs revealed that the primary I0 photoproduct observed in Hhal PYP is not populated in some other PYP homologs. This observation has implications for the mechanism of chromophore photoisomerization at the start of the functional photocycle of PYP. These results illustrate how studies of different members of a protein family can lead to novel insights into basic mechanisms underlying protein function

Here, there and everywhere: Resistin-like molecules in infection, inflammation, and metabolic disorders.

PubMed

Pine, Gabrielle M; Batugedara, Hashini M; Nair, Meera G

2018-06-01

The Resistin-Like Molecules (RELM) α, β, and γ and their namesake, resistin, share structural and sequence homology but exhibit significant diversity in expression and function within their mammalian host. RELM proteins are expressed in a wide range of diseases, such as: microbial infections (eg. bacterial and helminth), inflammatory diseases (eg. asthma, fibrosis) and metabolic disorders (eg. diabetes). While the expression pattern and molecular regulation of RELM proteins are well characterized, much controversy remains over their proposed functions, with evidence of host-protective and pathogenic roles. Moreover, the receptors for RELM proteins are unclear, although three receptors for resistin, decorin, adenylyl cyclase-associated protein 1 (CAP1), and Toll-like Receptor 4 (TLR4) have recently been proposed. In this review, we will first summarize the molecular regulation of the RELM gene family, including transcription regulation and tissue expression in humans and mouse disease models. Second, we will outline the function and receptor-mediated signaling associated with RELM proteins. Finally, we will discuss recent studies suggesting that, despite early misconceptions that these proteins are pathogenic, RELM proteins have a more nuanced and potentially beneficial role for the host in certain disease settings. Copyright © 2018 Elsevier Ltd. All rights reserved.

Evolution of the arginase fold and functional diversity

PubMed Central

Dowling, Daniel P.; Costanzo, Luigi Di; Gennadios, Heather A.; Christianson, David W.

2009-01-01

The large number of protein structures deposited in the Protein Data Bank allows for the identification of novel structural superfamilies based on conservation of fold in addition to conservation of amino acid sequence. Since sequence diverges more rapidly than fold in protein evolution, proteins with little or no significant sequence identity are occasionally observed to adopt similar folds, thereby reflecting unanticipated evolutionary relationships. Here, we review the unique α/β fold first observed in the manganese metalloenzyme rat liver arginase, consisting of a parallel 8 stranded β-sheet surrounded by several helices, and its evolutionary relationship with the zinc-requiring and/or iron-requiring histone deacetylases and acetylpolyamine amidohydrolases. Structural comparisons reveal key features of the core α/β fold that contribute to the divergent metal ion specificity and stoichiometry required for the chemical and biological functions of these enzymes. PMID:18360740

Diversity in the origins of proteostasis networks- a driver for protein function in evolution

PubMed Central

Powers, Evan T.; Balch, William E.

2013-01-01

Although a protein’s primary sequence largely determines its function, proteins can adopt different folding states in response to changes in the environment, some of which may be deleterious to the organism. All organisms, including Bacteria, Archaea and Eukarya, have evolved a protein homeostasis network, or proteostasis network, that consists of chaperones and folding factors, degradation components, signalling pathways and specialized compartmentalized modules that manage protein folding in response to environmental stimuli and variation. Surveying the origins of proteostasis networks reveals that they have co-evolved with the proteome to regulate the physiological state of the cell, reflecting the unique stresses that different cells or organisms experience, and that they have a key role in driving evolution by closely managing the link between the phenotype and the genotype. PMID:23463216

The Ser/Thr Protein Kinase Protein-Protein Interaction Map of M. tuberculosis.

PubMed

Wu, Fan-Lin; Liu, Yin; Jiang, He-Wei; Luan, Yi-Zhao; Zhang, Hai-Nan; He, Xiang; Xu, Zhao-Wei; Hou, Jing-Li; Ji, Li-Yun; Xie, Zhi; Czajkowsky, Daniel M; Yan, Wei; Deng, Jiao-Yu; Bi, Li-Jun; Zhang, Xian-En; Tao, Sheng-Ce

2017-08-01

Mycobacterium tuberculosis (Mtb) is the causative agent of tuberculosis, the leading cause of death among all infectious diseases. There are 11 eukaryotic-like serine/threonine protein kinases (STPKs) in Mtb, which are thought to play pivotal roles in cell growth, signal transduction and pathogenesis. However, their underlying mechanisms of action remain largely uncharacterized. In this study, using a Mtb proteome microarray, we have globally identified the binding proteins in Mtb for all of the STPKs, and constructed the first STPK protein interaction (KPI) map that includes 492 binding proteins and 1,027 interactions. Bioinformatics analysis showed that the interacting proteins reflect diverse functions, including roles in two-component system, transcription, protein degradation, and cell wall integrity. Functional investigations confirmed that PknG regulates cell wall integrity through key components of peptidoglycan (PG) biosynthesis, e.g. MurC. The global STPK-KPIs network constructed here is expected to serve as a rich resource for understanding the key signaling pathways in Mtb, thus facilitating drug development and effective control of Mtb. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

A structural-alphabet-based strategy for finding structural motifs across protein families

PubMed Central

Wu, Chih Yuan; Chen, Yao Chi; Lim, Carmay

2010-01-01

Proteins with insignificant sequence and overall structure similarity may still share locally conserved contiguous structural segments; i.e. structural/3D motifs. Most methods for finding 3D motifs require a known motif to search for other similar structures or functionally/structurally crucial residues. Here, without requiring a query motif or essential residues, a fully automated method for discovering 3D motifs of various sizes across protein families with different folds based on a 16-letter structural alphabet is presented. It was applied to structurally non-redundant proteins bound to DNA, RNA, obligate/non-obligate proteins as well as free DNA-binding proteins (DBPs) and proteins with known structures but unknown function. Its usefulness was illustrated by analyzing the 3D motifs found in DBPs. A non-specific motif was found with a ‘corner’ architecture that confers a stable scaffold and enables diverse interactions, making it suitable for binding not only DNA but also RNA and proteins. Furthermore, DNA-specific motifs present ‘only’ in DBPs were discovered. The motifs found can provide useful guidelines in detecting binding sites and computational protein redesign. PMID:20525797

'2TM proteins': an antigenically diverse superfamily with variable functions and export pathways.

PubMed

Kaur, Jasweer; Hora, Rachna

2018-01-01

Malaria is a disease that affects millions of people annually. An intracellular habitat and lack of protein synthesizing machinery in erythrocytes pose numerous difficulties for survival of the human pathogen Plasmodium falciparum . The parasite refurbishes the infected red blood cell (iRBC) by synthesis and export of several proteins in an attempt to suffice its metabolic needs and evade the host immune response. Immune evasion is largely mediated by surface display of highly polymorphic protein families known as variable surface antigens. These include the two trans-membrane (2TM) superfamily constituted by multicopy repetitive interspersed family (RIFINs), subtelomeric variable open reading frame (STEVORs) and Plasmodium falciparum Maurer's cleft two trans-membrane proteins present only in P. falciparum and some simian infecting Plasmodium species. Their hypervariable region flanked by 2TM domains exposed on the iRBC surface is believed to generate antigenic diversity. Though historically named "2TM superfamily," several A-type RIFINs and some STEVORs assume one trans-membrane topology. RIFINs and STEVORs share varied functions in different parasite life cycle stages like rosetting, alteration of iRBC rigidity and immune evasion. Additionally, a member of the STEVOR family has been implicated in merozoite invasion. Differential expression of these families in laboratory strains and clinical isolates propose them to be important for host cell survival and defense. The role of RIFINs in modulation of host immune response and presence of protective antibodies against these surface exposed molecules in patient sera highlights them as attractive targets of antimalarial therapies and vaccines. 2TM proteins are Plasmodium export elements positive, and several of these are exported to the infected erythrocyte surface after exiting through the classical secretory pathway within parasites. Cleaved and modified proteins are trafficked after packaging in vesicles to reach Maurer's clefts, while information regarding delivery to the iRBC surface is sparse. Expression and export timing of the RIFIN and Plasmodium falciparum erythrocyte membrane protein1 families correspond to each other. Here, we have compiled and comprehended detailed information regarding orthologues, domain architecture, surface topology, functions and trafficking of members of the "2TM superfamily." Considering the large repertoire of proteins included in the 2TM superfamily and recent advances defining their function in malaria biology, a surge in research carried out on this important protein superfamily is likely.

CRISPR-Cas: evolution of an RNA-based adaptive immunity system in prokaryotes.

PubMed

Koonin, Eugene V; Makarova, Kira S

2013-05-01

The CRISPR-Cas (clustered regularly interspaced short palindromic repeats, CRISPR-associated genes) is an adaptive immunity system in bacteria and archaea that functions via a distinct self-non-self recognition mechanism that is partially analogous to the mechanism of eukaryotic RNA interference (RNAi). The CRISPR-Cas system incorporates fragments of virus or plasmid DNA into the CRISPR repeat cassettes and employs the processed transcripts of these spacers as guide RNAs to cleave the cognate foreign DNA or RNA. The Cas proteins, however, are not homologous to the proteins involved in RNAi and comprise numerous, highly diverged families. The majority of the Cas proteins contain diverse variants of the RNA recognition motif (RRM), a widespread RNA-binding domain. Despite the fast evolution that is typical of the cas genes, the presence of diverse versions of the RRM in most Cas proteins provides for a simple scenario for the evolution of the three distinct types of CRISPR-cas systems. In addition to several proteins that are directly implicated in the immune response, the cas genes encode a variety of proteins that are homologous to prokaryotic toxins that typically possess nuclease activity. The predicted toxins associated with CRISPR-Cas systems include the essential Cas2 protein, proteins of COG1517 that, in addition to a ligand-binding domain and a helix-turn-helix domain, typically contain different nuclease domains and several other predicted nucleases. The tight association of the CRISPR-Cas immunity systems with predicted toxins that, upon activation, would induce dormancy or cell death suggests that adaptive immunity and dormancy/suicide response are functionally coupled. Such coupling could manifest in the persistence state being induced and potentially providing conditions for more effective action of the immune system or in cell death being triggered when immunity fails.

Distinct domains of the AVRPM3A2/F2 avirulence protein from wheat powdery mildew are involved in immune receptor recognition and putative effector function

USDA-ARS?s Scientific Manuscript database

Recognition of the AVRPM3A2/F2 avirulence protein from powdery mildew by the wheat PM3A/F immune receptor induces a hypersensitive response after coexpression in Nicotiana benthamiana. The molecular determinants of this interaction and how they shape natural AvrPm3a2/f2 allelic diversity is unknown....

"Phylogenetic and evolutionary analysis of functional divergence among Gamma glutamyl transpeptidase (GGT) subfamilies".

