Electron Spin Relaxation: The Role of Spin-Orbit Coupling in Organic Semiconductors

NASA Astrophysics Data System (ADS)

Willis, M.; Nuccio, L.; Schulz, L.; Gillin, W.; Kreouzis, T.; Pratt, F.; Lord, J.; Heeney, M.; Fratini, S.; Bernhard, C.; Drew, A.

2012-02-01

Rapid development of organic materials has lead to their availability in commercial products. Until now, the spin degree of freedom has not generally been used in organic materials. As well as engineering difficulties, there are fundamental questions with respect to the electron spin relaxation (eSR) mechanisms in organic molecules. Muons used as a microscopic spin probe, localized to a single molecule, can access information needed to identify the relevant model for eSR. In this presentation I will introduce the ALC-MuSR technique describing how eSR can be extracted and the expected effects. I will show how the technique has been applied to small organic molecules such as the group III Quinolate series and functionalized molecules with a pentacene-like backbone. Lastly I will present the Z-number and temperature dependence in these organic molecules and show strong evidence for a spin-orbit based eSR mechanism.

Simultaneous optimization of biomolecular energy function on features from small molecules and macromolecules

PubMed Central

Park, Hahnbeom; Bradley, Philip; Greisen, Per; Liu, Yuan; Mulligan, Vikram Khipple; Kim, David E.; Baker, David; DiMaio, Frank

2017-01-01

Most biomolecular modeling energy functions for structure prediction, sequence design, and molecular docking, have been parameterized using existing macromolecular structural data; this contrasts molecular mechanics force fields which are largely optimized using small-molecule data. In this study, we describe an integrated method that enables optimization of a biomolecular modeling energy function simultaneously against small-molecule thermodynamic data and high-resolution macromolecular structural data. We use this approach to develop a next-generation Rosetta energy function that utilizes a new anisotropic implicit solvation model, and an improved electrostatics and Lennard-Jones model, illustrating how energy functions can be considerably improved in their ability to describe large-scale energy landscapes by incorporating both small-molecule and macromolecule data. The energy function improves performance in a wide range of protein structure prediction challenges, including monomeric structure prediction, protein-protein and protein-ligand docking, protein sequence design, and prediction of the free energy changes by mutation, while reasonably recapitulating small-molecule thermodynamic properties. PMID:27766851

Evaluation of Kirkwood-Buff integrals via finite size scaling: a large scale molecular dynamics study

NASA Astrophysics Data System (ADS)

Dednam, W.; Botha, A. E.

2015-01-01

Solvation of bio-molecules in water is severely affected by the presence of co-solvent within the hydration shell of the solute structure. Furthermore, since solute molecules can range from small molecules, such as methane, to very large protein structures, it is imperative to understand the detailed structure-function relationship on the microscopic level. For example, it is useful know the conformational transitions that occur in protein structures. Although such an understanding can be obtained through large-scale molecular dynamic simulations, it is often the case that such simulations would require excessively large simulation times. In this context, Kirkwood-Buff theory, which connects the microscopic pair-wise molecular distributions to global thermodynamic properties, together with the recently developed technique, called finite size scaling, may provide a better method to reduce system sizes, and hence also the computational times. In this paper, we present molecular dynamics trial simulations of biologically relevant low-concentration solvents, solvated by aqueous co-solvent solutions. In particular we compare two different methods of calculating the relevant Kirkwood-Buff integrals. The first (traditional) method computes running integrals over the radial distribution functions, which must be obtained from large system-size NVT or NpT simulations. The second, newer method, employs finite size scaling to obtain the Kirkwood-Buff integrals directly by counting the particle number fluctuations in small, open sub-volumes embedded within a larger reservoir that can be well approximated by a much smaller simulation cell. In agreement with previous studies, which made a similar comparison for aqueous co-solvent solutions, without the additional solvent, we conclude that the finite size scaling method is also applicable to the present case, since it can produce computationally more efficient results which are equivalent to the more costly radial distribution function method.

Small molecule dual-inhibitors of TRPV4 and TRPA1 for attenuation of inflammation and pain

PubMed Central

Kanju, Patrick; Chen, Yong; Lee, Whasil; Yeo, Michele; Lee, Suk Hee; Romac, Joelle; Shahid, Rafiq; Fan, Ping; Gooden, David M.; Simon, Sidney A.; Spasojevic, Ivan; Mook, Robert A.; Liddle, Rodger A.; Guilak, Farshid; Liedtke, Wolfgang B.

2016-01-01

TRPV4 ion channels represent osmo-mechano-TRP channels with pleiotropic function and wide-spread expression. One of the critical functions of TRPV4 in this spectrum is its involvement in pain and inflammation. However, few small-molecule inhibitors of TRPV4 are available. Here we developed TRPV4-inhibitory molecules based on modifications of a known TRPV4-selective tool-compound, GSK205. We not only increased TRPV4-inhibitory potency, but surprisingly also generated two compounds that potently co-inhibit TRPA1, known to function as chemical sensor of noxious and irritant signaling. We demonstrate TRPV4 inhibition by these compounds in primary cells with known TRPV4 expression - articular chondrocytes and astrocytes. Importantly, our novel compounds attenuate pain behavior in a trigeminal irritant pain model that is known to rely on TRPV4 and TRPA1. Furthermore, our novel dual-channel blocker inhibited inflammation and pain-associated behavior in a model of acute pancreatitis – known to also rely on TRPV4 and TRPA1. Our results illustrate proof of a novel concept inherent in our prototype compounds of a drug that targets two functionally-related TRP channels, and thus can be used to combat isoforms of pain and inflammation in-vivo that involve more than one TRP channel. This approach could provide a novel paradigm for treating other relevant health conditions. PMID:27247148

Evaluation of “Credit Card” Libraries for Inhibition of HIV-1 gp41 Fusogenic Core Formation

PubMed Central

Xu, Yang; Lu, Hong; Kennedy, Jack P.; Yan, Xuxia; McAllister, Laura; Yamamoto, Noboru; Moss, Jason A.; Boldt, Grant E.; Jiang, Shibo; Janda, Kim D.

2008-01-01

Protein-protein interactions are of critical importance in biological systems and small molecule modulators of such protein recognition and intervention processes are of particular interests. To investigate this area of research, we have synthesized small molecule libraries that can disrupt a number of biologically relevant protein-protein interactions. These library members are designed upon planar motifs, appended with a variety of chemical functions, which we have termed as “credit-card” structures. From two of our “credit-card” libraries, a series of molecules were uncovered which act as inhibitors against the HIV-1 gp41 fusogenic 6-helix bundle core formation, viral antigen p24 formation and cell-cell fusion at low micromolar concentrations. From the high-throughput screening assays we utilized, a selective index (SI) value of 4.2 was uncovered for compound 2261, which bodes well for future structure activity investigations and the design of more potent gp41 inhibitors. PMID:16827565

Profiling protein function with small molecule microarrays

PubMed Central

Winssinger, Nicolas; Ficarro, Scott; Schultz, Peter G.; Harris, Jennifer L.

2002-01-01

The regulation of protein function through posttranslational modification, local environment, and protein–protein interaction is critical to cellular function. The ability to analyze on a genome-wide scale protein functional activity rather than changes in protein abundance or structure would provide important new insights into complex biological processes. Herein, we report the application of a spatially addressable small molecule microarray to an activity-based profile of proteases in crude cell lysates. The potential of this small molecule-based profiling technology is demonstrated by the detection of caspase activation upon induction of apoptosis, characterization of the activated caspase, and inhibition of the caspase-executed apoptotic phenotype using the small molecule inhibitor identified in the microarray-based profile. PMID:12167675

Solid-phase assays for small molecule screening using sol-gel entrapped proteins.

PubMed

Lebert, Julie M; Forsberg, Erica M; Brennan, John D

2008-04-01

With compound libraries exceeding one million compounds, the ability to quickly and effectively screen these compounds against relevant pharmaceutical targets has become crucial. Solid-phase assays present several advantages over solution-based methods. For example, a higher degree of miniaturization can be achieved, functional- and affinity-based studies are possible, and a variety of detection methods can be used. Unfortunately, most protein immobilization methods are either too harsh or require recombinant proteins and thus are not amenable to delicate proteins such as kinases and membrane-bound receptors. Sol-gel encapsulation of proteins in an inorganic silica matrix has emerged as a novel solid-phase assay platform. In this minireview, we discuss the development of sol-gel derived protein microarrays and sol-gel based monolithic bioaffinity columns for the high-throughput screening of small molecule libraries and mixtures.

Small Molecule Chemical Probes of MicroRNA Function

PubMed Central

Velagapudi, Sai Pradeep; Vummidi, Balayeshwanth R.; Disney, Matthew D.

2015-01-01

MicroRNAs (miRNAs) are small, non-coding RNAs that control protein expression. Aberrant miRNA expression has been linked to various human diseases, and thus miRNAs have been explored as diagnostic markers and therapeutic targets. Although it is challenging to target RNA with small molecules in general, there have been successful campaigns that have identified small molecule modulators of miRNA function by targeting various pathways. For example, small molecules that modulate transcription and target nuclease processing sites in miRNA precursors have been identified. Herein, we describe challenges in developing chemical probes that target miRNAs and highlight aspects of miRNA cellular biology elucidated by using small molecule chemical probes. We expect that this area will expand dramatically in the near future as strides are made to understand small molecule recognition of RNA from a fundamental perspective. PMID:25500006

Biosynthesis of Lincosamide Antibiotics: Reactions Associated with Degradation and Detoxification Pathways Play a Constructive Role.

PubMed

Zhang, Daozhong; Tang, Zhijun; Liu, Wen

2018-06-19

Natural products typically are small molecules produced by living organisms. These products possess a wide variety of biological activities and thus have historically played a critical role in medicinal chemistry and chemical biology either as chemotherapeutic agents or as useful tools. Natural products are not synthesized for use by human beings; rather, living organisms produce them in response to various biochemical processes and environmental concerns, both internal and external. These processes/concerns are often dynamic and thus motivate the diversification, optimization, and selection of small molecules in line with changes in biological function. Consequently, the interactions between living organisms and their environments serve as an engine that drives coevolution of natural products and their biological functions and ultimately programs the constant theme of small-molecule development in nature based on biosynthesis generality and specificity. Following this theme, we herein review the biosynthesis of lincosamide antibiotics and dissect the process through which nature creates an unusual eight-carbon aminosugar (lincosamide) and then functionalizes this common high-carbon chain-containing sugar core with diverse l-proline derivatives and sulfur appendages to form individual members, including the clinically useful anti-infective agent lincomycin A and its naturally occurring analogues celesticetin and Bu-2545. The biosynthesis of lincosamide antibiotics is unique in that it results from an intersection of anabolic and catabolic chemistry. Many reactions that are usually involved in degradation and detoxification play a constructive role in biosynthetic processes. Formation of the trans-4-propyl-l-proline residue in lincomycin A biosynthesis requires an oxidation-associated degradation-like pathway composed of heme peroxidase-catalyzed ortho-hydroxylation and non-heme 2,3-dioxygenase-catalyzed extradiol cleavage for l-tyrosine processing prior to the building-up process. Mycothiol (MSH) and ergothioneine (EGT), two small-molecule thiols that are known for their redox-relevant roles in protection against various endogenous and exogenous stresses, function through two unusual S-glycosylations to mediate an eight-carbon aminosugar transfer, activation, and modification during the molecular assembly and tailoring processes in lincosamide antibiotic biosynthesis. Related intermediates include an MSH S-conjugate, mercapturic acid, and a thiomethyl product, which are reminiscent of intermediates found in thiol-mediated detoxification metabolism. In these biosynthetic pathways, "old" protein folds can result in "new" enzymatic activity, such as the DinB-2 fold protein for thiol exchange between EGT and MSH, the γ-glutamyltranspeptidase homologue for C-C bond cleavage, and the pyridoxal-5'-phosphate-dependent enzyme for diverse S-functionalization, generating interest in how nature develops remarkably diverse biochemical functions using a limited range of protein scaffolds. These findings highlight what we can learn from natural product biosynthesis, the recognition of its generality and specificity, and the natural theme of the development of bioactive small molecules, which enables the diversification process to advance and expand small-molecule functions.