PubMed

Verma, Ved Vrat; Gupta, Rani; Goel, Manisha

2015-09-14

γ-glutamyltranspeptidase (GGT) is a bi-substrate enzyme conserved in all three domains of life. It catalyzes the cleavage and transfer of γ-glutamyl moiety of glutathione to either water (hydrolysis) or substrates like peptides (transpeptidation). GGTs exhibit great variability in their enzyme kinetics although the mechanism of catalysis is conserved. Recently, GGT has been shown to be a virulence factor in microbes like Helicobacter pylori and Bacillus anthracis. In mammalian cells also, GGT inhibition prior to chemotherapy has been shown to sensitize tumors to the therapy. Therefore, lately both bacterial and eukaryotic GGTs have emerged as potential drug targets, but the efforts directed towards finding suitable inhibitors have not yielded any significant results yet. We propose that delineating the residues responsible for the functional diversity associated with these proteins could help in design of species/clade specific inhibitors. In the present study, we have carried out phylogenetic analysis on a set of 47 GGT-like proteins to address the functional diversity. These proteins segregate into various subfamilies, forming separate clades on the tree. Sequence conservation and motif prediction studies show that even though most of the highly conserved residues have been characterized biochemically in previous studies, a significant number of novel putative sites and motifs are discovered that vary in a clade specific manner. Many of the putative sites predicted during the functional divergence type I and type II analysis, lie close to the known catalytic residues and line the walls of the substrate binding cavity, reinforcing their role in modulating the substrate specificity, catalytic rates and stability of this protein. The study offers interesting insights into the evolution of GGT-like proteins in pathogenic vs. non-pathogenic bacteria, archaea and eukaryotes. Our analysis delineates residues that are highly specific to each GGT subfamily. We propose that these sites not only explain the differences in stability and catalytic variability of various GGTs but can also aid in design of specific inhibitors against particular GGTs. Thus, apart from the commonly used in-silico inhibitor screening approaches, evolutionary analysis identifying the functional divergence hotspots in GGT proteins could augment the structure based drug design approaches.

1,003 reference genomes of bacterial and archaeal isolates expand coverage of the tree of life

DOE PAGES

Mukherjee, Supratim; Seshadri, Rekha; Varghese, Neha J.; ...

2017-06-12

We present 1,003 reference genomes that were sequenced as part of the Genomic Encyclopedia of Bacteria and Archaea (GEBA) initiative, selected to maximize sequence coverage of phylogenetic space. These genomes double the number of existing type strains and expand their overall phylogenetic diversity by 25%. Comparative analyses with previously available finished and draft genomes reveal a 10.5% increase in novel protein families as a function of phylogenetic diversity. The GEBA genomes recruit 25 million previously unassigned metagenomic proteins from 4,650 samples, improving their phylogenetic and functional interpretation. We identify numerous biosynthetic clusters and experimentally validate a divergent phenazine cluster withmore » potential new chemical structure and antimicrobial activity. This Resource is the largest single release of reference genomes to date. Bacterial and archaeal isolate sequence space is still far from saturated, and future endeavors in this direction will continue to be a valuable resource for scientific discovery.« less

Cracking the ANP32 whips: Important functions, unequal requirement, and hints at disease implications

PubMed Central

Reilly, Patrick T; Yu, Yun; Hamiche, Ali; Wang, Lishun

2014-01-01

The acidic (leucine-rich) nuclear phosphoprotein 32 kDa (ANP32) family is composed of small, evolutionarily conserved proteins characterized by an N-terminal leucine-rich repeat domain and a C-terminal low-complexity acidic region. The mammalian family members (ANP32A, ANP32B, and ANP32E) are ascribed physiologically diverse functions including chromatin modification and remodelling, apoptotic caspase modulation, protein phosphatase inhibition, as well as regulation of intracellular transport. In addition to reviewing the widespread literature on the topic, we present a concept of the ANP32s as having a whip-like structure. We also present hypotheses that ANP32C and other intronless sequences should not currently be considered bona fide family members, that their disparate necessity in development may be due to compensatory mechanisms, that their contrasting roles in cancer are likely context-dependent, along with an underlying hypothesis that ANP32s represent an important node of physiological regulation by virtue of their diverse biochemical activities. PMID:25156960

Mechanical design of translocating motor proteins.

PubMed

Hwang, Wonmuk; Lang, Matthew J

2009-01-01

Translocating motors generate force and move along a biofilament track to achieve diverse functions including gene transcription, translation, intracellular cargo transport, protein degradation, and muscle contraction. Advances in single molecule manipulation experiments, structural biology, and computational analysis are making it possible to consider common mechanical design principles of these diverse families of motors. Here, we propose a mechanical parts list that include track, energy conversion machinery, and moving parts. Energy is supplied not just by burning of a fuel molecule, but there are other sources or sinks of free energy, by binding and release of a fuel or products, or similarly between the motor and the track. Dynamic conformational changes of the motor domain can be regarded as controlling the flow of free energy to and from the surrounding heat reservoir. Multiple motor domains are organized in distinct ways to achieve motility under imposed physical constraints. Transcending amino acid sequence and structure, physically and functionally similar mechanical parts may have evolved as nature's design strategy for these molecular engines.

Mechanical Design of Translocating Motor Proteins

PubMed Central

Lang, Matthew J.

2013-01-01

Translocating motors generate force and move along a biofilament track to achieve diverse functions including gene transcription, translation, intracellular cargo transport, protein degradation, and muscle contraction. Advances in single molecule manipulation experiments, structural biology, and computational analysis are making it possible to consider common mechanical design principles of these diverse families of motors. Here, we propose a mechanical parts list that include track, energy conversion machinery, and moving parts. Energy is supplied not just by burning of a fuel molecule, but there are other sources or sinks of free energy, by binding and release of a fuel or products, or similarly between the motor and the track. Dynamic conformational changes of the motor domain can be regarded as controlling the flow of free energy to and from the surrounding heat reservoir. Multiple motor domains are organized in distinct ways to achieve motility under imposed physical constraints. Transcending amino acid sequence and structure, physically and functionally similar mechanical parts may have evolved as nature’s design strategy for these molecular engines. PMID:19452133

1,003 reference genomes of bacterial and archaeal isolates expand coverage of the tree of life

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mukherjee, Supratim; Seshadri, Rekha; Varghese, Neha J.

We present 1,003 reference genomes that were sequenced as part of the Genomic Encyclopedia of Bacteria and Archaea (GEBA) initiative, selected to maximize sequence coverage of phylogenetic space. These genomes double the number of existing type strains and expand their overall phylogenetic diversity by 25%. Comparative analyses with previously available finished and draft genomes reveal a 10.5% increase in novel protein families as a function of phylogenetic diversity. The GEBA genomes recruit 25 million previously unassigned metagenomic proteins from 4,650 samples, improving their phylogenetic and functional interpretation. We identify numerous biosynthetic clusters and experimentally validate a divergent phenazine cluster withmore » potential new chemical structure and antimicrobial activity. This Resource is the largest single release of reference genomes to date. Bacterial and archaeal isolate sequence space is still far from saturated, and future endeavors in this direction will continue to be a valuable resource for scientific discovery.« less

Achieving high confidence protein annotations in a sea of unknowns

NASA Astrophysics Data System (ADS)

Timmins-Schiffman, E.; May, D. H.; Noble, W. S.; Nunn, B. L.; Mikan, M.; Harvey, H. R.

2016-02-01

Increased sensitivity of mass spectrometry (MS) technology allows deep and broad insight into community functional analyses. Metaproteomics holds the promise to reveal functional responses of natural microbial communities, whereas metagenomics alone can only hint at potential functions. The complex datasets resulting from ocean MS have the potential to inform diverse realms of the biological, chemical, and physical ocean sciences, yet the extent of bacterial functional diversity and redundancy has not been fully explored. To take advantage of these impressive datasets, we need a clear bioinformatics pipeline for metaproteomics peptide identification and annotation with a database that can provide confident identifications. Researchers must consider whether it is sufficient to leverage the vast quantities of available ocean sequence data or if they must invest in site-specific metagenomic sequencing. We have sequenced, to our knowledge, the first western arctic metagenomes from the Bering Strait and the Chukchi Sea. We have addressed the long standing question: Is a metagenome required to accurately complete metaproteomics and assess the biological distribution of metabolic functions controlling nutrient acquisition in the ocean? Two different protein databases were constructed from 1) a site-specific metagenome and 2) subarctic/arctic groups available in NCBI's non-redundant database. Multiple proteomic search strategies were employed, against each individual database and against both databases combined, to determine the algorithm and approach that yielded the balance of high sensitivity and confident identification. Results yielded over 8200 confidently identified proteins. Our comparison of these results allows us to quantify the utility of investing resources in a metagenome versus using the constantly expanding and immediately available public databases for metaproteomic studies.

Differential regulation of cellular functions by the C-termini of transmembrane 4 L six family proteins in 2- or 3-dimensional environment.

PubMed

Cheong, Jin-Gyu; Song, Dae-Geun; Song, Haeng Eun; Berditchevski, Fedor; Nam, Seo Hee; Jung, Jae Woo; Kim, Hye-Jin; Kim, Ji Eon; Kim, Somi; Ryu, Jihye; Cho, Chang Yun; Lee, Kyung-Min; Lee, Jung Weon

2017-02-21

The transmembrane 4 L six family proteins TM4SF1, TM4SF4, and TM4SF5 share 40-50% overall sequence identity, but their C-terminus identity is limited. It may be likely that the C-termini of the members are important and unique for own regulatory functions. We thus examined how the TM4SF5 C-terminus affected cellular functions differentially from other family members. Using colon cancer cells expressing wildtype (WT), C-terminus-deleted, or chimeric mutants, diverse cellular functions were explored in 2-dimensional (2D) and 3-dimensional (3D) condition. The C-termini of the proteins were relatively comparable with respect to 2D cell proliferation, although each C-terminal-deletion mutant exhibited increased proliferation relative to the WT. Using chimeric constructs, we found that the TM4SF5 C-terminus was critical for regulating the diverse metastatic functions of TM4SF5, and could positively replace the C-termini of other family members. Replacement of the TM4SF1 or TM4SF4 C-terminus with that of TM4SF5 increased spheroids growth, transwell migration, and invasive dissemination from spheroids in 3D collagen gels. TM4SF5-mediated effects required its extracellular loop 2 linked to the C-terminus via the transmembrane domain 4, with causing c-Src activation. Altogether, the C-terminus of TM4SF5 appears to mediate pro-migratory roles, depending on a structural relay from the second extracellular loop to the C-terminus.

Evolution of the rodent eosinophil-associated RNase gene family by rapid gene sorting and positive selection

PubMed Central

Zhang, Jianzhi; Dyer, Kimberly D.; Rosenberg, Helene F.

2000-01-01

The mammalian RNase A superfamily comprises a diverse array of ribonucleolytic proteins that have a variety of biochemical activities and physiological functions. Two rapidly evolving RNases of higher primates are of particular interest as they are major secretory proteins of eosinophilic leukocytes and have been found to possess anti-pathogen activities in vitro. To understand how these RNases acquired this function during evolution and to develop animal models for the study of their functions in vivo, it is necessary to investigate these genes in many species. Here, we report the sequences of 38 functional genes and 23 pseudogenes of the eosinophil-associated RNase (EAR) family from 5 rodent species. Our phylogenetic analysis of these genes showed a clear pattern of evolution by a rapid birth-and-death process and gene sorting, a process characterized by rapid gene duplication and deactivation occurring differentially among lineages. This process ultimately generates distinct or only partially overlapping inventories of the genes, even in closely related species. Positive Darwinian selection also contributed to the diversification of these EAR genes. The striking similarity between the evolutionary patterns of the EAR genes and those of the major histocompatibility complex, immunoglobulin, and T cell receptor genes stands in strong support of the hypothesis that host-defense and generation of diversity are among the primary physiological function of the rodent EARs. The discovery of a large number of divergent EARs suggests the intriguing possibility that these proteins have been specifically tailored to fight against distinct rodent pathogens. PMID:10758160

Amyloid Fibrils as Building Blocks for Natural and Artificial Functional Materials.

PubMed

Knowles, Tuomas P J; Mezzenga, Raffaele

2016-08-01

Proteinaceous materials based on the amyloid core structure have recently been discovered at the origin of biological functionality in a remarkably diverse set of roles, and attention is increasingly turning towards such structures as the basis of artificial self-assembling materials. These roles contrast markedly with the original picture of amyloid fibrils as inherently pathological structures. Here we outline the salient features of this class of functional materials, both in the context of the functional roles that have been revealed for amyloid fibrils in nature, as well as in relation to their potential as artificial materials. We discuss how amyloid materials exemplify the emergence of function from protein self-assembly at multiple length scales. We focus on the connections between mesoscale structure and material function, and demonstrate how the natural examples of functional amyloids illuminate the potential applications for future artificial protein based materials. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Annotating Protein Functional Residues by Coupling High-Throughput Fitness Profile and Homologous-Structure Analysis.

PubMed

Du, Yushen; Wu, Nicholas C; Jiang, Lin; Zhang, Tianhao; Gong, Danyang; Shu, Sara; Wu, Ting-Ting; Sun, Ren

2016-11-01

Identification and annotation of functional residues are fundamental questions in protein sequence analysis. Sequence and structure conservation provides valuable information to tackle these questions. It is, however, limited by the incomplete sampling of sequence space in natural evolution. Moreover, proteins often have multiple functions, with overlapping sequences that present challenges to accurate annotation of the exact functions of individual residues by conservation-based methods. Using the influenza A virus PB1 protein as an example, we developed a method to systematically identify and annotate functional residues. We used saturation mutagenesis and high-throughput sequencing to measure the replication capacity of single nucleotide mutations across the entire PB1 protein. After predicting protein stability upon mutations, we identified functional PB1 residues that are essential for viral replication. To further annotate the functional residues important to the canonical or noncanonical functions of viral RNA-dependent RNA polymerase (vRdRp), we performed a homologous-structure analysis with 16 different vRdRp structures. We achieved high sensitivity in annotating the known canonical polymerase functional residues. Moreover, we identified a cluster of noncanonical functional residues located in the loop region of the PB1 β-ribbon. We further demonstrated that these residues were important for PB1 protein nuclear import through the interaction with Ran-binding protein 5. In summary, we developed a systematic and sensitive method to identify and annotate functional residues that are not restrained by sequence conservation. Importantly, this method is generally applicable to other proteins about which homologous-structure information is available. To fully comprehend the diverse functions of a protein, it is essential to understand the functionality of individual residues. Current methods are highly dependent on evolutionary sequence conservation, which is usually limited by sampling size. Sequence conservation-based methods are further confounded by structural constraints and multifunctionality of proteins. Here we present a method that can systematically identify and annotate functional residues of a given protein. We used a high-throughput functional profiling platform to identify essential residues. Coupling it with homologous-structure comparison, we were able to annotate multiple functions of proteins. We demonstrated the method with the PB1 protein of influenza A virus and identified novel functional residues in addition to its canonical function as an RNA-dependent RNA polymerase. Not limited to virology, this method is generally applicable to other proteins that can be functionally selected and about which homologous-structure information is available. Copyright © 2016 Du et al.

Site-directed protein recombination as a shortest-path problem.

PubMed

Endelman, Jeffrey B; Silberg, Jonathan J; Wang, Zhen-Gang; Arnold, Frances H

2004-07-01

Protein function can be tuned using laboratory evolution, in which one rapidly searches through a library of proteins for the properties of interest. In site-directed recombination, n crossovers are chosen in an alignment of p parents to define a set of p(n + 1) peptide fragments. These fragments are then assembled combinatorially to create a library of p(n+1) proteins. We have developed a computational algorithm to enrich these libraries in folded proteins while maintaining an appropriate level of diversity for evolution. For a given set of parents, our algorithm selects crossovers that minimize the average energy of the library, subject to constraints on the length of each fragment. This problem is equivalent to finding the shortest path between nodes in a network, for which the global minimum can be found efficiently. Our algorithm has a running time of O(N(3)p(2) + N(2)n) for a protein of length N. Adjusting the constraints on fragment length generates a set of optimized libraries with varying degrees of diversity. By comparing these optima for different sets of parents, we rapidly determine which parents yield the lowest energy libraries.

Plant Cation-Chloride Cotransporters (CCC): Evolutionary Origins and Functional Insights

PubMed Central

Gilliham, Matthew

2018-01-01

Genomes of unicellular and multicellular green algae, mosses, grasses and dicots harbor genes encoding cation-chloride cotransporters (CCC). CCC proteins from the plant kingdom have been comparatively less well investigated than their animal counterparts, but proteins from both plants and animals have been shown to mediate ion fluxes, and are involved in regulation of osmotic processes. In this review, we show that CCC proteins from plants form two distinct phylogenetic clades (CCC1 and CCC2). Some lycophytes and bryophytes possess members from each clade, most land plants only have members of the CCC1 clade, and green algae possess only the CCC2 clade. It is currently unknown whether CCC1 and CCC2 proteins have similar or distinct functions, however they are both more closely related to animal KCC proteins compared to NKCCs. Existing heterologous expression systems that have been used to functionally characterize plant CCC proteins, namely yeast and Xenopus laevis oocytes, have limitations that are discussed. Studies from plants exposed to chemical inhibitors of animal CCC protein function are reviewed for their potential to discern CCC function in planta. Thus far, mutations in plant CCC genes have been evaluated only in two species of angiosperms, and such mutations cause a diverse array of phenotypes—seemingly more than could simply be explained by localized disruption of ion transport alone. We evaluate the putative roles of plant CCC proteins and suggest areas for future investigation. PMID:29415511

Members of the YjgF/YER057c/UK114 family of proteins inhibit phosphoribosylamine synthesis in vitro.

PubMed

Lambrecht, Jennifer A; Browne, Beth Ann; Downs, Diana M

2010-11-05

The YjgF/YER057c/UK114 family of proteins is highly conserved across all three domains of life and currently lacks a consensus biochemical function. Analysis of Salmonella enterica strains lacking yjgF has led to a working model in which YjgF functions to remove potentially toxic secondary products of cellular enzymes. Strains lacking yjgF synthesize the thiamine precursor phosphoribosylamine (PRA) by a TrpD-dependent mechanism that is not present in wild-type strains. Here, PRA synthesis was reconstituted in vitro with anthranilate phosphoribosyltransferase (TrpD), threonine dehydratase (IlvA), threonine, and phosphoribosyl pyrophosphate. TrpD-dependent PRA formation in vitro was inhibited by S. enterica YjgF and the human homolog UK114. Thus, the work herein describes the first biochemical assay for diverse members of the highly conserved YjgF/YER057c/UK114 family of proteins and provides a means to dissect the cellular functions of these proteins.

Probing the mutational interplay between primary and promiscuous protein functions: a computational-experimental approach.

PubMed

Garcia-Seisdedos, Hector; Ibarra-Molero, Beatriz; Sanchez-Ruiz, Jose M

2012-01-01

Protein promiscuity is of considerable interest due its role in adaptive metabolic plasticity, its fundamental connection with molecular evolution and also because of its biotechnological applications. Current views on the relation between primary and promiscuous protein activities stem largely from laboratory evolution experiments aimed at increasing promiscuous activity levels. Here, on the other hand, we attempt to assess the main features of the simultaneous modulation of the primary and promiscuous functions during the course of natural evolution. The computational/experimental approach we propose for this task involves the following steps: a function-targeted, statistical coupling analysis of evolutionary data is used to determine a set of positions likely linked to the recruitment of a promiscuous activity for a new function; a combinatorial library of mutations on this set of positions is prepared and screened for both, the primary and the promiscuous activities; a partial-least-squares reconstruction of the full combinatorial space is carried out; finally, an approximation to the Pareto set of variants with optimal primary/promiscuous activities is derived. Application of the approach to the emergence of folding catalysis in thioredoxin scaffolds reveals an unanticipated scenario: diverse patterns of primary/promiscuous activity modulation are possible, including a moderate (but likely significant in a biological context) simultaneous enhancement of both activities. We show that this scenario can be most simply explained on the basis of the conformational diversity hypothesis, although alternative interpretations cannot be ruled out. Overall, the results reported may help clarify the mechanisms of the evolution of new functions. From a different viewpoint, the partial-least-squares-reconstruction/Pareto-set-prediction approach we have introduced provides the computational basis for an efficient directed-evolution protocol aimed at the simultaneous enhancement of several protein features and should therefore open new possibilities in the engineering of multi-functional enzymes.

Probing the Mutational Interplay between Primary and Promiscuous Protein Functions: A Computational-Experimental Approach

PubMed Central

Garcia-Seisdedos, Hector; Ibarra-Molero, Beatriz; Sanchez-Ruiz, Jose M.