Small Molecule Inhibitors of AI-2 Signaling in Bacteria: State-of-the-Art and Future Perspectives for Anti-Quorum Sensing Agents

PubMed Central

Guo, Min; Gamby, Sonja; Zheng, Yue; Sintim, Herman O.

2013-01-01

Bacteria respond to different small molecules that are produced by other neighboring bacteria. These molecules, called autoinducers, are classified as intraspecies (i.e., molecules produced and perceived by the same bacterial species) or interspecies (molecules that are produced and sensed between different bacterial species). AI-2 has been proposed as an interspecies autoinducer and has been shown to regulate different bacterial physiology as well as affect virulence factor production and biofilm formation in some bacteria, including bacteria of clinical relevance. Several groups have embarked on the development of small molecules that could be used to perturb AI-2 signaling in bacteria, with the ultimate goal that these molecules could be used to inhibit bacterial virulence and biofilm formation. Additionally, these molecules have the potential to be used in synthetic biology applications whereby these small molecules are used as inputs to switch on and off AI-2 receptors. In this review, we highlight the state-of-the-art in the development of small molecules that perturb AI-2 signaling in bacteria and offer our perspective on the future development and applications of these classes of molecules. PMID:23994835

Developing an Efficient and General Strategy for Immobilization of Small Molecules onto Microarrays Using Isocyanate Chemistry.

PubMed

Zhu, Chenggang; Zhu, Xiangdong; Landry, James P; Cui, Zhaomeng; Li, Quanfu; Dang, Yongjun; Mi, Lan; Zheng, Fengyun; Fei, Yiyan

2016-03-16

Small-molecule microarray (SMM) is an effective platform for identifying lead compounds from large collections of small molecules in drug discovery, and efficient immobilization of molecular compounds is a pre-requisite for the success of such a platform. On an isocyanate functionalized surface, we studied the dependence of immobilization efficiency on chemical residues on molecular compounds, terminal residues on isocyanate functionalized surface, lengths of spacer molecules, and post-printing treatment conditions, and we identified a set of optimized conditions that enable us to immobilize small molecules with significantly improved efficiencies, particularly for those molecules with carboxylic acid residues that are known to have low isocyanate reactivity. We fabricated microarrays of 3375 bioactive compounds on isocyanate functionalized glass slides under these optimized conditions and confirmed that immobilization percentage is over 73%.

Antibody-Mediated Small Molecule Detection Using Programmable DNA-Switches.

PubMed

Rossetti, Marianna; Ippodrino, Rudy; Marini, Bruna; Palleschi, Giuseppe; Porchetta, Alessandro

2018-06-13

The development of rapid, cost-effective, and single-step methods for the detection of small molecules is crucial for improving the quality and efficiency of many applications ranging from life science to environmental analysis. Unfortunately, current methodologies still require multiple complex, time-consuming washing and incubation steps, which limit their applicability. In this work we present a competitive DNA-based platform that makes use of both programmable DNA-switches and antibodies to detect small target molecules. The strategy exploits both the advantages of proximity-based methods and structure-switching DNA-probes. The platform is modular and versatile and it can potentially be applied for the detection of any small target molecule that can be conjugated to a nucleic acid sequence. Here the rational design of programmable DNA-switches is discussed, and the sensitive, rapid, and single-step detection of different environmentally relevant small target molecules is demonstrated.

Small molecule chemical probes of microRNA function.

PubMed

Velagapudi, Sai Pradeep; Vummidi, Balayeshwanth R; Disney, Matthew D

2015-02-01

MicroRNAs (miRNAs) are small, non-coding RNAs that control protein expression. Aberrant miRNA expression has been linked to various human diseases, and thus miRNAs have been explored as diagnostic markers and therapeutic targets. Although it is challenging to target RNA with small molecules in general, there have been successful campaigns that have identified small molecule modulators of miRNA function by targeting various pathways. For example, small molecules that modulate transcription and target nuclease processing sites in miRNA precursors have been identified. Herein, we describe challenges in developing chemical probes that target miRNAs and highlight aspects of miRNA cellular biology elucidated by using small molecule chemical probes. We expect that this area will expand dramatically in the near future as progress is made in understanding small molecule recognition of RNA. Copyright © 2014. Published by Elsevier Ltd.

Salting Constants of Small Organic Molecules in Aerosol-Relevant Salts and Application to Aerosol Formation in the Southeastern United States

NASA Astrophysics Data System (ADS)

Waxman, E.; Carlton, A. M. G.; Ziemann, P. J.; Volkamer, R. M.

2014-12-01

Secondary organic aerosol (SOA) formation from small water-soluble molecules such as glyoxal and methyl glyoxal is a topic of emerging interest. Results from recent field campaigns, e.g. Waxman et al. (2013, GRL) and Knote et al. (2014, ACP), show that these molecules can form significant SOA mass as a result of 'salting-in'. Salting-in happens when a molecule's solubility increases with salt concentration and salting-out is the reverse. Salting effects modify the solubility exponentially with increasing salt concentration, and thus the effective Henry's law constant can strongly modify partitioning, and multiphase chemical reaction rates in aerosol water. Moreover, the solubility in aerosol water cannot easily inferred based on the solubility in cloud water, as the salting effects could change the solubility by a factor of 104 or more. In this work, we have devised and applied a novel experimental setup to measure salting constants using an ion trap mass spectrometer. We focus on small, water soluble molecules like methyl glyoxal and similar compounds and measure salting constants for aerosol-relevant salts including ammonium sulfate, ammonium nitrate, and sodium chloride. The Setschenow salting-constant values are then used to parameterize the effects of salting in CMAQ. We present a series of sensitivity studies of the effects that inorganic aerosols have on the SOA formation from small soluble molecules in the southeastern United States.

Calculations of electric dipole moments and static dipole polarizabilities based on the two-component normalized elimination of the small component method.

PubMed

Yoshizawa, Terutaka; Zou, Wenli; Cremer, Dieter

2016-11-14

The analytical energy gradient and Hessian of the two-component Normalized Elimination of the Small Component (2c-NESC) method with regard to the components of the electric field are derived and used to calculate spin-orbit coupling (SOC) corrected dipole moments and dipole polarizabilities of molecules, which contain elements with high atomic number. Calculated 2c-NESC dipole moments and isotropic polarizabilities agree well with the corresponding four-component-Dirac Hartree-Fock or density functional theory values. SOC corrections for the electrical properties are in general small, but become relevant for the accurate prediction of these properties when the molecules in question contain sixth and/or seventh period elements (e.g., the SO effect for At 2 is about 10% of the 2c-NESC polarizability). The 2c-NESC changes in the electric molecular properties are rationalized in terms of spin-orbit splitting and SOC-induced mixing of frontier orbitals with the same j = l + s quantum numbers.

Calculations of electric dipole moments and static dipole polarizabilities based on the two-component normalized elimination of the small component method

NASA Astrophysics Data System (ADS)

Yoshizawa, Terutaka; Zou, Wenli; Cremer, Dieter

2016-11-01

The analytical energy gradient and Hessian of the two-component Normalized Elimination of the Small Component (2c-NESC) method with regard to the components of the electric field are derived and used to calculate spin-orbit coupling (SOC) corrected dipole moments and dipole polarizabilities of molecules, which contain elements with high atomic number. Calculated 2c-NESC dipole moments and isotropic polarizabilities agree well with the corresponding four-component-Dirac Hartree-Fock or density functional theory values. SOC corrections for the electrical properties are in general small, but become relevant for the accurate prediction of these properties when the molecules in question contain sixth and/or seventh period elements (e.g., the SO effect for At2 is about 10% of the 2c-NESC polarizability). The 2c-NESC changes in the electric molecular properties are rationalized in terms of spin-orbit splitting and SOC-induced mixing of frontier orbitals with the same j = l + s quantum numbers.

Importance of Being Nernst: Synaptic Activity and Functional Relevance in Stem Cell-derived Neurons

DTIC Science & Technology

2015-07-26

neurodevelopmental stages. In some cases these factors can be controlled very precisely, such as by the addition of small molecules to promote exit from...neurogenesis[43]. These include markers of the different stages of neurodevelopment , starting from a stem cell state and expressing characteristics of a...neuroligin-3 mutations associated with autism cause post-synaptic dysfunction in iNs when co-cultured with primary neurons[163]. The iN field is still

The perception of strigolactones in vascular plants.

PubMed

Lumba, Shelley; Holbrook-Smith, Duncan; McCourt, Peter

2017-05-17

Small-molecule hormones play central roles in plant development, ranging from cellular differentiation and organ formation to developmental response instruction in changing environments. A recently discovered collection of related small molecules collectively called strigolactones are of particular interest, as these hormones also function as ecological communicators between plants and fungi and between parasitic plants and their hosts. Advances from model plant systems have begun to unravel how, as a hormone, strigolactone is perceived and transduced. In this Review, we summarize this information and examine how understanding strigolactone hormone signaling is leading to insights into parasitic plant infections. We specifically focus on how the development of chemical probes can be used in combination with model plant systems to dissect strigolactone's perception in the parasitic plant Striga hermonthica. This information is particularly relevant since Striga is considered one of the largest impediments to food security in sub-Saharan Africa.

Rational design of chemical genetic probes of RNA function and lead therapeutics targeting repeating transcripts.

PubMed

Disney, Matthew D

2013-12-01

RNA is an important yet vastly underexploited target for small molecule chemical probes or lead therapeutics. Small molecules have been used successfully to modulate the function of the bacterial ribosome, viral RNAs and riboswitches. These RNAs are either highly expressed or can be targeted using substrate mimicry, a mainstay in the design of enzyme inhibitors. However, most cellular RNAs are neither highly expressed nor have a lead small molecule inhibitor, a significant challenge for drug discovery efforts. Herein, I describe the design of small molecules targeting expanded repeating transcripts that cause myotonic muscular dystrophy (DM). These test cases illustrate the challenges of designing small molecules that target RNA and the advantages of targeting repeating transcripts. Lastly, I discuss how small molecules might be more advantageous than oligonucleotides for targeting RNA. Copyright © 2013 Elsevier Ltd. All rights reserved.

SPLINTS: small-molecule protein ligand interface stabilizers.

PubMed

Fischer, Eric S; Park, Eunyoung; Eck, Michael J; Thomä, Nicolas H

2016-04-01

Regulatory protein-protein interactions are ubiquitous in biology, and small molecule protein-protein interaction inhibitors are an important focus in drug discovery. Remarkably little attention has been given to the opposite strategy-stabilization of protein-protein interactions, despite the fact that several well-known therapeutics act through this mechanism. From a structural perspective, we consider representative examples of small molecules that induce or stabilize the association of protein domains to inhibit, or alter, signaling for nuclear hormone, GTPase, kinase, phosphatase, and ubiquitin ligase pathways. These SPLINTS (small-molecule protein ligand interface stabilizers) drive interactions that are in some cases physiologically relevant, and in others entirely adventitious. The diverse structural mechanisms employed suggest approaches for a broader and systematic search for such compounds in drug discovery. Copyright © 2016 Elsevier Ltd. All rights reserved.

High-Resolution Photoionization, Photoelectron and Photodissociation Studies. Determination of Accurate Energetic and Spectroscopic Database for Combustion Radicals and Molecules

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

Ng, Cheuk-Yiu

2016-04-25

The main goal of this research program was to obtain accurate thermochemical and spectroscopic data, such as ionization energies (IEs), 0 K bond dissociation energies, 0 K heats of formation, and spectroscopic constants for radicals and molecules and their ions of relevance to combustion chemistry. Two unique, generally applicable vacuum ultraviolet (VUV) laser photoion-photoelectron apparatuses have been developed in our group, which have used for high-resolution photoionization, photoelectron, and photodissociation studies for many small molecules of combustion relevance.

Precision Electrophile Tagging in Caenorhabditis elegans.