2012-01-01

Protein promiscuity is of considerable interest due its role in adaptive metabolic plasticity, its fundamental connection with molecular evolution and also because of its biotechnological applications. Current views on the relation between primary and promiscuous protein activities stem largely from laboratory evolution experiments aimed at increasing promiscuous activity levels. Here, on the other hand, we attempt to assess the main features of the simultaneous modulation of the primary and promiscuous functions during the course of natural evolution. The computational/experimental approach we propose for this task involves the following steps: a function-targeted, statistical coupling analysis of evolutionary data is used to determine a set of positions likely linked to the recruitment of a promiscuous activity for a new function; a combinatorial library of mutations on this set of positions is prepared and screened for both, the primary and the promiscuous activities; a partial-least-squares reconstruction of the full combinatorial space is carried out; finally, an approximation to the Pareto set of variants with optimal primary/promiscuous activities is derived. Application of the approach to the emergence of folding catalysis in thioredoxin scaffolds reveals an unanticipated scenario: diverse patterns of primary/promiscuous activity modulation are possible, including a moderate (but likely significant in a biological context) simultaneous enhancement of both activities. We show that this scenario can be most simply explained on the basis of the conformational diversity hypothesis, although alternative interpretations cannot be ruled out. Overall, the results reported may help clarify the mechanisms of the evolution of new functions. From a different viewpoint, the partial-least-squares-reconstruction/Pareto-set-prediction approach we have introduced provides the computational basis for an efficient directed-evolution protocol aimed at the simultaneous enhancement of several protein features and should therefore open new possibilities in the engineering of multi-functional enzymes. PMID:22719242

Protein homology model refinement by large-scale energy optimization.

PubMed

Park, Hahnbeom; Ovchinnikov, Sergey; Kim, David E; DiMaio, Frank; Baker, David

2018-03-20

Proteins fold to their lowest free-energy structures, and hence the most straightforward way to increase the accuracy of a partially incorrect protein structure model is to search for the lowest-energy nearby structure. This direct approach has met with little success for two reasons: first, energy function inaccuracies can lead to false energy minima, resulting in model degradation rather than improvement; and second, even with an accurate energy function, the search problem is formidable because the energy only drops considerably in the immediate vicinity of the global minimum, and there are a very large number of degrees of freedom. Here we describe a large-scale energy optimization-based refinement method that incorporates advances in both search and energy function accuracy that can substantially improve the accuracy of low-resolution homology models. The method refined low-resolution homology models into correct folds for 50 of 84 diverse protein families and generated improved models in recent blind structure prediction experiments. Analyses of the basis for these improvements reveal contributions from both the improvements in conformational sampling techniques and the energy function.

Metabolic functions of FABPs— mechanisms and therapeutic implications

PubMed Central

Hotamisligil, Gökhan S.; Bernlohr, David A.

2015-01-01

Intracellular and extracellular interactions with proteins enables the functional and mechanistic diversity of lipids. Fatty acid-binding proteins (FABPs) were originally described as intracellular proteins that can affect lipid fluxes, metabolism and signalling within cells. As the functions of this protein family have been further elucidated, it has become evident that they are critical mediators of metabolism and inflammatory processes, both locally and systemically, and therefore are potential therapeutic targets for immunometabolic diseases. In particular, genetic deficiency and small molecule-mediated inhibition of FABP4 (also known as aP2) and FABP5 can potently improve glucose homeostasis and reduce atherosclerosis in mouse models. Further research has shown that in addition to their intracellular roles, some FABPs are found outside the cells, and FABP4 undergoes regulated, vesicular secretion. The circulating form of FABP4 has crucial hormonal functions in systemic metabolism. In this Review we discuss the roles and regulation of both intracellular and extracellular FABP actions, highlighting new insights that might direct drug discovery efforts and opportunities for management of chronic metabolic diseases. PMID:26260145

Evolution and functional divergence of the anoctamin family of membrane proteins

PubMed Central

2010-01-01

Background The anoctamin family of transmembrane proteins are found in all eukaryotes and consists of 10 members in vertebrates. Ano1 and ano2 were observed to have Ca2+ activated Cl- channel activity. Recent findings however have revealed that ano6, and ano7 can also produce chloride currents, although with different properties. In contrast, ano9 and ano10 suppress baseline Cl- conductance when co-expressed with ano1 thus suggesting that different anoctamins can interfere with each other. In order to elucidate intrinsic functional diversity, and underlying evolutionary mechanism among anoctamins, we performed comprehensive bioinformatics analysis of anoctamin gene family. Results Our results show that anoctamin protein paralogs evolved from several gene duplication events followed by functional divergence of vertebrate anoctamins. Most of the amino acid replacements responsible for the functional divergence were fixed by adaptive evolution and this seem to be a common pattern in anoctamin gene family evolution. Strong purifying selection and the loss of many gene duplication products indicate rigid structure-function relationships among anoctamins. Conclusions Our study suggests that anoctamins have evolved by series of duplication events, and that they are constrained by purifying selection. In addition we identified a number of protein domains, and amino acid residues which contribute to predicted functional divergence. Hopefully, this work will facilitate future functional characterization of the anoctamin membrane protein family. PMID:20964844

MitProNet: A Knowledgebase and Analysis Platform of Proteome, Interactome and Diseases for Mammalian Mitochondria

PubMed Central

Mao, Song; Chai, Xiaoqiang; Hu, Yuling; Hou, Xugang; Tang, Yiheng; Bi, Cheng; Li, Xiao

2014-01-01

Mitochondrion plays a central role in diverse biological processes in most eukaryotes, and its dysfunctions are critically involved in a large number of diseases and the aging process. A systematic identification of mitochondrial proteomes and characterization of functional linkages among mitochondrial proteins are fundamental in understanding the mechanisms underlying biological functions and human diseases associated with mitochondria. Here we present a database MitProNet which provides a comprehensive knowledgebase for mitochondrial proteome, interactome and human diseases. First an inventory of mammalian mitochondrial proteins was compiled by widely collecting proteomic datasets, and the proteins were classified by machine learning to achieve a high-confidence list of mitochondrial proteins. The current version of MitProNet covers 1124 high-confidence proteins, and the remainders were further classified as middle- or low-confidence. An organelle-specific network of functional linkages among mitochondrial proteins was then generated by integrating genomic features encoded by a wide range of datasets including genomic context, gene expression profiles, protein-protein interactions, functional similarity and metabolic pathways. The functional-linkage network should be a valuable resource for the study of biological functions of mitochondrial proteins and human mitochondrial diseases. Furthermore, we utilized the network to predict candidate genes for mitochondrial diseases using prioritization algorithms. All proteins, functional linkages and disease candidate genes in MitProNet were annotated according to the information collected from their original sources including GO, GEO, OMIM, KEGG, MIPS, HPRD and so on. MitProNet features a user-friendly graphic visualization interface to present functional analysis of linkage networks. As an up-to-date database and analysis platform, MitProNet should be particularly helpful in comprehensive studies of complicated biological mechanisms underlying mitochondrial functions and human mitochondrial diseases. MitProNet is freely accessible at http://bio.scu.edu.cn:8085/MitProNet. PMID:25347823

Exploring the Arabidopsis proteome: influence of protein solubilization buffers on proteome coverage.

PubMed

Marondedze, Claudius; Wong, Aloysius; Groen, Arnoud; Serrano, Natalia; Jankovic, Boris; Lilley, Kathryn; Gehring, Christoph; Thomas, Ludivine

2014-12-31

The study of proteomes provides new insights into stimulus-specific responses of protein synthesis and turnover, and the role of post-translational modifications at the systems level. Due to the diverse chemical nature of proteins and shortcomings in the analytical techniques used in their study, only a partial display of the proteome is achieved in any study, and this holds particularly true for plant proteomes. Here we show that different solubilization and separation methods have profound effects on the resulting proteome. In particular, we observed that the type of detergents employed in the solubilization buffer preferentially enriches proteins in different functional categories. These include proteins with a role in signaling, transport, response to temperature stimuli and metabolism. This data may offer a functional bias on comparative analysis studies. In order to obtain a broader coverage, we propose a two-step solubilization protocol with first a detergent-free buffer and then a second step utilizing a combination of two detergents to solubilize proteins.

On the nature of a glassy state of matter in a hydrated protein: Relation to protein function.

PubMed

Teeter, M M; Yamano, A; Stec, B; Mohanty, U

2001-09-25

Diverse biochemical and biophysical experiments indicate that all proteins, regardless of size or origin, undergo a dynamic transition near 200 K. The cause of this shift in dynamic behavior, termed a "glass transition," and its relation to protein function are important open questions. One explanation postulated for the transition is solidification of correlated motions in proteins below the transition. We verified this conjecture by showing that crambin's radius of gyration (Rg) remains constant below approximately 180 K. We show that both atom position and dynamics of protein and solvent are physically coupled, leading to a novel cooperative state. This glassy state is identified by negative slopes of the Debye-Waller (B) factor vs. temperature. It is composed of multisubstate side chains and solvent. Based on generalization of Adam-Gibbs' notion of a cooperatively rearranging region and decrease of the total entropy with temperature, we calculate the slope of the Debye-Waller factor. The results are in accord with experiment.

A Global Map of Lipid-Binding Proteins and Their Ligandability in Cells.

PubMed

Niphakis, Micah J; Lum, Kenneth M; Cognetta, Armand B; Correia, Bruno E; Ichu, Taka-Aki; Olucha, Jose; Brown, Steven J; Kundu, Soumajit; Piscitelli, Fabiana; Rosen, Hugh; Cravatt, Benjamin F

2015-06-18

Lipids play central roles in physiology and disease, where their structural, metabolic, and signaling functions often arise from interactions with proteins. Here, we describe a set of lipid-based chemical proteomic probes and their global interaction map in mammalian cells. These interactions involve hundreds of proteins from diverse functional classes and frequently occur at sites of drug action. We determine the target profiles for several drugs across the lipid-interaction proteome, revealing that its ligandable content extends far beyond traditionally defined categories of druggable proteins. In further support of this finding, we describe a selective ligand for the lipid-binding protein nucleobindin-1 (NUCB1) and show that this compound perturbs the hydrolytic and oxidative metabolism of endocannabinoids in cells. The described chemical proteomic platform thus provides an integrated path to both discover and pharmacologically characterize a wide range of proteins that participate in lipid pathways in cells. Copyright © 2015 Elsevier Inc. All rights reserved.

Exploring the Arabidopsis Proteome: Influence of Protein Solubilization Buffers on Proteome Coverage

PubMed Central

Marondedze, Claudius; Wong, Aloysius; Groen, Arnoud; Serrano, Natalia; Jankovic, Boris; Lilley, Kathryn; Gehring, Christoph; Thomas, Ludivine

2014-01-01

The study of proteomes provides new insights into stimulus-specific responses of protein synthesis and turnover, and the role of post-translational modifications at the systems level. Due to the diverse chemical nature of proteins and shortcomings in the analytical techniques used in their study, only a partial display of the proteome is achieved in any study, and this holds particularly true for plant proteomes. Here we show that different solubilization and separation methods have profound effects on the resulting proteome. In particular, we observed that the type of detergents employed in the solubilization buffer preferentially enriches proteins in different functional categories. These include proteins with a role in signaling, transport, response to temperature stimuli and metabolism. This data may offer a functional bias on comparative analysis studies. In order to obtain a broader coverage, we propose a two-step solubilization protocol with first a detergent-free buffer and then a second step utilizing a combination of two detergents to solubilize proteins. PMID:25561235

Transduction of Redox Signaling by Electrophile-Protein Reactions

PubMed Central

Rudolph, Tanja K.; Freeman, Bruce A.

2014-01-01

Over the last 50 years, the posttranslational modification (PTM) of proteins has emerged as a central mechanism for cells to regulate metabolism, growth, differentiation, cell-cell interactions, and immune responses. By influencing protein structure and function, PTM leads to a multiplication of proteome diversity. Redox-dependent PTMs, mediated by environmental and endogenously generated reactive species, induce cell signaling responses and can have toxic effects in organisms. PTMs induced by the electrophilic by-products of redox reactions most frequently occur at protein thiols; other nucleophilic amino acids serve as less favorable targets. Advances in mass spectrometry and affinity-chemistry strategies have improved the detection of electrophile-induced protein modifications both in vitro and in vivo and have revealed a high degree of amino acid and protein selectivity of electrophilic PTM. The identification of biological targets of electrophiles has motivated further study of the functional impact of various PTM reactions on specific signaling pathways and how this might affect organisms. PMID:19797270

How many human proteoforms are there?

PubMed

Aebersold, Ruedi; Agar, Jeffrey N; Amster, I Jonathan; Baker, Mark S; Bertozzi, Carolyn R; Boja, Emily S; Costello, Catherine E; Cravatt, Benjamin F; Fenselau, Catherine; Garcia, Benjamin A; Ge, Ying; Gunawardena, Jeremy; Hendrickson, Ronald C; Hergenrother, Paul J; Huber, Christian G; Ivanov, Alexander R; Jensen, Ole N; Jewett, Michael C; Kelleher, Neil L; Kiessling, Laura L; Krogan, Nevan J; Larsen, Martin R; Loo, Joseph A; Ogorzalek Loo, Rachel R; Lundberg, Emma; MacCoss, Michael J; Mallick, Parag; Mootha, Vamsi K; Mrksich, Milan; Muir, Tom W; Patrie, Steven M; Pesavento, James J; Pitteri, Sharon J; Rodriguez, Henry; Saghatelian, Alan; Sandoval, Wendy; Schlüter, Hartmut; Sechi, Salvatore; Slavoff, Sarah A; Smith, Lloyd M; Snyder, Michael P; Thomas, Paul M; Uhlén, Mathias; Van Eyk, Jennifer E; Vidal, Marc; Walt, David R; White, Forest M; Williams, Evan R; Wohlschlager, Therese; Wysocki, Vicki H; Yates, Nathan A; Young, Nicolas L; Zhang, Bing

2018-02-14

Despite decades of accumulated knowledge about proteins and their post-translational modifications (PTMs), numerous questions remain regarding their molecular composition and biological function. One of the most fundamental queries is the extent to which the combinations of DNA-, RNA- and PTM-level variations explode the complexity of the human proteome. Here, we outline what we know from current databases and measurement strategies including mass spectrometry-based proteomics. In doing so, we examine prevailing notions about the number of modifications displayed on human proteins and how they combine to generate the protein diversity underlying health and disease. We frame central issues regarding determination of protein-level variation and PTMs, including some paradoxes present in the field today. We use this framework to assess existing data and to ask the question, "How many distinct primary structures of proteins (proteoforms) are created from the 20,300 human genes?" We also explore prospects for improving measurements to better regularize protein-level biology and efficiently associate PTMs to function and phenotype.

Short- and long-term memory are modulated by multiple isoforms of the fragile X mental retardation protein.

PubMed

Banerjee, Paromita; Schoenfeld, Brian P; Bell, Aaron J; Choi, Catherine H; Bradley, Michael P; Hinchey, Paul; Kollaros, Maria; Park, Jae H; McBride, Sean M J; Dockendorff, Thomas C

2010-05-12

The diversity of protein isoforms arising from alternative splicing is thought to modulate fine-tuning of synaptic plasticity. Fragile X mental retardation protein (FMRP), a neuronal RNA binding protein, exists in isoforms as a result of alternative splicing, but the contribution of these isoforms to neural plasticity are not well understood. We show that two isoforms of Drosophila melanogaster FMRP (dFMR1) have differential roles in mediating neural development and behavior functions conferred by the dfmr1 gene. These isoforms differ in the presence of a protein interaction module that is related to prion domains and is functionally conserved between FMRPs. Expression of both isoforms is necessary for optimal performance in tests of short- and long-term memory of courtship training. The presence or absence of the protein interaction domain may govern the types of ribonucleoprotein (RNP) complexes dFMR1 assembles into, with different RNPs regulating gene expression in a manner necessary for establishing distinct phases of memory formation.

7TMRmine: a Web server for hierarchical mining of 7TMR proteins

PubMed Central

Lu, Guoqing; Wang, Zhifang; Jones, Alan M; Moriyama, Etsuko N

2009-01-01

Background Seven-transmembrane region-containing receptors (7TMRs) play central roles in eukaryotic signal transduction. Due to their biomedical importance, thorough mining of 7TMRs from diverse genomes has been an active target of bioinformatics and pharmacogenomics research. The need for new and accurate 7TMR/GPCR prediction tools is paramount with the accelerated rate of acquisition of diverse sequence information. Currently available and often used protein classification methods (e.g., profile hidden Markov Models) are highly accurate for identifying their membership information among already known 7TMR subfamilies. However, these alignment-based methods are less effective for identifying remote similarities, e.g., identifying proteins from highly divergent or possibly new 7TMR families. In this regard, more sensitive (e.g., alignment-free) methods are needed to complement the existing protein classification methods. A better strategy would be to combine different classifiers, from more specific to more sensitive methods, to identify a broader spectrum of 7TMR protein candidates. Description We developed a Web server, 7TMRmine, by integrating alignment-free and alignment-based classifiers specifically trained to identify candidate 7TMR proteins as well as transmembrane (TM) prediction methods. This new tool enables researchers to easily assess the distribution of GPCR functionality in diverse genomes or individual newly-discovered proteins. 7TMRmine is easily customized and facilitates exploratory analysis of diverse genomes. Users can integrate various alignment-based, alignment-free, and TM-prediction methods in any combination and in any hierarchical order. Sixteen classifiers (including two TM-prediction methods) are available on the 7TMRmine Web server. Not only can the 7TMRmine tool be used for 7TMR mining, but also for general TM-protein analysis. Users can submit protein sequences for analysis, or explore pre-analyzed results for multiple genomes. The server currently includes prediction results and the summary statistics for 68 genomes. Conclusion 7TMRmine facilitates the discovery of 7TMR proteins. By combining prediction results from different classifiers in a multi-level filtering process, prioritized sets of 7TMR candidates can be obtained for further investigation. 7TMRmine can be also used as a general TM-protein classifier. Comparisons of TM and 7TMR protein distributions among 68 genomes revealed interesting differences in evolution of these protein families among major eukaryotic phyla. PMID:19538753

Functions of Intracellular Retinoid Binding-Proteins.

PubMed

Napoli, Joseph L

Multiple binding and transport proteins facilitate many aspects of retinoid biology through effects on retinoid transport, cellular uptake, metabolism, and nuclear delivery. These include the serum retinol binding protein sRBP (aka Rbp4), the plasma membrane sRBP receptor Stra6, and the intracellular retinoid binding-proteins such as cellular retinol-binding proteins (CRBP) and cellular retinoic acid binding-proteins (CRABP). sRBP transports the highly lipophilic retinol through an aqueous medium. The major intracellular retinol-binding protein, CRBP1, likely enhances efficient retinoid use by providing a sink to facilitate retinol uptake from sRBP through the plasma membrane or via Stra6, delivering retinol or retinal to select enzymes that generate retinyl esters or retinoic acid, and protecting retinol/retinal from excess catabolism or opportunistic metabolism. Intracellular retinoic acid binding-proteins (CRABP1 and 2, and FABP5) seem to have more diverse functions distinctive to each, such as directing retinoic acid to catabolism, delivering retinoic acid to specific nuclear receptors, and generating non-canonical actions. Gene ablation of intracellular retinoid binding-proteins does not cause embryonic lethality or gross morphological defects. Metabolic and functional defects manifested in knockouts of CRBP1, CRBP2 and CRBP3, however, illustrate their essentiality to health, and in the case of CRBP2, to survival during limited dietary vitamin A. Future studies should continue to address the specific molecular interactions that occur between retinoid binding-proteins and their targets and their precise physiologic contributions to retinoid homeostasis and function.

Biodiversity enhances reef fish biomass and resistance to climate change.

PubMed

Duffy, J Emmett; Lefcheck, Jonathan S; Stuart-Smith, Rick D; Navarrete, Sergio A; Edgar, Graham J

2016-05-31

Fishes are the most diverse group of vertebrates, play key functional roles in aquatic ecosystems, and provide protein for a billion people, especially in the developing world. Those functions are compromised by mounting pressures on marine biodiversity and ecosystems. Because of its economic and food value, fish biomass production provides an unusually direct link from biodiversity to critical ecosystem services. We used the Reef Life Survey's global database of 4,556 standardized fish surveys to test the importance of biodiversity to fish production relative to 25 environmental drivers. Temperature, biodiversity, and human influence together explained 47% of the global variation in reef fish biomass among sites. Fish species richness and functional diversity were among the strongest predictors of fish biomass, particularly for the large-bodied species and carnivores preferred by fishers, and these biodiversity effects were robust to potentially confounding influences of sample abundance, scale, and environmental correlations. Warmer temperatures increased biomass directly, presumably by raising metabolism, and indirectly by increasing diversity, whereas temperature variability reduced biomass. Importantly, diversity and climate interact, with biomass of diverse communities less affected by rising and variable temperatures than species-poor communities. Biodiversity thus buffers global fish biomass from climate change, and conservation of marine biodiversity can stabilize fish production in a changing ocean.

Biodiversity enhances reef fish biomass and resistance to climate change

PubMed Central

Duffy, J. Emmett; Lefcheck, Jonathan S.; Navarrete, Sergio A.; Edgar, Graham J.

2016-01-01

Fishes are the most diverse group of vertebrates, play key functional roles in aquatic ecosystems, and provide protein for a billion people, especially in the developing world. Those functions are compromised by mounting pressures on marine biodiversity and ecosystems. Because of its economic and food value, fish biomass production provides an unusually direct link from biodiversity to critical ecosystem services. We used the Reef Life Survey’s global database of 4,556 standardized fish surveys to test the importance of biodiversity to fish production relative to 25 environmental drivers. Temperature, biodiversity, and human influence together explained 47% of the global variation in reef fish biomass among sites. Fish species richness and functional diversity were among the strongest predictors of fish biomass, particularly for the large-bodied species and carnivores preferred by fishers, and these biodiversity effects were robust to potentially confounding influences of sample abundance, scale, and environmental correlations. Warmer temperatures increased biomass directly, presumably by raising metabolism, and indirectly by increasing diversity, whereas temperature variability reduced biomass. Importantly, diversity and climate interact, with biomass of diverse communities less affected by rising and variable temperatures than species-poor communities. Biodiversity thus buffers global fish biomass from climate change, and conservation of marine biodiversity can stabilize fish production in a changing ocean. PMID:27185921

Functional diversification of sea urchin ABCC1 (MRP1) by alternative splicing.