PubMed

Long, Marcus J C; Urul, Daniel A; Chawla, Shivansh; Lin, Hong-Yu; Zhao, Yi; Haegele, Joseph A; Wang, Yiran; Aye, Yimon

2018-01-16

Adduction of an electrophile to privileged sensor proteins and the resulting phenotypically dominant responses are increasingly appreciated as being essential for metazoan health. Functional similarities between the biological electrophiles and electrophilic pharmacophores commonly found in covalent drugs further fortify the translational relevance of these small-molecule signals. Genetically encodable or small-molecule-based fluorescent reporters and redox proteomics have revolutionized the observation and profiling of cellular redox states and electrophile-sensor proteins, respectively. However, precision mapping between specific redox-modified targets and specific responses has only recently begun to be addressed, and systems tractable to both genetic manipulation and on-target redox signaling in vivo remain largely limited. Here we engineer transgenic Caenorhabditis elegans expressing functional HaloTagged fusion proteins and use this system to develop a generalizable light-controlled approach to tagging a prototypical electrophile-sensor protein with native electrophiles in vivo. The method circumvents issues associated with low uptake/distribution and toxicity/promiscuity. Given the validated success of C. elegans in aging studies, this optimized platform offers a new lens with which to scrutinize how on-target electrophile signaling influences redox-dependent life span regulation.

Precision Electrophile Tagging in Caenorhabditis elegans

PubMed Central

2017-01-01

Adduction of an electrophile to privileged sensor proteins and the resulting phenotypically dominant responses are increasingly appreciated as being essential for metazoan health. Functional similarities between the biological electrophiles and electrophilic pharmacophores commonly found in covalent drugs further fortify the translational relevance of these small-molecule signals. Genetically encodable or small-molecule-based fluorescent reporters and redox proteomics have revolutionized the observation and profiling of cellular redox states and electrophile–sensor proteins, respectively. However, precision mapping between specific redox-modified targets and specific responses has only recently begun to be addressed, and systems tractable to both genetic manipulation and on-target redox signaling in vivo remain largely limited. Here we engineer transgenic Caenorhabditis elegans expressing functional HaloTagged fusion proteins and use this system to develop a generalizable light-controlled approach to tagging a prototypical electrophile–sensor protein with native electrophiles in vivo. The method circumvents issues associated with low uptake/distribution and toxicity/promiscuity. Given the validated success of C. elegans in aging studies, this optimized platform offers a new lens with which to scrutinize how on-target electrophile signaling influences redox-dependent life span regulation. PMID:28857552

Methods to enable the design of bioactive small molecules targeting RNA

PubMed Central

Disney, Matthew D.; Yildirim, Ilyas; Childs-Disney, Jessica L.

2014-01-01

RNA is an immensely important target for small molecule therapeutics or chemical probes of function. However, methods that identify, annotate, and optimize RNA-small molecule interactions that could enable the design of compounds that modulate RNA function are in their infancies. This review describes recent approaches that have been developed to understand and optimize RNA motif-small molecule interactions, including Structure-Activity Relationships Through Sequencing (StARTS), quantitative structure-activity relationships (QSAR), chemical similarity searching, structure-based design and docking, and molecular dynamics (MD) simulations. Case studies described include the design of small molecules targeting RNA expansions, the bacterial A-site, viral RNAs, and telomerase RNA. These approaches can be combined to afford a synergistic method to exploit the myriad of RNA targets in the transcriptome. PMID:24357181

Methods to enable the design of bioactive small molecules targeting RNA.

PubMed

Disney, Matthew D; Yildirim, Ilyas; Childs-Disney, Jessica L

2014-02-21

RNA is an immensely important target for small molecule therapeutics or chemical probes of function. However, methods that identify, annotate, and optimize RNA-small molecule interactions that could enable the design of compounds that modulate RNA function are in their infancies. This review describes recent approaches that have been developed to understand and optimize RNA motif-small molecule interactions, including structure-activity relationships through sequencing (StARTS), quantitative structure-activity relationships (QSAR), chemical similarity searching, structure-based design and docking, and molecular dynamics (MD) simulations. Case studies described include the design of small molecules targeting RNA expansions, the bacterial A-site, viral RNAs, and telomerase RNA. These approaches can be combined to afford a synergistic method to exploit the myriad of RNA targets in the transcriptome.

Tuning dissociation using isoelectronically doped graphene and hexagonal boron nitride: Water and other small molecules

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

Al-Hamdani, Yasmine S.; Department of Chemistry, University College London, 20 Gordon Street, London WC1H 0AJ; Alfè, Dario

2016-04-21

Novel uses for 2-dimensional materials like graphene and hexagonal boron nitride (h-BN) are being frequently discovered especially for membrane and catalysis applications. Still however, a great deal remains to be understood about the interaction of environmentally and industrially relevant molecules such as water with these materials. Taking inspiration from advances in hybridising graphene and h-BN, we explore using density functional theory, the dissociation of water, hydrogen, methane, and methanol on graphene, h-BN, and their isoelectronic doped counterparts: BN doped graphene and C doped h-BN. We find that doped surfaces are considerably more reactive than their pristine counterparts and by comparingmore » the reactivity of several small molecules, we develop a general framework for dissociative adsorption. From this a particularly attractive consequence of isoelectronic doping emerges: substrates can be doped to enhance their reactivity specifically towards either polar or non-polar adsorbates. As such, these substrates are potentially viable candidates for selective catalysts and membranes, with the implication that a range of tuneable materials can be designed.« less

Tuning dissociation using isoelectronically doped graphene and hexagonal boron nitride: Water and other small molecules

NASA Astrophysics Data System (ADS)

Al-Hamdani, Yasmine S.; Alfè, Dario; von Lilienfeld, O. Anatole; Michaelides, Angelos

2016-04-01

Novel uses for 2-dimensional materials like graphene and hexagonal boron nitride (h-BN) are being frequently discovered especially for membrane and catalysis applications. Still however, a great deal remains to be understood about the interaction of environmentally and industrially relevant molecules such as water with these materials. Taking inspiration from advances in hybridising graphene and h-BN, we explore using density functional theory, the dissociation of water, hydrogen, methane, and methanol on graphene, h-BN, and their isoelectronic doped counterparts: BN doped graphene and C doped h-BN. We find that doped surfaces are considerably more reactive than their pristine counterparts and by comparing the reactivity of several small molecules, we develop a general framework for dissociative adsorption. From this a particularly attractive consequence of isoelectronic doping emerges: substrates can be doped to enhance their reactivity specifically towards either polar or non-polar adsorbates. As such, these substrates are potentially viable candidates for selective catalysts and membranes, with the implication that a range of tuneable materials can be designed.

Small-Molecule-Directed Hepatocyte-Like Cell Differentiation of Human Pluripotent Stem Cells.

PubMed

Mathapati, Santosh; Siller, Richard; Impellizzeri, Agata A R; Lycke, Max; Vegheim, Karianne; Almaas, Runar; Sullivan, Gareth J

2016-08-17

Hepatocyte-like cells (HLCs) generated in vitro from human pluripotent stem cells (hPSCs) provide an invaluable resource for basic research, regenerative medicine, drug screening, toxicology, and modeling of liver disease and development. This unit describes a small-molecule-driven protocol for in vitro differentiation of hPSCs into HLCs without the use of growth factors. hPSCs are coaxed through a developmentally relevant route via the primitive streak to definitive endoderm (DE) using the small molecule CHIR99021 (a Wnt agonist), replacing the conventional growth factors Wnt3A and activin A. The small-molecule-derived DE is then differentiated to hepatoblast-like cells in the presence of dimethyl sulfoxide. The resulting hepatoblasts are then differentiated to HLCs with N-hexanoic-Tyr, Ile-6 aminohexanoic amide (Dihexa, a hepatocyte growth factor agonist) and dexamethasone. The protocol provides an efficient and reproducible procedure for differentiation of hPSCs into HLCs utilizing small molecules. © 2016 by John Wiley & Sons, Inc. Copyright © 2016 John Wiley & Sons, Inc.

Mathematical Modeling of E6-p53 interactions in Cervical Cancer

PubMed

Khattak, Faryal; Haseeb, Muhammad; Fazal, Sahar; Bhatti, A I; Ullah, Mukhtar

2017-04-01

Background: Cervical cancer is the third most common cancer in women throughout the world. The human papillomavirus (HPV) E6 viral protein plays an essential role in proteasomal degradation of the cancer suppressant protein p53. As a result, p53 negative regulation and apoptosis relevant activities are abrogated, facilitating development of cervical cancer. Methods: A mathematical model of E6-p53 interactions was developed using mathematical laws. In-silico simulations were carried out on CellDesigner and as a test case the small molecule drug RITA was considered for its ability to rescue the functions of tumor suppressor p53 by inhibiting E6 mediated proteasomal degradation. Results: Using a computational model we scrutinized how p53 responds to RITA, and chemical reactions of this small molecule drug were incorporated to perceive the full effects. The evolved strategy allowed the p53 response and rescue of its tumor suppressor function to be delineated, RITA being found to block p53 interactions with E6 associated proteins. Conclusion: We could develop a model of E6-p53 interactions with incorporation of actions of the small molecule drug RITA. Suppression of E6 associated proteins by RITA induces accumulation of tumor suppressant p53. Using CellDesigner to encode the model ensured that it can be easily modified and extended as more data become available. This strategy should play an effective role in the development of therapies against cancer. Creative Commons Attribution License

Mathematical Modeling of E6-p53 interactions in Cervical Cancer

PubMed Central

Khattak, Faryal; Haseeb, Muhammad; Fazal, Sahar; Bhatti, AI; Ullah, Mukhtar

2017-01-01

Background: Cervical cancer is the third most common cancer in women throughout the world. The human papillomavirus (HPV) E6 viral protein plays an essential role in proteasomal degradation of the cancer suppressant protein p53. As a result, p53 negative regulation and apoptosis relevant activities are abrogated, facilitating development of cervical cancer. Methods: A mathematical model of E6-p53 interactions was developed using mathematical laws. In-silico simulations were carried out on CellDesigner and as a test case the small molecule drug RITA was considered for its ability to rescue the functions of tumor suppressor p53 by inhibiting E6 mediated proteasomal degradation. Results: Using a computational model we scrutinized how p53 responds to RITA, and chemical reactions of this small molecule drug were incorporated to perceive the full effects. The evolved strategy allowed the p53 response and rescue of its tumor suppressor function to be delineated, RITA being found to block p53 interactions with E6 associated proteins. Conclusion: We could develop a model of E6-p53 interactions with incorporation of actions of the small molecule drug RITA. Suppression of E6 associated proteins by RITA induces accumulation of tumor suppressant p53. Using CellDesigner to encode the model ensured that it can be easily modified and extended as more data become available. This strategy should play an effective role in the development of therapies against cancer. PMID:28547941

Inforna 2.0: A Platform for the Sequence-Based Design of Small Molecules Targeting Structured RNAs.

PubMed

Disney, Matthew D; Winkelsas, Audrey M; Velagapudi, Sai Pradeep; Southern, Mark; Fallahi, Mohammad; Childs-Disney, Jessica L

2016-06-17

The development of small molecules that target RNA is challenging yet, if successful, could advance the development of chemical probes to study RNA function or precision therapeutics to treat RNA-mediated disease. Previously, we described Inforna, an approach that can mine motifs (secondary structures) within target RNAs, which is deduced from the RNA sequence, and compare them to a database of known RNA motif-small molecule binding partners. Output generated by Inforna includes the motif found in both the database and the desired RNA target, lead small molecules for that target, and other related meta-data. Lead small molecules can then be tested for binding and affecting cellular (dys)function. Herein, we describe Inforna 2.0, which incorporates all known RNA motif-small molecule binding partners reported in the scientific literature, a chemical similarity searching feature, and an improved user interface and is freely available via an online web server. By incorporation of interactions identified by other laboratories, the database has been doubled, containing 1936 RNA motif-small molecule interactions, including 244 unique small molecules and 1331 motifs. Interestingly, chemotype analysis of the compounds that bind RNA in the database reveals features in small molecule chemotypes that are privileged for binding. Further, this updated database expanded the number of cellular RNAs to which lead compounds can be identified.

Recent Developments in β-Cell Differentiation of Pluripotent Stem Cells Induced by Small and Large Molecules

PubMed Central

Kumar, S. Suresh; Alarfaj, Abdullah A.; Munusamy, Murugan A.; Singh, A. J. A. Ranjith; Peng, I-Chia; Priya, Sivan Padma; Hamat, Rukman Awang; Higuchi, Akon

2014-01-01

Human pluripotent stem cells, including human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs), hold promise as novel therapeutic tools for diabetes treatment because of their self-renewal capacity and ability to differentiate into beta (β)-cells. Small and large molecules play important roles in each stage of β-cell differentiation from both hESCs and hiPSCs. The small and large molecules that are described in this review have significantly advanced efforts to cure diabetic disease. Lately, effective protocols have been implemented to induce hESCs and human mesenchymal stem cells (hMSCs) to differentiate into functional β-cells. Several small molecules, proteins, and growth factors promote pancreatic differentiation from hESCs and hMSCs. These small molecules (e.g., cyclopamine, wortmannin, retinoic acid, and sodium butyrate) and large molecules (e.g. activin A, betacellulin, bone morphogentic protein (BMP4), epidermal growth factor (EGF), fibroblast growth factor (FGF), keratinocyte growth factor (KGF), hepatocyte growth factor (HGF), noggin, transforming growth factor (TGF-α), and WNT3A) are thought to contribute from the initial stages of definitive endoderm formation to the final stages of maturation of functional endocrine cells. We discuss the importance of such small and large molecules in uniquely optimized protocols of β-cell differentiation from stem cells. A global understanding of various small and large molecules and their functions will help to establish an efficient protocol for β-cell differentiation. PMID:25526563

Cheminformatics-aided discovery of small-molecule Protein-Protein Interaction (PPI) dual inhibitors of Tumor Necrosis Factor (TNF) and Receptor Activator of NF-κB Ligand (RANKL).

PubMed

Melagraki, Georgia; Ntougkos, Evangelos; Rinotas, Vagelis; Papaneophytou, Christos; Leonis, Georgios; Mavromoustakos, Thomas; Kontopidis, George; Douni, Eleni; Afantitis, Antreas; Kollias, George

2017-04-01

We present an in silico drug discovery pipeline developed and applied for the identification and virtual screening of small-molecule Protein-Protein Interaction (PPI) compounds that act as dual inhibitors of TNF and RANKL through the trimerization interface. The cheminformatics part of the pipeline was developed by combining structure-based with ligand-based modeling using the largest available set of known TNF inhibitors in the literature (2481 small molecules). To facilitate virtual screening, the consensus predictive model was made freely available at: http://enalos.insilicotox.com/TNFPubChem/. We thus generated a priority list of nine small molecules as candidates for direct TNF function inhibition. In vitro evaluation of these compounds led to the selection of two small molecules that act as potent direct inhibitors of TNF function, with IC50 values comparable to those of a previously-described direct inhibitor (SPD304), but with significantly reduced toxicity. These molecules were also identified as RANKL inhibitors and validated in vitro with respect to this second functionality. Direct binding of the two compounds was confirmed both for TNF and RANKL, as well as their ability to inhibit the biologically-active trimer forms. Molecular dynamics calculations were also carried out for the two small molecules in each protein to offer additional insight into the interactions that govern TNF and RANKL complex formation. To our knowledge, these compounds, namely T8 and T23, constitute the second and third published examples of dual small-molecule direct function inhibitors of TNF and RANKL, and could serve as lead compounds for the development of novel treatments for inflammatory and autoimmune diseases.

Interferometric Scattering Microscopy for the Study of Molecular Motors

PubMed Central

Andrecka, J.; Takagi, Y.; Mickolajczyk, K.J.; Lippert, L.G.; Sellers, J.R.; Hancock, W.O.; Goldman, Y.E.; Kukura, P.

2016-01-01

Our understanding of molecular motor function has been greatly improved by the development of imaging modalities, which enable real-time observation of their motion at the single-molecule level. Here, we describe the use of a new method, interferometric scattering microscopy, for the investigation of motor protein dynamics by attaching and tracking the motion of metallic nanoparticle labels as small as 20 nm diameter. Using myosin-5, kinesin-1, and dynein as examples, we describe the basic assays, labeling strategies, and principles of data analysis. Our approach is relevant not only for motor protein dynamics but also provides a general tool for single-particle tracking with high spatiotemporal precision, which overcomes the limitations of single-molecule fluorescence methods. PMID:27793291

Small-molecule control of protein function through Staudinger reduction

NASA Astrophysics Data System (ADS)

Luo, Ji; Liu, Qingyang; Morihiro, Kunihiko; Deiters, Alexander

2016-11-01

Using small molecules to control the function of proteins in live cells with complete specificity is highly desirable, but challenging. Here we report a small-molecule switch that can be used to control protein activity. The approach uses a phosphine-mediated Staudinger reduction to activate protein function. Genetic encoding of an ortho-azidobenzyloxycarbonyl amino acid using a pyrrolysyl transfer RNA synthetase/tRNACUA pair in mammalian cells enables the site-specific introduction of a small-molecule-removable protecting group into the protein of interest. Strategic placement of this group renders the protein inactive until deprotection through a bioorthogonal Staudinger reduction delivers the active wild-type protein. This developed methodology was applied to the conditional control of several cellular processes, including bioluminescence (luciferase), fluorescence (enhanced green fluorescent protein), protein translocation (nuclear localization sequence), DNA recombination (Cre) and gene editing (Cas9).

[Innovative application of small molecules to influence -pathogenicity of dental plaque].

PubMed

Janus, M M; Volgenant, C M C; Krom, B P

2018-05-01

Current preventive measures against infectious oral diseases are mainly focussed on plaque removal and promoting a healthy lifestyle. This in vitro study investigated a third preventive method: maintaining healthy dental plaque with the use of small molecules. As a model of dental plaque, in vitro biofilms were cultivated under conditions that induce pathogenic characteristics. The effect of erythritol and other small molecules on the pathogenic characteristics and bacterial composition of the biofilm was evaluated. The artificial sweetener erythritol and the molecule 3-Oxo-N-(2-oxycyclohexyl)dodecanamide (3-Oxo-N) had no clinically relevant effect on total biofilm formation. Erythritol did, however, lower the gingivitis related protease activity of the biofilm, while 3-Oxo-N blocked the caries related lactic acid accumulation. Furthermore, both substances ensured the biofilm maintained a young, non-pathogenic microbial composition. This shows it is possible to influence the dental plaque in a positive manner in vitro with the help of small molecules. Further research is necessary before this manipulation of dental plaque can be applied.

Role of p21-activated kinases in cardiovascular development and function.

PubMed

Kelly, Mollie L; Astsaturov, Artyom; Chernoff, Jonathan

2013-11-01

p21-activated kinases (Paks) are a group of six serine/threonine kinases (Pak1-6) that are involved in a variety of biological processes. Recently, Paks, more specifically Pak1, -2, and -4, have been shown to play important roles in cardiovascular development and function in a range of model organisms including zebrafish and mice. These functions include proper morphogenesis and conductance of the heart, cardiac contractility, and development and integrity of the vasculature. The mechanisms underlying these effects are not fully known, but they likely differ among the various Pak isoforms and include both kinase-dependent and -independent functions. In this review, we discuss aspects of Pak function relevant to cardiovascular biology as well as potential therapeutic implications of small-molecule Pak inhibitors in cardiovascular disease.

Exploration of Novel Chemical Space: Synthesis and in vitro Evaluation of N-Functionalized Tertiary Sulfonimidamides.

PubMed

Izzo, Flavia; Schäfer, Martina; Lienau, Philip; Ganzer, Ursula; Stockman, Robert; Lücking, Ulrich

2018-05-04

An unprecedented set of structurally diverse sulfonimidamides (47 compounds) has been prepared by various N-functionalization reactions of tertiary =NH sulfonimidamide 2 aa. These N-functionalization reactions of model compound 2 aa include arylation, alkylation, trifluoromethylation, cyanation, sulfonylation, alkoxycarbonylation (carbamate formation) and aminocarbonylation (urea formation). Small molecule X-ray analyses of selected N-functionalized products are reported. To gain further insight into the properties of sulfonimidamides relevant to medicinal chemistry, a variety of structurally diverse reaction products were tested in selected in vitro assays. The described N-functionalization reactions provide a short and efficient approach to structurally diverse sulfonimidamides which have been the subject of recent, growing interest in the life sciences. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Small functional groups for controlled differentiation of hydrogel-encapsulated human mesenchymal stem cells

NASA Astrophysics Data System (ADS)

Benoit, Danielle S. W.; Schwartz, Michael P.; Durney, Andrew R.; Anseth, Kristi S.

2008-10-01

Cell-matrix interactions have critical roles in regeneration, development and disease. The work presented here demonstrates that encapsulated human mesenchymal stem cells (hMSCs) can be induced to differentiate down osteogenic and adipogenic pathways by controlling their three-dimensional environment using tethered small-molecule chemical functional groups. Hydrogels were formed using sufficiently low concentrations of tether molecules to maintain constant physical characteristics, encapsulation of hMSCs in three dimensions prevented changes in cell morphology, and hMSCs were shown to differentiate in normal growth media, indicating that the small-molecule functional groups induced differentiation. To our knowledge, this is the first example where synthetic matrices are shown to control induction of multiple hMSC lineages purely through interactions with small-molecule chemical functional groups tethered to the hydrogel material. Strategies using simple chemistry to control complex biological processes would be particularly powerful as they could make production of therapeutic materials simpler, cheaper and more easily controlled.

Studying the Immunomodulatory Effects of Small Molecule Ras-Inhibitors in Animal Models of Rheumatoid Arthritis

DTIC Science & Technology

2015-10-01

Studying the Immunomodulatory Effects of Small Molecule Ras-Inhibitors in Animal Models of Rheumatoid Arthritis 5a. CONTRACT NUMBER 5b. GRANT NUMBER...Importantly, T cells from patients with Rheumatoid Arthritis (RA) display augmented activation of the Ras/Raf/MEK/ERK1/2 signaling pathway, and...analysis. The proposed project is highly relevant to the FY13 PRMRP topic area of Rheumatoid Arthritis (RA). The short-term impact of our research will

Group specific internal standard technology (GSIST) for simultaneous identification and quantification of small molecules

DOEpatents

Adamec, Jiri; Yang, Wen-Chu; Regnier, Fred E

2014-01-14

Reagents and methods are provided that permit simultaneous analysis of multiple diverse small molecule analytes present in a complex mixture. Samples are labeled with chemically identical but isotopically distince forms of the labeling reagent, and analyzed using mass spectrometry. A single reagent simultaneously derivatizes multiple small molecule analytes having different reactive functional groups.

Free-standing few-layered graphene oxide films: selective, steady and lasting permeation of organic molecules with adjustable speeds

NASA Astrophysics Data System (ADS)

Huang, Tao; An, Qi; Luan, Xinglong; Zhang, Qian; Zhang, Yihe

2016-01-01

A variety of small molecules with diameters around 1 nm possess a range of functions, such as antibiotic, antimicrobic, anticoagulant, pesticidal and chemotherapy effects, making these molecules especially useful in various applications ranging from medical treatment to environmental microbiological control. However, the long-term steady delivery (release or permeation) of these small molecules with adjustable and controllable speeds has remained an especially challenging task. In this study, we prepared covalently cross-linked free-standing few-layered GO films using a layer-by-layer technique in combination with photochemical cross-linkages, and achieved a controlled release of positively charged, negatively charged, and zwitterionic small molecules with adjustable and controllable speeds. The steady delivery of the small molecule lasted up to 9 days. Other functionalities, such as graphene-enhanced Raman spectra and electrochemical properties that could also be integrated or employed in delivery systems, were also studied for our films. We expect the special molecular delivery properties of our films to lead to new possibilities in drug/fertilizer delivery and environmental microbiological control applications.A variety of small molecules with diameters around 1 nm possess a range of functions, such as antibiotic, antimicrobic, anticoagulant, pesticidal and chemotherapy effects, making these molecules especially useful in various applications ranging from medical treatment to environmental microbiological control. However, the long-term steady delivery (release or permeation) of these small molecules with adjustable and controllable speeds has remained an especially challenging task. In this study, we prepared covalently cross-linked free-standing few-layered GO films using a layer-by-layer technique in combination with photochemical cross-linkages, and achieved a controlled release of positively charged, negatively charged, and zwitterionic small molecules with adjustable and controllable speeds. The steady delivery of the small molecule lasted up to 9 days. Other functionalities, such as graphene-enhanced Raman spectra and electrochemical properties that could also be integrated or employed in delivery systems, were also studied for our films. We expect the special molecular delivery properties of our films to lead to new possibilities in drug/fertilizer delivery and environmental microbiological control applications. Electronic supplementary information (ESI) available: AFM images of GO and GO films, UV-vis spectra of delayed release, and permeation fidelities. See DOI: 10.1039/c5nr08129g

Engineered kinesin motor proteins amenable to small-molecule inhibition

PubMed Central

Engelke, Martin F.; Winding, Michael; Yue, Yang; Shastry, Shankar; Teloni, Federico; Reddy, Sanjay; Blasius, T. Lynne; Soppina, Pushpanjali; Hancock, William O.; Gelfand, Vladimir I.; Verhey, Kristen J.