PubMed

Gökirmak, Tufan; Campanale, Joseph P; Reitzel, Adam M; Shipp, Lauren E; Moy, Gary W; Hamdoun, Amro

2016-06-01

The multidrug resistance protein (MRP) family encodes a diverse repertoire of ATP-binding cassette (ABC) transporters with multiple roles in development, disease, and homeostasis. Understanding MRP evolution is central to unraveling their roles in these diverse processes. Sea urchins occupy an important phylogenetic position for understanding the evolution of vertebrate proteins and have been an important invertebrate model system for study of ABC transporters. We used phylogenetic analyses to examine the evolution of MRP transporters and functional approaches to identify functional forms of sea urchin MRP1 (also known as SpABCC1). SpABCC1, the only MRP homolog in sea urchins, is co-orthologous to human MRP1, MRP3, and MRP6 (ABCC1, ABCC3, and ABCC6) transporters. However, efflux assays revealed that alternative splicing of exon 22, a region critical for substrate interactions, could diversify functions of sea urchin MRP1. Phylogenetic comparisons also indicate that while MRP1, MRP3, and MRP6 transporters potentially arose from a single transporter in basal deuterostomes, alternative splicing appears to have been the major mode of functional diversification in invertebrates, while duplication may have served a more important role in vertebrates. These results provide a deeper understanding of the evolutionary origins of MRP transporters and the potential mechanisms used to diversify their functions in different groups of animals. Copyright © 2016 the American Physiological Society.

Functional diversification of sea urchin ABCC1 (MRP1) by alternative splicing

PubMed Central

Gökirmak, Tufan; Campanale, Joseph P.; Reitzel, Adam M.; Shipp, Lauren E.; Moy, Gary W.

2016-01-01

The multidrug resistance protein (MRP) family encodes a diverse repertoire of ATP-binding cassette (ABC) transporters with multiple roles in development, disease, and homeostasis. Understanding MRP evolution is central to unraveling their roles in these diverse processes. Sea urchins occupy an important phylogenetic position for understanding the evolution of vertebrate proteins and have been an important invertebrate model system for study of ABC transporters. We used phylogenetic analyses to examine the evolution of MRP transporters and functional approaches to identify functional forms of sea urchin MRP1 (also known as SpABCC1). SpABCC1, the only MRP homolog in sea urchins, is co-orthologous to human MRP1, MRP3, and MRP6 (ABCC1, ABCC3, and ABCC6) transporters. However, efflux assays revealed that alternative splicing of exon 22, a region critical for substrate interactions, could diversify functions of sea urchin MRP1. Phylogenetic comparisons also indicate that while MRP1, MRP3, and MRP6 transporters potentially arose from a single transporter in basal deuterostomes, alternative splicing appears to have been the major mode of functional diversification in invertebrates, while duplication may have served a more important role in vertebrates. These results provide a deeper understanding of the evolutionary origins of MRP transporters and the potential mechanisms used to diversify their functions in different groups of animals. PMID:27053522

Intragenome Diversity of Gene Families Encoding Toxin-like Proteins in Venomous Animals.

PubMed

Rodríguez de la Vega, Ricardo C; Giraud, Tatiana

2016-11-01

The evolution of venoms is the story of how toxins arise and of the processes that generate and maintain their diversity. For animal venoms these processes include recruitment for expression in the venom gland, neofunctionalization, paralogous expansions, and functional divergence. The systematic study of these processes requires the reliable identification of the venom components involved in antagonistic interactions. High-throughput sequencing has the potential of uncovering the entire set of toxins in a given organism, yet the existence of non-venom toxin paralogs and the misleading effects of partial census of the molecular diversity of toxins make necessary to collect complementary evidence to distinguish true toxins from their non-venom paralogs. Here, we analyzed the whole genomes of two scorpions, one spider and one snake, aiming at the identification of the full repertoires of genes encoding toxin-like proteins. We classified the entire set of protein-coding genes into paralogous groups and monotypic genes, identified genes encoding toxin-like proteins based on known toxin families, and quantified their expression in both venom-glands and pooled tissues. Our results confirm that genes encoding toxin-like proteins are part of multigene families, and that these families arise by recruitment events from non-toxin genes followed by limited expansions of the toxin-like protein coding genes. We also show that failing to account for sequence similarity with non-toxin proteins has a considerable misleading effect that can be greatly reduced by comparative transcriptomics. Our study overall contributes to the understanding of the evolutionary dynamics of proteins involved in antagonistic interactions. © The Author 2016. Published by Oxford University Press on behalf of the Society for Integrative and Comparative Biology. All rights reserved. For permissions please email: journals.permissions@oup.com.

The BAR Domain Proteins: Molding Membranes in Fission, Fusion, and Phagy

PubMed Central

Ren, Gang; Vajjhala, Parimala; Lee, Janet S.; Winsor, Barbara; Munn, Alan L.

2006-01-01

The Bin1/amphiphysin/Rvs167 (BAR) domain proteins are a ubiquitous protein family. Genes encoding members of this family have not yet been found in the genomes of prokaryotes, but within eukaryotes, BAR domain proteins are found universally from unicellular eukaryotes such as yeast through to plants, insects, and vertebrates. BAR domain proteins share an N-terminal BAR domain with a high propensity to adopt α-helical structure and engage in coiled-coil interactions with other proteins. BAR domain proteins are implicated in processes as fundamental and diverse as fission of synaptic vesicles, cell polarity, endocytosis, regulation of the actin cytoskeleton, transcriptional repression, cell-cell fusion, signal transduction, apoptosis, secretory vesicle fusion, excitation-contraction coupling, learning and memory, tissue differentiation, ion flux across membranes, and tumor suppression. What has been lacking is a molecular understanding of the role of the BAR domain protein in each process. The three-dimensional structure of the BAR domain has now been determined and valuable insight has been gained in understanding the interactions of BAR domains with membranes. The cellular roles of BAR domain proteins, characterized over the past decade in cells as distinct as yeasts, neurons, and myocytes, can now be understood in terms of a fundamental molecular function of all BAR domain proteins: to sense membrane curvature, to bind GTPases, and to mold a diversity of cellular membranes. PMID:16524918

Identification of a Unique Fe-S Cluster Binding Site in a Glycyl-Radical Type Microcompartment Shell Protein

PubMed Central

Thompson, Michael C.; Wheatley, Nicole M.; Jorda, Julien; Sawaya, Michael R.; Gidaniyan, Soheil D.; Ahmed, Hoda; Yang, Zhongyu; McCarty, Krystal N.; Whitelegge, Julian P.; Yeates, Todd O.

2014-01-01

Recently, progress has been made toward understanding the functional diversity of bacterial microcompartment (MCP) systems, which serve as protein-based metabolic organelles in diverse microbes. New types of MCPs have been identified, including the glycyl-radical propanediol (Grp) MCP. Within these elaborate protein complexes, BMC-domain shell proteins assemble to form a polyhedral barrier that encapsulates the enzymatic contents of the MCP. Interestingly, the Grp MCP contains a number of shell proteins with unusual sequence features. GrpU is one such shell protein, whose amino acid sequence is particularly divergent from other members of the BMC-domain superfamily of proteins that effectively defines all MCPs. Expression, purification, and subsequent characterization of the protein showed, unexpectedly, that it binds an iron-sulfur cluster. We determined X-ray crystal structures of two GrpU orthologs, providing the first structural insight into the homohexameric BMC-domain shell proteins of the Grp system. The X-ray structures of GrpU, both obtained in the apo form, combined with spectroscopic analyses and computational modeling, show that the metal cluster resides in the central pore of the BMC shell protein at a position of broken 6-fold symmetry. The result is a structurally polymorphic iron-sulfur cluster binding site that appears to be unique among metalloproteins studied to date. PMID:25102080

Structural Analysis of PTM Hotspots (SAPH-ire)--A Quantitative Informatics Method Enabling the Discovery of Novel Regulatory Elements in Protein Families.

PubMed

Dewhurst, Henry M; Choudhury, Shilpa; Torres, Matthew P

2015-08-01

Predicting the biological function potential of post-translational modifications (PTMs) is becoming increasingly important in light of the exponential increase in available PTM data from high-throughput proteomics. We developed structural analysis of PTM hotspots (SAPH-ire)--a quantitative PTM ranking method that integrates experimental PTM observations, sequence conservation, protein structure, and interaction data to allow rank order comparisons within or between protein families. Here, we applied SAPH-ire to the study of PTMs in diverse G protein families, a conserved and ubiquitous class of proteins essential for maintenance of intracellular structure (tubulins) and signal transduction (large and small Ras-like G proteins). A total of 1728 experimentally verified PTMs from eight unique G protein families were clustered into 451 unique hotspots, 51 of which have a known and cited biological function or response. Using customized software, the hotspots were analyzed in the context of 598 unique protein structures. By comparing distributions of hotspots with known versus unknown function, we show that SAPH-ire analysis is predictive for PTM biological function. Notably, SAPH-ire revealed high-ranking hotspots for which a functional impact has not yet been determined, including phosphorylation hotspots in the N-terminal tails of G protein gamma subunits--conserved protein structures never before reported as regulators of G protein coupled receptor signaling. To validate this prediction we used the yeast model system for G protein coupled receptor signaling, revealing that gamma subunit-N-terminal tail phosphorylation is activated in response to G protein coupled receptor stimulation and regulates protein stability in vivo. These results demonstrate the utility of integrating protein structural and sequence features into PTM prioritization schemes that can improve the analysis and functional power of modification-specific proteomics data. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

The alpha subunit of Go interacts with promyelocytic leukemia zinc finger protein and modulates its functions.

PubMed

Won, Jung Hee; Park, Jung Sik; Ju, Hyun Hee; Kim, Soyeon; Suh-Kim, Haeyoung; Ghil, Sung Ho

2008-05-01

Heterotrimeric GTP-binding proteins (G proteins) mediate signal transduction generated by neurotransmitters and hormones. Go, a member of the Go/Gi family, is the most abundant heterotrimeric G protein in the brain. Most mechanistic analyses on Go activation demonstrate that its action is mediated by the Gbetagamma dimer; downstream effectors for its alpha subunit (Goalpha) have not been clearly defined. Here, we employ the yeast two-hybrid system to screen for Goalpha-interacting partners in a cDNA library from human fetal brain. The transcription factor promyelocytic leukemia zinc finger protein (PLZF) specifically bound to Goalpha. Interactions between PLZF and Goalpha were confirmed using in vitro and in vivo affinity binding assays. Activated Goalpha interacted directly with PLZF, and enhanced its function as a transcriptional and cell growth suppressor. Notably, PLZF activity was additionally promoted by the Go/ialpha-coupled cannabinoid receptor (CB) in HL60 cells endogenously expressing CB and PLZF. These results collectively suggest that Goalpha modulates the function of PLZF via direct interactions. Our novel findings provide insights into the diverse cellular roles of Goalpha and its coupled receptor.