2016-01-01

The human genome encodes 45 kinesin motor proteins that drive cell division, cell motility, intracellular trafficking and ciliary function. Determining the cellular function of each kinesin would benefit from specific small-molecule inhibitors. However, screens have yielded only a few specific inhibitors. Here we present a novel chemical-genetic approach to engineer kinesin motors that can carry out the function of the wild-type motor yet can also be efficiently inhibited by small, cell-permeable molecules. Using kinesin-1 as a prototype, we develop two independent strategies to generate inhibitable motors, and characterize the resulting inhibition in single-molecule assays and in cells. We further apply these two strategies to create analogously inhibitable kinesin-3 motors. These inhibitable motors will be of great utility to study the functions of specific kinesins in a dynamic manner in cells and animals. Furthermore, these strategies can be used to generate inhibitable versions of any motor protein of interest. PMID:27045608

Discovery of Small Molecules that Inhibit the Disordered Protein, p27Kip1

PubMed Central

Iconaru, Luigi I.; Ban, David; Bharatham, Kavitha; Ramanathan, Arvind; Zhang, Weixing; Shelat, Anang A.; Zuo, Jian; Kriwacki, Richard W.

2015-01-01

Disordered proteins are highly prevalent in biological systems, they control myriad signaling and regulatory processes, and their levels and/or cellular localization are often altered in human disease. In contrast to folded proteins, disordered proteins, due to conformational heterogeneity and dynamics, are not considered viable drug targets. We challenged this paradigm by identifying through NMR-based screening small molecules that bound specifically, albeit weakly, to the disordered cell cycle regulator, p27Kip1 (p27). Two groups of molecules bound to sites created by transient clusters of aromatic residues within p27. Conserved chemical features within these two groups of small molecules exhibited complementarity to their binding sites within p27, establishing structure-activity relationships for small molecule:disordered protein interactions. Finally, one compound counteracted the Cdk2/cyclin A inhibitory function of p27 in vitro, providing proof-of-principle that small molecules can inhibit the function of a disordered protein (p27) through sequestration in a conformation incapable of folding and binding to a natural regulatory target (Cdk2/cyclin A). PMID:26507530

Discovery of Small Molecules that Inhibit the Disordered Protein, p27 Kip1

DOE PAGES

Iconaru, Luigi I.; Ban, David; Bharatham, Kavitha; ...

2015-10-28

In disordered proteins we see that they are highly prevalent in biological systems. They control myriad signaling and regulatory processes, and their levels and/or cellular localization are often altered in human disease. In contrast to folded proteins, disordered proteins, due to conformational heterogeneity and dynamics, are not considered viable drug targets. We challenged this paradigm by identifying through NMR-based screening small molecules that bound specifically, albeit weakly, to the disordered cell cycle regulator, p27 Kip1 (p27). Moreover, two groups of molecules bound to sites created by transient clusters of aromatic residues within p27. Conserved chemical features within these two groupsmore » of small molecules exhibited complementarity to their binding sites within p27, establishing structure-activity relationships for small molecule: disordered protein interactions. Finally, one compound counteracted the Cdk2/cyclin A inhibitory function of p27 in vitro, providing proof-of- principle that small molecules can inhibit the function of a disordered protein (p27) through sequestration in a conformation incapable of folding and binding to a natural regulatory target (Cdk2/cyclin A).« less

Use of mRNA expression signatures to discover small molecule inhibitors of skeletal muscle atrophy

PubMed Central

Adams, Christopher M.; Ebert, Scott M.; Dyle, Michael C.

2017-01-01

Purpose of review Here, we discuss a recently developed experimental strategy for discovering small molecules with potential to prevent and treat skeletal muscle atrophy. Recent findings Muscle atrophy involves and requires widespread changes in skeletal muscle gene expression, which generate complex but measurable patterns of positive and negative changes in skeletal muscle mRNA levels (a.k.a. mRNA expression signatures of muscle atrophy). Many bioactive small molecules generate their own characteristic mRNA expression signatures, and by identifying small molecules whose signatures approximate mirror images of muscle atrophy signatures, one may identify small molecules with potential to prevent and/or reverse muscle atrophy. Unlike a conventional drug discovery approach, this strategy does not rely on a predefined molecular target but rather exploits the complexity of muscle atrophy to identify small molecules that counter the entire spectrum of pathological changes in atrophic muscle. We discuss how this strategy has been used to identify two natural compounds, ursolic acid and tomatidine, that reduce muscle atrophy and improve skeletal muscle function. Summary Discovery strategies based on mRNA expression signatures can elucidate new approaches for preserving and restoring muscle mass and function. PMID:25807353

Use of mRNA expression signatures to discover small molecule inhibitors of skeletal muscle atrophy.

PubMed

Adams, Christopher M; Ebert, Scott M; Dyle, Michael C

2015-05-01

Here, we discuss a recently developed experimental strategy for discovering small molecules with potential to prevent and treat skeletal muscle atrophy. Muscle atrophy involves and requires widespread changes in skeletal muscle gene expression, which generate complex but measurable patterns of positive and negative changes in skeletal muscle mRNA levels (a.k.a. mRNA expression signatures of muscle atrophy). Many bioactive small molecules generate their own characteristic mRNA expression signatures, and by identifying small molecules whose signatures approximate mirror images of muscle atrophy signatures, one may identify small molecules with potential to prevent and/or reverse muscle atrophy. Unlike a conventional drug discovery approach, this strategy does not rely on a predefined molecular target but rather exploits the complexity of muscle atrophy to identify small molecules that counter the entire spectrum of pathological changes in atrophic muscle. We discuss how this strategy has been used to identify two natural compounds, ursolic acid and tomatidine, that reduce muscle atrophy and improve skeletal muscle function. Discovery strategies based on mRNA expression signatures can elucidate new approaches for preserving and restoring muscle mass and function.

In pursuit of small molecule chemistry for calcium-permeable non-selective TRPC channels – mirage or pot of gold?

PubMed Central

Bon, Robin S; Beech, David J

2013-01-01

The primary purpose of this review is to address the progress towards small molecule modulators of human Transient Receptor Potential Canonical proteins (TRPC1, TRPC3, TRPC4, TRPC5, TRPC6 and TRPC7). These proteins generate channels for calcium and sodium ion entry. They are relevant to many mammalian cell types including acinar gland cells, adipocytes, astrocytes, cardiac myocytes, cochlea hair cells, endothelial cells, epithelial cells, fibroblasts, hepatocytes, keratinocytes, leukocytes, mast cells, mesangial cells, neurones, osteoblasts, osteoclasts, platelets, podocytes, smooth muscle cells, skeletal muscle and tumour cells. There are broad-ranging positive roles of the channels in cell adhesion, migration, proliferation, survival and turning, vascular permeability, hypertrophy, wound-healing, hypo-adiponectinaemia, angiogenesis, neointimal hyperplasia, oedema, thrombosis, muscle endurance, lung hyper-responsiveness, glomerular filtration, gastrointestinal motility, pancreatitis, seizure, innate fear, motor coordination, saliva secretion, mast cell degranulation, cancer cell drug resistance, survival after myocardial infarction, efferocytosis, hypo-matrix metalloproteinase, vasoconstriction and vasodilatation. Known small molecule stimulators of the channels include hyperforin, genistein and rosiglitazone, but there is more progress with inhibitors, some of which have promising potency and selectivity. The inhibitors include 2-aminoethoxydiphenyl borate, 2-aminoquinolines, 2-aminothiazoles, fatty acids, isothiourea derivatives, naphthalene sulfonamides, N-phenylanthranilic acids, phenylethylimidazoles, piperazine/piperidine analogues, polyphenols, pyrazoles and steroids. A few of these agents are starting to be useful as tools for determining the physiological and pathophysiological functions of TRPC channels. We suggest that the pursuit of small molecule modulators for TRPC channels is important but that it requires substantial additional effort and investment before we can reap the rewards of highly potent and selective pharmacological modulators. PMID:23763262

High-throughput platform assay technology for the discovery of pre-microrna-selective small molecule probes.

PubMed

Lorenz, Daniel A; Song, James M; Garner, Amanda L

2015-01-21

MicroRNAs (miRNA) play critical roles in human development and disease. As such, the targeting of miRNAs is considered attractive as a novel therapeutic strategy. A major bottleneck toward this goal, however, has been the identification of small molecule probes that are specific for select RNAs and methods that will facilitate such discovery efforts. Using pre-microRNAs as proof-of-concept, herein we report a conceptually new and innovative approach for assaying RNA-small molecule interactions. Through this platform assay technology, which we term catalytic enzyme-linked click chemistry assay or cat-ELCCA, we have designed a method that can be implemented in high throughput, is virtually free of false readouts, and is general for all nucleic acids. Through cat-ELCCA, we envision the discovery of selective small molecule ligands for disease-relevant miRNAs to promote the field of RNA-targeted drug discovery and further our understanding of the role of miRNAs in cellular biology.

A semantic web ontology for small molecules and their biological targets.

PubMed

Choi, Jooyoung; Davis, Melissa J; Newman, Andrew F; Ragan, Mark A

2010-05-24

A wide range of data on sequences, structures, pathways, and networks of genes and gene products is available for hypothesis testing and discovery in biological and biomedical research. However, data describing the physical, chemical, and biological properties of small molecules have not been well-integrated with these resources. Semantically rich representations of chemical data, combined with Semantic Web technologies, have the potential to enable the integration of small molecule and biomolecular data resources, expanding the scope and power of biomedical and pharmacological research. We employed the Semantic Web technologies Resource Description Framework (RDF) and Web Ontology Language (OWL) to generate a Small Molecule Ontology (SMO) that represents concepts and provides unique identifiers for biologically relevant properties of small molecules and their interactions with biomolecules, such as proteins. We instanced SMO using data from three public data sources, i.e., DrugBank, PubChem and UniProt, and converted to RDF triples. Evaluation of SMO by use of predetermined competency questions implemented as SPARQL queries demonstrated that data from chemical and biomolecular data sources were effectively represented and that useful knowledge can be extracted. These results illustrate the potential of Semantic Web technologies in chemical, biological, and pharmacological research and in drug discovery.

Exporters for Production of Amino Acids and Other Small Molecules.

PubMed

Eggeling, Lothar

Microbes are talented catalysts to synthesize valuable small molecules in their cytosol. However, to make full use of their skills - and that of metabolic engineers - the export of intracellularly synthesized molecules to the culture medium has to be considered. This step is as essential as is each step for the synthesis of the favorite molecule of the metabolic engineer, but is frequently not taken into account. To export small molecules via the microbial cell envelope, a range of different types of carrier proteins is recognized to be involved, which are primary active carriers, secondary active carriers, or proteins increasing diffusion. Relevant export may require just one carrier as is the case with L-lysine export by Corynebacterium glutamicum or involve up to four carriers as known for L-cysteine excretion by Escherichia coli. Meanwhile carriers for a number of small molecules of biotechnological interest are recognized, like for production of peptides, nucleosides, diamines, organic acids, or biofuels. In addition to carriers involved in amino acid excretion, such carriers and their impact on product formation are described, as well as the relatedness of export carriers which may serve as a hint to identify further carriers required to improve product formation by engineering export.

Challenges and Opportunities for Small-Molecule Fluorescent Probes in Redox Biology Applications.

PubMed

Jiang, Xiqian; Wang, Lingfei; Carroll, Shaina L; Chen, Jianwei; Wang, Meng C; Wang, Jin

2018-02-16

The concentrations of reactive oxygen/nitrogen species (ROS/RNS) are critical to various biochemical processes. Small-molecule fluorescent probes have been widely used to detect and/or quantify ROS/RNS in many redox biology studies and serve as an important complementary to protein-based sensors with unique applications. Recent Advances: New sensing reactions have emerged in probe development, allowing more selective and quantitative detection of ROS/RNS, especially in live cells. Improvements have been made in sensing reactions, fluorophores, and bioavailability of probe molecules. In this review, we will not only summarize redox-related small-molecule fluorescent probes but also lay out the challenges of designing probes to help redox biologists independently evaluate the quality of reported small-molecule fluorescent probes, especially in the chemistry literature. We specifically highlight the advantages of reversibility in sensing reactions and its applications in ratiometric probe design for quantitative measurements in living cells. In addition, we compare the advantages and disadvantages of small-molecule probes and protein-based probes. The low physiological relevant concentrations of most ROS/RNS call for new sensing reactions with better selectivity, kinetics, and reversibility; fluorophores with high quantum yield, wide wavelength coverage, and Stokes shifts; and structural design with good aqueous solubility, membrane permeability, low protein interference, and organelle specificity. Antioxid. Redox Signal. 00, 000-000.

Function of reactive oxygen species during animal development: passive or active?

PubMed

Covarrubias, Luis; Hernández-García, David; Schnabel, Denhí; Salas-Vidal, Enrique; Castro-Obregón, Susana

2008-08-01

Oxidative stress is considered causal of aging and pathological cell death, however, very little is known about its function in the natural processes that support the formation of an organism. It is generally thought that cells must continuously protect themselves from the possible damage caused by reactive oxygen species (ROS) (passive ROS function). However, presently, ROS are recognized as physiologically relevant molecules that mediate cell responses to a variety of stimuli, and the activities of several molecules, some developmentally relevant, are directly or indirectly regulated by oxidative stress (active ROS function). Here we review recent data that are suggestive of specific ROS functions during development of animals, particularly mammals.

Molecular sled sequences are common in mammalian proteins.

PubMed

Xiong, Kan; Blainey, Paul C

2016-03-18

Recent work revealed a new class of molecular machines called molecular sleds, which are small basic molecules that bind and slide along DNA with the ability to carry cargo along DNA. Here, we performed biochemical and single-molecule flow stretching assays to investigate the basis of sliding activity in molecular sleds. In particular, we identified the functional core of pVIc, the first molecular sled characterized; peptide functional groups that control sliding activity; and propose a model for the sliding activity of molecular sleds. We also observed widespread DNA binding and sliding activity among basic polypeptide sequences that implicate mammalian nuclear localization sequences and many cell penetrating peptides as molecular sleds. These basic protein motifs exhibit weak but physiologically relevant sequence-nonspecific DNA affinity. Our findings indicate that many mammalian proteins contain molecular sled sequences and suggest the possibility that substantial undiscovered sliding activity exists among nuclear mammalian proteins. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

Allosteric ligands for the pharmacologically dark receptors GPR68 and GPR65

PubMed Central

Huang, Xi-Ping; Karpiak, Joel; Kroeze, Wesley K.; Zhu, Hu; Chen, Xin; Moy, Sheryl S.; Saddoris, Kara A.; Nikolova, Viktoriya; Farrell, Martilias S.; Wang, Sheng; Mangano, Thomas J.; Deshpande, Deepak A.; Jiang, Alice; Penn, Raymond B.; Jin, Jian; Koller, Beverly H.; Kenakin, Terry; Shoichet, Brian K.; Roth, Bryan L.

2016-01-01

At least 120 non-olfactory G protein-coupled receptors in the human genome are ”orphans” for which endogenous ligands are unknown, and many have no selective ligands, hindering elucidation of their biological functions and clinical relevance. Among these is GPR68, a proton receptor that lacks small molecule modulators for probing its biology. Yeast-based screens against GPR68 identified the benzodiazepine drug lorazepam as a non-selective GPR68 positive allosteric modulator. Over 3000 GPR68 homology models were refined to recognize lorazepam in a putative allosteric site. Docking 3.1 million molecules predicted new GPR68 modulators many of which were confirmed in functional assays. One potent GPR68 modulator—ogerin– suppressed recall in fear conditioning in wild-type, but not in GPR68 knockout mice. The same approach led to the discovery of allosteric agonists and negative allosteric modulators for GPR65. Combining physical and structure-based screening may be broadly useful for ligand discovery for understudied and orphan GPCRs. PMID:26550826

Application of encoded library technology (ELT) to a protein-protein interaction target: discovery of a potent class of integrin lymphocyte function-associated antigen 1 (LFA-1) antagonists.

PubMed

Kollmann, Christopher S; Bai, Xiaopeng; Tsai, Ching-Hsuan; Yang, Hongfang; Lind, Kenneth E; Skinner, Steven R; Zhu, Zhengrong; Israel, David I; Cuozzo, John W; Morgan, Barry A; Yuki, Koichi; Xie, Can; Springer, Timothy A; Shimaoka, Motomu; Evindar, Ghotas

2014-04-01

The inhibition of protein-protein interactions remains a challenge for traditional small molecule drug discovery. Here we describe the use of DNA-encoded library technology for the discovery of small molecules that are potent inhibitors of the interaction between lymphocyte function-associated antigen 1 and its ligand intercellular adhesion molecule 1. A DNA-encoded library with a potential complexity of 4.1 billion compounds was exposed to the I-domain of the target protein and the bound ligands were affinity selected, yielding an enriched small-molecule hit family. Compounds representing this family were synthesized without their DNA encoding moiety and found to inhibit the lymphocyte function-associated antigen 1/intercellular adhesion molecule-1 interaction with submicromolar potency in both ELISA and cell adhesion assays. Re-synthesized compounds conjugated to DNA or a fluorophore were demonstrated to bind to cells expressing the target protein. Copyright © 2014 Elsevier Ltd. All rights reserved.

Screening small-molecule compound microarrays for protein ligands without fluorescence labeling with a high-throughput scanning microscope.

PubMed

Fei, Yiyan; Landry, James P; Sun, Yungshin; Zhu, Xiangdong; Wang, Xiaobing; Luo, Juntao; Wu, Chun-Yi; Lam, Kit S

2010-01-01

We describe a high-throughput scanning optical microscope for detecting small-molecule compound microarrays on functionalized glass slides. It is based on measurements of oblique-incidence reflectivity difference and employs a combination of a y-scan galvometer mirror and an x-scan translation stage with an effective field of view of 2 cm x 4 cm. Such a field of view can accommodate a printed small-molecule compound microarray with as many as 10,000 to 20,000 targets. The scanning microscope is capable of measuring kinetics as well as endpoints of protein-ligand reactions simultaneously. We present the experimental results on solution-phase protein reactions with small-molecule compound microarrays synthesized from one-bead, one-compound combinatorial chemistry and immobilized on a streptavidin-functionalized glass slide.

Screening small-molecule compound microarrays for protein ligands without fluorescence labeling with a high-throughput scanning microscope

PubMed Central

Fei, Yiyan; Landry, James P.; Sun, Yungshin; Zhu, Xiangdong; Wang, Xiaobing; Luo, Juntao; Wu, Chun-Yi; Lam, Kit S.

2010-01-01

We describe a high-throughput scanning optical microscope for detecting small-molecule compound microarrays on functionalized glass slides. It is based on measurements of oblique-incidence reflectivity difference and employs a combination of a y-scan galvometer mirror and an x-scan translation stage with an effective field of view of 2 cm×4 cm. Such a field of view can accommodate a printed small-molecule compound microarray with as many as 10,000 to 20,000 targets. The scanning microscope is capable of measuring kinetics as well as endpoints of protein-ligand reactions simultaneously. We present the experimental results on solution-phase protein reactions with small-molecule compound microarrays synthesized from one-bead, one-compound combinatorial chemistry and immobilized on a streptavidin-functionalized glass slide. PMID:20210464

Generation of small molecules to interfere with regulated necrosis.

PubMed

Degterev, Alexei; Linkermann, Andreas

2016-06-01

Interference with regulated necrosis for clinical purposes carries broad therapeutic relevance and, if successfully achieved, has a potential to revolutionize everyday clinical routine. Necrosis was interpreted as something that no clinician might ever be able to prevent due to the unregulated nature of this form of cell death. However, given our growing understanding of the existence of regulated forms of necrosis and the roles of key enzymes of these pathways, e.g., kinases, peroxidases, etc., the possibility emerges to identify efficient and selective small molecule inhibitors of pathologic necrosis. Here, we review the published literature on small molecule inhibition of regulated necrosis and provide an outlook on how combination therapy may be most effective in treatment of necrosis-associated clinical situations like stroke, myocardial infarction, sepsis, cancer and solid organ transplantation.

Computational active site analysis of molecular pathways to improve functional classification of enzymes.

PubMed

Ozyurt, A Sinem; Selby, Thomas L

2008-07-01

This study describes a method to computationally assess the function of homologous enzymes through small molecule binding interaction energy. Three experimentally determined X-ray structures and four enzyme models from ornithine cyclo-deaminase, alanine dehydrogenase, and mu-crystallin were used in combination with nine small molecules to derive a function score (FS) for each enzyme-model combination. While energy values varied for a single molecule-enzyme combination due to differences in the active sites, we observe that the binding energies for the entire pathway were proportional for each set of small molecules investigated. This proportionality of energies for a reaction pathway appears to be dependent on the amino acids in the active site and their direct interactions with the small molecules, which allows a function score (FS) to be calculated to assess the specificity of each enzyme. Potential of mean force (PMF) calculations were used to obtain the energies, and the resulting FS values demonstrate that a measurement of function may be obtained using differences between these PMF values. Additionally, limitations of this method are discussed based on: (a) larger substrates with significant conformational flexibility; (b) low homology enzymes; and (c) open active sites. This method should be useful in accurately predicting specificity for single enzymes that have multiple steps in their reactions and in high throughput computational methods to accurately annotate uncharacterized proteins based on active site interaction analysis. 2008 Wiley-Liss, Inc.

Gauge-origin dependence in electronic g-tensor calculations

NASA Astrophysics Data System (ADS)

Glasbrenner, Michael; Vogler, Sigurd; Ochsenfeld, Christian

2018-06-01

We present a benchmark study on the gauge-origin dependence of the electronic g-tensor using data from unrestricted density functional theory calculations with the spin-orbit mean field ansatz. Our data suggest in accordance with previous studies that g-tensor calculations employing a common gauge-origin are sufficiently accurate for small molecules; however, for extended molecules, the introduced errors can become relevant and significantly exceed the basis set error. Using calculations with the spin-orbit mean field ansatz and gauge-including atomic orbitals as a reference, we furthermore show that the accuracy and reliability of common gauge-origin approaches in larger molecules depends strongly on the locality of the spin density distribution. We propose a new pragmatic ansatz for choosing the gauge-origin which takes the spin density distribution into account and gives reasonably accurate values for molecules with a single localized spin center. For more general cases like molecules with several spatially distant spin centers, common gauge-origin approaches are shown to be insufficient for consistently achieving high accuracy. Therefore the computation of g-tensors using distributed gauge-origin methods like gauge-including atomic orbitals is considered as the ideal approach and is recommended for larger molecular systems.

Computational biology of RNA interactions.