Current Progress in Tonoplast Proteomics Reveals Insights into the Function of the Large Central Vacuole

PubMed Central

Trentmann, Oliver; Haferkamp, Ilka

2013-01-01

Vacuoles of plants fulfill various biologically important functions, like turgor generation and maintenance, detoxification, solute sequestration, or protein storage. Different types of plant vacuoles (lytic versus protein storage) are characterized by different functional properties apparently caused by a different composition/abundance and regulation of transport proteins in the surrounding membrane, the tonoplast. Proteome analyses allow the identification of vacuolar proteins and provide an informative basis for assigning observed transport processes to specific carriers or channels. This review summarizes techniques required for vacuolar proteome analyses, like e.g., isolation of the large central vacuole or tonoplast membrane purification. Moreover, an overview about diverse published vacuolar proteome studies is provided. It becomes evident that qualitative proteomes from different plant species represent just the tip of the iceberg. During the past few years, mass spectrometry achieved immense improvement concerning its accuracy, sensitivity, and application. As a consequence, modern tonoplast proteome approaches are suited for detecting alterations in membrane protein abundance in response to changing environmental/physiological conditions and help to clarify the regulation of tonoplast transport processes. PMID:23459586

ATP-independent diffusion of double-stranded RNA binding proteins

PubMed Central

Koh, Hye Ran; Kidwell, Mary Anne; Ragunathan, Kaushik; Doudna, Jennifer A.; Myong, Sua

2013-01-01

The proteins harboring double-stranded RNA binding domains (dsRBDs) play diverse functional roles such as RNA localization, splicing, editing, export, and translation, yet mechanistic basis and functional significance of dsRBDs remain unclear. To unravel this enigma, we investigated transactivation response RNA binding protein (TRBP) consisting of three dsRBDs, which functions in HIV replication, protein kinase R(PKR)–mediated immune response, and RNA silencing. Here we report an ATP-independent diffusion activity of TRBP exclusively on dsRNA in a length-dependent manner. The first two dsRBDs of TRBP are essential for diffusion, whereas the third dsRBD is dispensable. Two homologs of TRBP, PKR activator and R3D1-L, displayed the same diffusion, implying a universality of the diffusion activity among this protein family. Furthermore, a Dicer–TRBP complex on dsRNA exhibited dynamic diffusion, which was correlated with Dicer’s catalytic activity. These results implicate the dsRNA-specific diffusion activity of TRBP that contributes to enhancing siRNA and miRNA processing by Dicer. PMID:23251028

Protein complexes, big data, machine learning and integrative proteomics: lessons learned over a decade of systematic analysis of protein interaction networks.

PubMed

Havugimana, Pierre C; Hu, Pingzhao; Emili, Andrew

2017-10-01

Elucidation of the networks of physical (functional) interactions present in cells and tissues is fundamental for understanding the molecular organization of biological systems, the mechanistic basis of essential and disease-related processes, and for functional annotation of previously uncharacterized proteins (via guilt-by-association or -correlation). After a decade in the field, we felt it timely to document our own experiences in the systematic analysis of protein interaction networks. Areas covered: Researchers worldwide have contributed innovative experimental and computational approaches that have driven the rapidly evolving field of 'functional proteomics'. These include mass spectrometry-based methods to characterize macromolecular complexes on a global-scale and sophisticated data analysis tools - most notably machine learning - that allow for the generation of high-quality protein association maps. Expert commentary: Here, we recount some key lessons learned, with an emphasis on successful workflows, and challenges, arising from our own and other groups' ongoing efforts to generate, interpret and report proteome-scale interaction networks in increasingly diverse biological contexts.

Functions of IQD proteins as hubs in cellular calcium and auxin signaling: A toolbox for shape formation and tissue-specification in plants?

PubMed

Bürstenbinder, Katharina; Mitra, Dipannita; Quegwer, Jakob

2017-06-03

Calcium (Ca 2+ ) ions play pivotal roles as second messengers in intracellular signal transduction, and coordinate many biological processes. Changes in intracellular Ca 2+ levels are perceived by Ca 2+ sensors such as calmodulin (CaM) and CaM-like (CML) proteins, which transduce Ca 2+ signals into cellular responses by regulation of diverse target proteins. Insights into molecular functions of CaM targets are thus essential to understand the molecular and cellular basis of Ca 2+ signaling. During the last decade, IQ67-domain (IQD) proteins emerged as the largest class of CaM targets in plants with mostly unknown functions. In the March issue of Plant Physiology, we presented the first comprehensive characterization of the 33-membered IQD family in Arabidopsis thaliana. We showed, by analysis of the subcellular localization of translational green fluorescent protein (GFP) fusion proteins, that most IQD members label microtubules (MTs), and additionally often localize to the cell nucleus or to membranes, where they recruit CaM Ca 2+ sensors. Important functions at MTs are supported by altered MT organization and plant growth in IQD gain-of-function lines. Because IQD proteins share structural hallmarks of scaffold proteins, we propose roles of IQDs in the assembly of macromolecular complexes to orchestrate Ca 2+ CaM signaling from membranes to the nucleus. Interestingly, expression of several IQDs is regulated by auxin, which suggests functions of IQDs as hubs in cellular auxin and calcium signaling to regulate plant growth and development.

Polypyrimidine Tract Binding Protein Homologs from Arabidopsis Are Key Regulators of Alternative Splicing with Implications in Fundamental Developmental Processes[W

PubMed Central

Rühl, Christina; Stauffer, Eva; Kahles, André; Wagner, Gabriele; Drechsel, Gabriele; Rätsch, Gunnar; Wachter, Andreas

2012-01-01

Alternative splicing (AS) generates transcript variants by variable exon/intron definition and massively expands transcriptome diversity. Changes in AS patterns have been found to be linked to manifold biological processes, yet fundamental aspects, such as the regulation of AS and its functional implications, largely remain to be addressed. In this work, widespread AS regulation by Arabidopsis thaliana Polypyrimidine tract binding protein homologs (PTBs) was revealed. In total, 452 AS events derived from 307 distinct genes were found to be responsive to the levels of the splicing factors PTB1 and PTB2, which predominantly triggered splicing of regulated introns, inclusion of cassette exons, and usage of upstream 5′ splice sites. By contrast, no major AS regulatory function of the distantly related PTB3 was found. Dependent on their position within the mRNA, PTB-regulated events can both modify the untranslated regions and give rise to alternative protein products. We find that PTB-mediated AS events are connected to diverse biological processes, and the functional implications of selected instances were further elucidated. Specifically, PTB misexpression changes AS of PHYTOCHROME INTERACTING FACTOR6, coinciding with altered rates of abscisic acid–dependent seed germination. Furthermore, AS patterns as well as the expression of key flowering regulators were massively changed in a PTB1/2 level-dependent manner. PMID:23192226

Immunoglobulin superfamily members encoded by viruses and their multiple roles in immune evasion.

PubMed

Farré, Domènec; Martínez-Vicente, Pablo; Engel, Pablo; Angulo, Ana

2017-05-01

Pathogens have developed a plethora of strategies to undermine host immune defenses in order to guarantee their survival. For large DNA viruses, these immune evasion mechanisms frequently rely on the expression of genes acquired from host genomes. Horizontally transferred genes include members of the immunoglobulin superfamily, whose products constitute the most diverse group of proteins of vertebrate genomes. Their promiscuous immunoglobulin domains, which comprise the building blocks of these molecules, are involved in a large variety of functions mediated by ligand-binding interactions. The flexible structural nature of the immunoglobulin domains makes them appealing targets for viral capture due to their capacity to generate high functional diversity. Here, we present an up-to-date review of immunoglobulin superfamily gene homologs encoded by herpesviruses, poxviruses, and adenoviruses, that include CD200, CD47, Fc receptors, interleukin-1 receptor 2, interleukin-18 binding protein, CD80, carcinoembryonic antigen-related cell adhesion molecules, and signaling lymphocyte activation molecules. We discuss their distinct structural attributes, binding properties, and functions, shaped by evolutionary pressures to disarm specific immune pathways. We include several novel genes identified from extensive genome database surveys. An understanding of the properties and modes of action of these viral proteins may guide the development of novel immune-modulatory therapeutic tools. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Unveiling the functional diversity of the Alpha-Beta hydrolase fold in plants

PubMed Central

Mindrebo, Jeffrey T.; Nartey, Charisse M.; Seto, Yoshiya; Burkart, Michael D.; Noel, Joseph P.

2017-01-01

The alpha/beta hydrolase (ABH) superfamily is a widespread and functionally malleable protein fold recognized for its diverse biochemical activities across all three domains of life. ABH enzymes possess unexpected catalytic activity in the green plant lineage through selective alterations in active site architecture and chemistry. Furthermore, the ABH fold serves as the core structure for phytohormone and ligand receptors in the gibberellin, strigolactone, and karrikin signaling pathways in plants. Despite recent discoveries, the ABH family is sparsely characterized in plants, a sessile kingdom known to evolve complex and specialized chemical adaptations as survival responses to widely varying biotic and abiotic ecologies. This review calls attention to the ABH superfamily in the plant kingdom to highlight the functional adaptability of the ABH fold. PMID:27662376

Antimicrobial Functions of Lactoferrin Promote Genetic Conflicts in Ancient Primates and Modern Humans.

PubMed

Barber, Matthew F; Kronenberg, Zev; Yandell, Mark; Elde, Nels C

2016-05-01

Lactoferrin is a multifunctional mammalian immunity protein that limits microbial growth through sequestration of nutrient iron. Additionally, lactoferrin possesses cationic protein domains that directly bind and inhibit diverse microbes. The implications for these dual functions on lactoferrin evolution and genetic conflicts with microbes remain unclear. Here we show that lactoferrin has been subject to recurrent episodes of positive selection during primate divergence predominately at antimicrobial peptide surfaces consistent with long-term antagonism by bacteria. An abundant lactoferrin polymorphism in human populations and Neanderthals also exhibits signatures of positive selection across primates, linking ancient host-microbe conflicts to modern human genetic variation. Rapidly evolving sites in lactoferrin further correspond to molecular interfaces with opportunistic bacterial pathogens causing meningitis, pneumonia, and sepsis. Because microbes actively target lactoferrin to acquire iron, we propose that the emergence of antimicrobial activity provided a pivotal mechanism of adaptation sparking evolutionary conflicts via acquisition of new protein functions.

The liver in regulation of iron homeostasis.

PubMed

Rishi, Gautam; Subramaniam, V Nathan

2017-09-01

The liver is one of the largest and most functionally diverse organs in the human body. In addition to roles in detoxification of xenobiotics, digestion, synthesis of important plasma proteins, gluconeogenesis, lipid metabolism, and storage, the liver also plays a significant role in iron homeostasis. Apart from being the storage site for excess body iron, it also plays a vital role in regulating the amount of iron released into the blood by enterocytes and macrophages. Since iron is essential for many important physiological and molecular processes, it increases the importance of liver in the proper functioning of the body's metabolism. This hepatic iron-regulatory function can be attributed to the expression of many liver-specific or liver-enriched proteins, all of which play an important role in the regulation of iron homeostasis. This review focuses on these proteins and their known roles in the regulation of body iron metabolism. Copyright © 2017 the American Physiological Society.

Pili and flagella biology, structure, and biotechnological applications.

PubMed

Van Gerven, Nani; Waksman, Gabriel; Remaut, Han

2011-01-01

Bacteria and Archaea expose on their outer surfaces a variety of thread-like proteinaceous organelles with which they interact with their environments. These structures are repetitive assemblies of covalently or non-covalently linked protein subunits, organized into filamentous polymers known as pili ("hair"), flagella ("whips") or injectisomes ("needles"). They serve different roles in cell motility, adhesion and host invasion, protein and DNA secretion and uptake, conductance, or cellular encapsulation. Here we describe the functional, morphological and genetic diversity of these bacterial filamentous protein structures. The organized, multi-copy build-up and/or the natural function of pili and flagella have lead to their biotechnological application as display and secretion tools, as therapeutic targets or as molecular motors. We review the documented and potential technological exploitation of bacterial surface filaments in light of their structural and functional traits. Copyright © 2011 Elsevier Inc. All rights reserved.

Collagens--structure, function, and biosynthesis.

PubMed

Gelse, K; Pöschl, E; Aigner, T

2003-11-28

The extracellular matrix represents a complex alloy of variable members of diverse protein families defining structural integrity and various physiological functions. The most abundant family is the collagens with more than 20 different collagen types identified so far. Collagens are centrally involved in the formation of fibrillar and microfibrillar networks of the extracellular matrix, basement membranes as well as other structures of the extracellular matrix. This review focuses on the distribution and function of various collagen types in different tissues. It introduces their basic structural subunits and points out major steps in the biosynthesis and supramolecular processing of fibrillar collagens as prototypical members of this protein family. A final outlook indicates the importance of different collagen types not only for the understanding of collagen-related diseases, but also as a basis for the therapeutical use of members of this protein family discussed in other chapters of this issue.

[GPCRs heterodimerization: a new way towards the discovery of function for the orphan receptors?].

PubMed

Levoye, Angélique; Jockers, Ralf

2007-01-01

G protein-coupled receptors (GPCRs), also called seven transmembrane domain (7TM) proteins, represent the largest family of cell surface receptors. GPCRs control a variety of physiological processes, are involved in multiple diseases and are major drug targets. Despite a vast effort of academic and industrial research, more than one hundred receptors remain orphans. These orphan GPCRs offer a great potential for drug discovery, as almost 60% of currently prescribed drugs target GPCRs. Deorphenization strategies have concentrated mainly on the identification of the natural ligands of these proteins. Recent advances have shown that orphan GPCRs, similar to orphan nuclear receptors, can regulate the function of non-orphan receptors by heterodimerization. These findings not only help to better understand the extraordinary diversity of GPCRs, but also open new perspectives for the identification of the function of these orphan receptors that hold great therapeutic potential.

Structural Elements Regulating AAA+ Protein Quality Control Machines.

PubMed

Chang, Chiung-Wen; Lee, Sukyeong; Tsai, Francis T F

2017-01-01

Members of the ATPases Associated with various cellular Activities (AAA+) superfamily participate in essential and diverse cellular pathways in all kingdoms of life by harnessing the energy of ATP binding and hydrolysis to drive their biological functions. Although most AAA+ proteins share a ring-shaped architecture, AAA+ proteins have evolved distinct structural elements that are fine-tuned to their specific functions. A central question in the field is how ATP binding and hydrolysis are coupled to substrate translocation through the central channel of ring-forming AAA+ proteins. In this mini-review, we will discuss structural elements present in AAA+ proteins involved in protein quality control, drawing similarities to their known role in substrate interaction by AAA+ proteins involved in DNA translocation. Elements to be discussed include the pore loop-1, the Inter-Subunit Signaling (ISS) motif, and the Pre-Sensor I insert (PS-I) motif. Lastly, we will summarize our current understanding on the inter-relationship of those structural elements and propose a model how ATP binding and hydrolysis might be coupled to polypeptide translocation in protein quality control machines.

Identification of novel phosphatidic acid-binding proteins in the rat brain.

PubMed

Park, ChiHu; Kang, Du-Seock; Shin, Geon-Hoon; Seo, Jeongkon; Kim, Hyein; Suh, Pann-Ghill; Bae, Chang-Dae; Shin, Joo-Ho

2015-05-19

Phosphatidic acid (PA) is an abundant negatively-charged phospholipid and has long been considered to be an important signaling molecule in diverse cellular events. Thus, the identification of proteins that specifically interact with PA is of considerable interest to understand the regulatory roles of PA. Herein, lipid-affinity purification and mass spectrometric analysis reveals 43 proteins, 19 known and 24 novel, as PA-binding proteins. A lipid-protein overlay assay confirmed that GDI1, PACSIN1, and DPYSL2 interact with not only with PA but also with other phospholipids. These results might be helpful for deciphering the functional effect of PA in the brain. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Coupling Protein Side-Chain and Backbone Flexibility Improves the Re-design of Protein-Ligand Specificity.

PubMed

Ollikainen, Noah; de Jong, René M; Kortemme, Tanja

2015-01-01

Interactions between small molecules and proteins play critical roles in regulating and facilitating diverse biological functions, yet our ability to accurately re-engineer the specificity of these interactions using computational approaches has been limited. One main difficulty, in addition to inaccuracies in energy functions, is the exquisite sensitivity of protein-ligand interactions to subtle conformational changes, coupled with the computational problem of sampling the large conformational search space of degrees of freedom of ligands, amino acid side chains, and the protein backbone. Here, we describe two benchmarks for evaluating the accuracy of computational approaches for re-engineering protein-ligand interactions: (i) prediction of enzyme specificity altering mutations and (ii) prediction of sequence tolerance in ligand binding sites. After finding that current state-of-the-art "fixed backbone" design methods perform poorly on these tests, we develop a new "coupled moves" design method in the program Rosetta that couples changes to protein sequence with alterations in both protein side-chain and protein backbone conformations, and allows for changes in ligand rigid-body and torsion degrees of freedom. We show significantly increased accuracy in both predicting ligand specificity altering mutations and binding site sequences. These methodological improvements should be useful for many applications of protein-ligand design. The approach also provides insights into the role of subtle conformational adjustments that enable functional changes not only in engineering applications but also in natural protein evolution.

Lysine acetylome profiling uncovers novel histone deacetylase substrate proteins in Arabidopsis.

PubMed

Hartl, Markus; Füßl, Magdalena; Boersema, Paul J; Jost, Jan-Oliver; Kramer, Katharina; Bakirbas, Ahmet; Sindlinger, Julia; Plöchinger, Magdalena; Leister, Dario; Uhrig, Glen; Moorhead, Greg Bg; Cox, Jürgen; Salvucci, Michael E; Schwarzer, Dirk; Mann, Matthias; Finkemeier, Iris

2017-10-23

Histone deacetylases have central functions in regulating stress defenses and development in plants. However, the knowledge about the deacetylase functions is largely limited to histones, although these enzymes were found in diverse subcellular compartments. In this study, we determined the proteome-wide signatures of the RPD3/HDA1 class of histone deacetylases in Arabidopsis Relative quantification of the changes in the lysine acetylation levels was determined on a proteome-wide scale after treatment of Arabidopsis leaves with deacetylase inhibitors apicidin and trichostatin A. We identified 91 new acetylated candidate proteins other than histones, which are potential substrates of the RPD3/HDA1-like histone deacetylases in Arabidopsis , of which at least 30 of these proteins function in nucleic acid binding. Furthermore, our analysis revealed that histone deacetylase 14 (HDA14) is the first organellar-localized RPD3/HDA1 class protein found to reside in the chloroplasts and that the majority of its protein targets have functions in photosynthesis. Finally, the analysis of HDA14 loss-of-function mutants revealed that the activation state of RuBisCO is controlled by lysine acetylation of RuBisCO activase under low-light conditions. © 2017 The Authors. Published under the terms of the CC BY 4.0 license.

The cryo-electron microscopy structure of huntingtin

NASA Astrophysics Data System (ADS)

Guo, Qiang; Bin Huang; Cheng, Jingdong; Seefelder, Manuel; Engler, Tatjana; Pfeifer, Günter; Oeckl, Patrick; Otto, Markus; Moser, Franziska; Maurer, Melanie; Pautsch, Alexander; Baumeister, Wolfgang; Fernández-Busnadiego, Rubén; Kochanek, Stefan

2018-03-01

Huntingtin (HTT) is a large (348 kDa) protein that is essential for embryonic development and is involved in diverse cellular activities such as vesicular transport, endocytosis, autophagy and the regulation of transcription. Although an integrative understanding of the biological functions of HTT is lacking, the large number of identified HTT interactors suggests that it serves as a protein-protein interaction hub. Furthermore, Huntington’s disease is caused by a mutation in the HTT gene, resulting in a pathogenic expansion of a polyglutamine repeat at the amino terminus of HTT. However, only limited structural information regarding HTT is currently available. Here we use cryo-electron microscopy to determine the structure of full-length human HTT in a complex with HTT-associated protein 40 (HAP40; encoded by three F8A genes in humans) to an overall resolution of 4 Å. HTT is largely α-helical and consists of three major domains. The amino- and carboxy-terminal domains contain multiple HEAT (huntingtin, elongation factor 3, protein phosphatase 2A and lipid kinase TOR) repeats arranged in a solenoid fashion. These domains are connected by a smaller bridge domain containing different types of tandem repeats. HAP40 is also largely α-helical and has a tetratricopeptide repeat-like organization. HAP40 binds in a cleft and contacts the three HTT domains by hydrophobic and electrostatic interactions, thereby stabilizing the conformation of HTT. These data rationalize previous biochemical results and pave the way for improved understanding of the diverse cellular functions of HTT.