PubMed

Dieterich, Christoph; Stadler, Peter F

2013-01-01

The biodiversity of the RNA world has been underestimated for decades. RNA molecules are key building blocks, sensors, and regulators of modern cells. The biological function of RNA molecules cannot be separated from their ability to bind to and interact with a wide space of chemical species, including small molecules, nucleic acids, and proteins. Computational chemists, physicists, and biologists have developed a rich tool set for modeling and predicting RNA interactions. These interactions are to some extent determined by the binding conformation of the RNA molecule. RNA binding conformations are approximated with often acceptable accuracy by sequence and secondary structure motifs. Secondary structure ensembles of a given RNA molecule can be efficiently computed in many relevant situations by employing a standard energy model for base pair interactions and dynamic programming techniques. The case of bi-molecular RNA-RNA interactions can be seen as an extension of this approach. However, unbiased transcriptome-wide scans for local RNA-RNA interactions are computationally challenging yet become efficient if the binding motif/mode is known and other external information can be used to confine the search space. Computational methods are less developed for proteins and small molecules, which bind to RNA with very high specificity. Binding descriptors of proteins are usually determined by in vitro high-throughput assays (e.g., microarrays or sequencing). Intriguingly, recent experimental advances, which are mostly based on light-induced cross-linking of binding partners, render in vivo binding patterns accessible yet require new computational methods for careful data interpretation. The grand challenge is to model the in vivo situation where a complex interplay of RNA binders competes for the same target RNA molecule. Evidently, bioinformaticians are just catching up with the impressive pace of these developments. Copyright © 2012 John Wiley & Sons, Ltd.

Molecular targets for small-molecule modulators of circadian clocks

PubMed Central

He, Baokun; Chen, Zheng

2016-01-01

Background Circadian clocks are endogenous timing systems that regulate various aspects of mammalian metabolism, physiology and behavior. Traditional chronotherapy refers to the administration of drugs in a defined circadian time window to achieve optimal pharmacokinetic and therapeutic efficacies. In recent years, substantial efforts have been dedicated to developing novel small-molecule modulators of circadian clocks. Methods Here, we review the recent progress in the identification of molecular targets of small-molecule clock modulators and their efficacies in clock-related disorders. Specifically, we examine the clock components and regulatory factors as possible molecular targets of small molecules, and we review several key clock-related disorders as promising venues for testing the preventive/therapeutic efficacies of these small molecules. Finally, we also discuss circadian regulation of drug metabolism. Results Small molecules can modulate the period, phase and/or amplitude of the circadian cycle. Core clock proteins, nuclear hormone receptors, and clock-related kinases and other epigenetic regulators are promising molecular targets for small molecules. Through these targets small molecules exert protective effects against clock-related disorders including the metabolic syndrome, immune disorders, sleep disorders and cancer. Small molecules can also modulate circadian drug metabolism and response to existing therapeutics. Conclusion Small-molecule clock modulators target clock components or diverse cellular pathways that functionally impinge upon the clock. Target identification of new small-molecule modulators will deepen our understanding of key regulatory nodes in the circadian network. Studies of clock modulators will facilitate their therapeutic applications, alone or in combination, for clock-related diseases. PMID:26750111

Functional porous composites by blending with solution-processable molecular pores.

PubMed

Jiang, S; Chen, L; Briggs, M E; Hasell, T; Cooper, A I

2016-05-25

We present a simple method for rendering non-porous materials porous by solution co-processing with organic cage molecules. This method can be used both for small functional molecules and for polymers, thus creating porous composites by molecular blending, rather than the more traditional approach of supporting functional molecules on pre-frabricated porous supports.

ACTP: A webserver for predicting potential targets and relevant pathways of autophagy-modulating compounds

PubMed Central

Ouyang, Liang; Cai, Haoyang; Liu, Bo

2016-01-01

Autophagy (macroautophagy) is well known as an evolutionarily conserved lysosomal degradation process for long-lived proteins and damaged organelles. Recently, accumulating evidence has revealed a series of small-molecule compounds that may activate or inhibit autophagy for therapeutic potential on human diseases. However, targeting autophagy for drug discovery still remains in its infancy. In this study, we developed a webserver called Autophagic Compound-Target Prediction (ACTP) (http://actp.liu-lab.com/) that could predict autophagic targets and relevant pathways for a given compound. The flexible docking of submitted small-molecule compound (s) to potential autophagic targets could be performed by backend reverse docking. The webpage would return structure-based scores and relevant pathways for each predicted target. Thus, these results provide a basis for the rapid prediction of potential targets/pathways of possible autophagy-activating or autophagy-inhibiting compounds without labor-intensive experiments. Moreover, ACTP will be helpful to shed light on identifying more novel autophagy-activating or autophagy-inhibiting compounds for future therapeutic implications. PMID:26824420

Targeting RNA in mammalian systems with small molecules.

PubMed

Donlic, Anita; Hargrove, Amanda E

2018-05-03

The recognition of RNA functions beyond canonical protein synthesis has challenged the central dogma of molecular biology. Indeed, RNA is now known to directly regulate many important cellular processes, including transcription, splicing, translation, and epigenetic modifications. The misregulation of these processes in disease has led to an appreciation of RNA as a therapeutic target. This potential was first recognized in bacteria and viruses, but discoveries of new RNA classes following the sequencing of the human genome have invigorated exploration of its disease-related functions in mammals. As stable structure formation is evolving as a hallmark of mammalian RNAs, the prospect of utilizing small molecules to specifically probe the function of RNA structural domains and their interactions is gaining increased recognition. To date, researchers have discovered bioactive small molecules that modulate phenotypes by binding to expanded repeats, microRNAs, G-quadruplex structures, and RNA splice sites in neurological disorders, cancers, and other diseases. The lessons learned from achieving these successes both call for additional studies and encourage exploration of the plethora of mammalian RNAs whose precise mechanisms of action remain to be elucidated. Efforts toward understanding fundamental principles of small molecule-RNA recognition combined with advances in methodology development should pave the way toward targeting emerging RNA classes such as long noncoding RNAs. Together, these endeavors can unlock the full potential of small molecule-based probing of RNA-regulated processes and enable us to discover new biology and underexplored avenues for therapeutic intervention in human disease. This article is categorized under: RNA Methods > RNA Analyses In Vitro and In Silico RNA Interactions with Proteins and Other Molecules > Small Molecule-RNA Interactions RNA in Disease and Development > RNA in Disease. © 2018 Wiley Periodicals, Inc.

Small molecule alteration of RNA sequence in cells and animals.

PubMed

Guan, Lirui; Luo, Yiling; Ja, William W; Disney, Matthew D

2017-10-18

RNA regulation and maintenance are critical for proper cell function. Small molecules that specifically alter RNA sequence would be exceptionally useful as probes of RNA structure and function or as potential therapeutics. Here, we demonstrate a photochemical approach for altering the trinucleotide expanded repeat causative of myotonic muscular dystrophy type 1 (DM1), r(CUG) exp . The small molecule, 2H-4-Ru, binds to r(CUG) exp and converts guanosine residues to 8-oxo-7,8-dihydroguanosine upon photochemical irradiation. We demonstrate targeted modification upon irradiation in cell culture and in Drosophila larvae provided a diet containing 2H-4-Ru. Our results highlight a general chemical biology approach for altering RNA sequence in vivo by using small molecules and photochemistry. Furthermore, these studies show that addition of 8-oxo-G lesions into RNA 3' untranslated regions does not affect its steady state levels. Copyright © 2017 Elsevier Ltd. All rights reserved.

From Vesicles to Protocells: The Roles of Amphiphilic Molecules

PubMed Central

Sakuma, Yuka; Imai, Masayuki

2015-01-01

It is very challenging to construct protocells from molecular assemblies. An important step in this challenge is the achievement of vesicle dynamics that are relevant to cellular functions, such as membrane trafficking and self-reproduction, using amphiphilic molecules. Soft matter physics will play an important role in the development of vesicles that have these functions. Here, we show that simple binary phospholipid vesicles have the potential to reproduce the relevant functions of adhesion, pore formation and self-reproduction of vesicles, by coupling the lipid geometries (spontaneous curvatures) and the phase separation. This achievement will elucidate the pathway from molecular assembly to cellular life. PMID:25738256

Fiber optic SERS-based plasmonics nanobiosensing in single living cells

NASA Astrophysics Data System (ADS)

Scaffidi, Jonathan P.; Gregas, Molly K.; Seewaldt, Victoria; Vo-Dinh, Tuan

2009-05-01

We describe the development of small molecule-sensitive plasmonics-active fiber-optic nanoprobes suitable for intracellular bioanalysis in single living human cells using surface-enhanced Raman scattering (SERS) detection. The practical utility of SERS-based fiber-optic nanoprobes is illustrated by measurements of intracellular pH in HMEC- 15/hTERT immortalized "normal" human mammary epithelial cells and PC-3 human prostate cancer cells. The results indicate that fiber-optic nanoprobe insertion and interrogation provide a sensitive and selective means to monitor biologically-relevant small molecules at the single cell level.

Mapping the Small Molecule Interactome by Mass Spectrometry.

PubMed

Flaxman, Hope A; Woo, Christina M

2018-01-16

Mapping small molecule interactions throughout the proteome provides the critical structural basis for functional analysis of their impact on biochemistry. However, translation of mass spectrometry-based proteomics methods to directly profile the interaction between a small molecule and the whole proteome is challenging because of the substoichiometric nature of many interactions, the diversity of covalent and noncovalent interactions involved, and the subsequent computational complexity associated with their spectral assignment. Recent advances in chemical proteomics have begun fill this gap to provide a structural basis for the breadth of small molecule-protein interactions in the whole proteome. Innovations enabling direct characterization of the small molecule interactome include faster, more sensitive instrumentation coupled to chemical conjugation, enrichment, and labeling methods that facilitate detection and assignment. These methods have started to measure molecular interaction hotspots due to inherent differences in local amino acid reactivity and binding affinity throughout the proteome. Measurement of the small molecule interactome is producing structural insights and methods for probing and engineering protein biochemistry. Direct structural characterization of the small molecule interactome is a rapidly emerging area pushing new frontiers in biochemistry at the interface of small molecules and the proteome.

Reaction-based small-molecule fluorescent probes for dynamic detection of ROS and transient redox changes in living cells and small animals.

PubMed

Lü, Rui

2017-09-01

Dynamic detection of transient redox changes in living cells and animals has broad implications for human health and disease diagnosis, because intracellular redox homeostasis regulated by reactive oxygen species (ROS) plays important role in cell functions, normal physiological functions and some serious human diseases (e.g., cancer, Alzheimer's disease, diabetes, etc.) usually have close relationship with the intracellular redox status. Small-molecule ROS-responsive fluorescent probes can act as powerful tools for dynamic detection of ROS and redox changes in living cells and animals through fluorescence imaging techniques; and great advances have been achieved recently in the design and synthesis of small-molecule ROS-responsive fluorescent probes. This article highlights up-to-date achievements in designing and using the reaction-based small-molecule fluorescent probes (with high sensitivity and selectivity to ROS and redox cycles) in the dynamic detection of ROS and transient redox changes in living cells and animals through fluorescence imaging. Copyright © 2017. Published by Elsevier Ltd.

Aptamer Recognition of Multiplexed Small-Molecule-Functionalized Substrates.

PubMed

Nakatsuka, Nako; Cao, Huan H; Deshayes, Stephanie; Melkonian, Arin Lucy; Kasko, Andrea M; Weiss, Paul S; Andrews, Anne M

2018-05-31

Aptamers are chemically synthesized oligonucleotides or peptides with molecular recognition capabilities. We investigated recognition of substrate-tethered small-molecule targets, using neurotransmitters as examples, and fluorescently labeled DNA aptamers. Substrate regions patterned via microfluidic channels with dopamine or L-tryptophan were selectively recognized by previously identified dopamine or L-tryptophan aptamers, respectively. The on-substrate dissociation constant determined for the dopamine aptamer was comparable to, though slightly greater than the previously determined solution dissociation constant. Using pre-functionalized neurotransmitter-conjugated oligo(ethylene glycol) alkanethiols and microfluidics patterning, we produced multiplexed substrates to capture and to sort aptamers. Substrates patterned with L-DOPA, L-DOPS, and L-5-HTP enabled comparison of the selectivity of the dopamine aptamer for different targets via simultaneous determination of in situ binding constants. Thus, beyond our previous demonstrations of recognition by protein binding partners (i.e., antibodies and G-protein-coupled receptors), strategically optimized small-molecule-functionalized substrates show selective recognition of nucleic acid binding partners. These substrates are useful for side-by-side target comparisons, and future identification and characterization of novel aptamers targeting neurotransmitters or other important small-molecules.

Screening of Small Molecule Interactor Library by Using In-Cell NMR Spectroscopy (SMILI-NMR)

PubMed Central

Xie, Jingjing; Thapa, Rajiv; Reverdatto, Sergey; Burz, David S.; Shekhtman, Alexander

2011-01-01

We developed an in-cell NMR assay for screening small molecule interactor libraries (SMILI-NMR) for compounds capable of disrupting or enhancing specific interactions between two or more components of a biomolecular complex. The method relies on the formation of a well-defined biocomplex and utilizes in-cell NMR spectroscopy to identify the molecular surfaces involved in the interaction at atomic scale resolution. Changes in the interaction surface caused by a small molecule interfering with complex formation are used as a read-out of the assay. The in-cell nature of the experimental protocol insures that the small molecule is capable of penetrating the cell membrane and specifically engaging the target molecule(s). Utility of the method was demonstrated by screening a small dipeptide library against the FKBP–FRB protein complex involved in cell cycle arrest. The dipeptide identified by SMILI-NMR showed biological activity in a functional assay in yeast. PMID:19422228

A fluid membrane enhances the velocity of cargo transport by small teams of kinesin-1

NASA Astrophysics Data System (ADS)

Li, Qiaochu; Tseng, Kuo-Fu; King, Stephen J.; Qiu, Weihong; Xu, Jing

2018-03-01

Kinesin-1 (hereafter referred to as kinesin) is a major microtubule-based motor protein for plus-end-directed intracellular transport in live cells. While the single-molecule functions of kinesin are well characterized, the physiologically relevant transport of membranous cargos by small teams of kinesins remains poorly understood. A key experimental challenge remains in the quantitative control of the number of motors driving transport. Here we utilized "motile fraction" to overcome this challenge and experimentally accessed transport by a single kinesin through the physiologically relevant transport by a small team of kinesins. We used a fluid lipid bilayer to model the cellular membrane in vitro and employed optical trapping to quantify the transport of membrane-enclosed cargos versus traditional membrane-free cargos under identical conditions. We found that coupling motors via a fluid membrane significantly enhances the velocity of cargo transport by small teams of kinesins. Importantly, enclosing a cargo in a fluid lipid membrane did not impact single-kinesin transport, indicating that membrane-dependent velocity enhancement for team-based transport arises from altered interactions between kinesins. Our study demonstrates that membrane-based coupling between motors is a key determinant of kinesin-based transport. Enhanced velocity may be critical for fast delivery of cargos in live cells.

Influence of thermocleavable functionality on organic field-effect transistor performance of small molecules

NASA Astrophysics Data System (ADS)

Mahale, Rajashree Y.; Dharmapurikar, Satej S.; Chini, Mrinmoy Kumar; Venugopalan, Vijay

2017-06-01

Diketopyrrolopyrrole based donor-acceptor-donor conjugated small molecules using ethylene dioxythiophene as a donor was synthesized. Electron deficient diketopyrrolopyrrole unit was substituted with thermocleavable (tert-butyl acetate) side chains. The thermal treatment of the molecules at 160 °C eliminated the tert-butyl ester group results in the formation of corresponding acid. Optical and theoretical studies revealed that the molecules adopted a change in molecular arrangement after thermolysis. The conjugated small molecules possessed p-channel charge transport characteristics in organic field effect transistors. The charge carrier mobility was increased after thermolysis of tert-butyl ester group to 5.07 × 10-5 cm2/V s.

Caenorhabditis elegans chemical biology: lessons from small molecules

USDA-ARS?s Scientific Manuscript database

How can we complement Caenorhabditis elegans genomics and proteomics with a comprehensive structural and functional annotation of its metabolome? Several lines of evidence indicate that small molecules of largely undetermined structure play important roles in C. elegans biology, including key pathw...

[Relevance of long non-coding RNAs in tumour biology].

PubMed

Nagy, Zoltán; Szabó, Diána Rita; Zsippai, Adrienn; Falus, András; Rácz, Károly; Igaz, Péter

2012-09-23

The discovery of the biological relevance of non-coding RNA molecules represents one of the most significant advances in contemporary molecular biology. It has turned out that a major fraction of the non-coding part of the genome is transcribed. Beside small RNAs (including microRNAs) more and more data are disclosed concerning long non-coding RNAs of 200 nucleotides to 100 kb length that are implicated in the regulation of several basic molecular processes (cell proliferation, chromatin functioning, microRNA-mediated effects, etc.). Some of these long non-coding RNAs have been associated with human tumours, including H19, HOTAIR, MALAT1, etc., the different expression of which has been noted in various neoplasms relative to healthy tissues. Long non-coding RNAs may represent novel markers of molecular diagnostics and they might even turn out to be targets of therapeutic intervention.

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

Iconaru, Luigi I.; Ban, David; Bharatham, Kavitha

In disordered proteins we see that they are highly prevalent in biological systems. They control myriad signaling and regulatory processes, and their levels and/or cellular localization are often altered in human disease. In contrast to folded proteins, disordered proteins, due to conformational heterogeneity and dynamics, are not considered viable drug targets. We challenged this paradigm by identifying through NMR-based screening small molecules that bound specifically, albeit weakly, to the disordered cell cycle regulator, p27 Kip1 (p27). Moreover, two groups of molecules bound to sites created by transient clusters of aromatic residues within p27. Conserved chemical features within these two groupsmore » of small molecules exhibited complementarity to their binding sites within p27, establishing structure-activity relationships for small molecule: disordered protein interactions. Finally, one compound counteracted the Cdk2/cyclin A inhibitory function of p27 in vitro, providing proof-of- principle that small molecules can inhibit the function of a disordered protein (p27) through sequestration in a conformation incapable of folding and binding to a natural regulatory target (Cdk2/cyclin A).« less

Mesoporous Silicate Materials in Sensing

PubMed Central

Melde, Brian J.; Johnson, Brandy J.; Charles, Paul T.

2008-01-01

Mesoporous silicas, especially those exhibiting ordered pore systems and uniform pore diameters, have shown great potential for sensing applications in recent years. Morphological control grants them versatility in the method of deployment whether as bulk powders, monoliths, thin films, or embedded in coatings. High surface areas and pore sizes greater than 2 nm make them effective as adsorbent coatings for humidity sensors. The pore networks also provide the potential for immobilization of enzymes within the materials. Functionalization of materials by silane grafting or through co-condensation of silicate precursors can be used to provide mesoporous materials with a variety of fluorescent probes as well as surface properties that aid in selective detection of specific analytes. This review will illustrate how mesoporous silicas have been applied to sensing changes in relative humidity, changes in pH, metal cations, toxic industrial compounds, volatile organic compounds, small molecules and ions, nitroenergetic compounds, and biologically relevant molecules. PMID:27873810

The intrinsic role of nanoconfinement in chemical equilibrium: evidence from DNA hybridization.

PubMed

Rubinovich, Leonid; Polak, Micha

2013-05-08

Recently we predicted that when a reaction involving a small number of molecules occurs in a nanometric-scale domain entirely segregated from the surrounding media, the nanoconfinement can shift the position of equilibrium toward products via reactant-product reduced mixing. In this Letter, we demonstrate how most-recently reported single-molecule fluorescence measurements of partial hybridization of ssDNA confined within nanofabricated chambers provide the first experimental confirmation of this entropic nanoconfinement effect. Thus, focusing separately on each occupancy-specific equilibrium constant, quantitatively reveals extra stabilization of the product upon decreasing the chamber occupancy or size. Namely, the DNA hybridization under nanoconfined conditions is significantly favored over the identical reaction occurring in bulk media with the same reactant concentrations. This effect, now directly verified for DNA, can be relevant to actual biological processes, as well as to diverse reactions occurring within molecular capsules, nanotubes, and other functional nanospaces.

Osmotic pressure induced tensile forces in tendon collagen

NASA Astrophysics Data System (ADS)

Masic, Admir; Bertinetti, Luca; Schuetz, Roman; Chang, Shu-Wei; Metzger, Till Hartmut; Buehler, Markus J.; Fratzl, Peter

2015-01-01

Water is an important component of collagen in tendons, but its role for the function of this load-carrying protein structure is poorly understood. Here we use a combination of multi-scale experimentation and computation to show that water is an integral part of the collagen molecule, which changes conformation upon water removal. The consequence is a shortening of the molecule that translates into tensile stresses in the range of several to almost 100 MPa, largely surpassing those of about 0.3 MPa generated by contractile muscles. Although a complete drying of collagen would be relevant for technical applications, such as the fabrication of leather or parchment, stresses comparable to muscle contraction already occur at small osmotic pressures common in biological environments. We suggest, therefore, that water-generated tensile stresses may play a role in living collagen-based materials such as tendon or bone.

9-ING-41, a small-molecule glycogen synthase kinase-3 inhibitor, is active in neuroblastoma.

PubMed

Ugolkov, Andrey V; Bondarenko, Gennadiy I; Dubrovskyi, Oleksii; Berbegall, Ana P; Navarro, Samuel; Noguera, Rosa; O'Halloran, Thomas V; Hendrix, Mary J; Giles, Francis J; Mazar, Andrew P

2018-05-25

Advanced stage neuroblastoma is a very aggressive pediatric cancer with limited treatment options and a high mortality rate. Glycogen synthase kinase-3β (GSK-3β) is a potential therapeutic target in neuroblastoma. Using immunohistochemical staining, we observed positive GSK-3β expression in 67% of human neuroblastomas (34 of 51 cases). Chemically distinct GSK-3 inhibitors (AR-A014418, TDZD-8, and 9-ING-41) suppressed the growth of neuroblastoma cells, whereas 9-ING-41, a clinically relevant small-molecule GSK-3β inhibitor with broad-spectrum preclinical antitumor activity, being the most potent. Inhibition of GSK-3 resulted in a decreased expression of the antiapoptotic molecule XIAP and an increase in neuroblastoma cell apoptosis. Mouse xenograft studies showed that the combination of clinically relevant doses of CPT-11 and 9-ING-41 led to greater antitumor effect than was observed with either agent alone. These data support the inclusion of patients with advanced neuroblastoma in clinical studies of 9-ING-41, especially in combination with CPT-11.

Extensive survey of molecules related to wood formation and gravity for space agriculture.

NASA Astrophysics Data System (ADS)

Motohashi, Kyohei; Tomita-Yokotani, Kaori; Baba, Keiichi; Furukawa, Jun; Sato, Seigo; Suzuki, Toshisada; Hashimoto, Hirofumi; Yamashita, Masamichi; Japanese Space Tree Working Group

Most, if not all, terrestrial subjects are under the influence of gravity. Since the gravitational force is proportional to the mass of subject, gravity is dominant for larger masses. The response of a plant against gravity is not an exception in this respect even it shows rather complicated features. For the angiosperm tree, its shape is determined by the forming tension wood, which induces more tensile stress in the xylem than in the normal wood. The mechanism of tension wood formation and its relevance to gravity have been extensively studied. Gibberellin is known to be responsible for this phenomenon in angiosperm tree, for example, the Japanese cherry tree, Prunus jamasakura. However, full understanding of the mechanisms has not yet been clarified. For an extensive survey of molecules related to tension wood formation, we induced an artificial tension wood formation and examined the tension wood formation by microscopic observations with double-staining. This enables the screening of really functional molecules in the space environment for future space agriculture. We demonstrated that Prunus incise is suitable for this research as a test material based on several reasons. We focused our attention in the region of the branch, i.e., the CosmoTree in CosmoBon, and established an experimental system to analyze the real functional factors of the tension wood. This study might ensure wood formation in a space environment and use woody plants as a material for space development. ("CosmoBon" is the Bonsai small tree for our space experiments. "CosmoTree" is a small branch/tree.)

Functional Conservation and Divergence of daf-22 Paralogs in Pristionchus pacificus Dauer Development.

PubMed

Markov, Gabriel V; Meyer, Jan M; Panda, Oishika; Artyukhin, Alexander B; Claaßen, Marc; Witte, Hanh; Schroeder, Frank C; Sommer, Ralf J

2016-10-01

Small-molecule signaling in nematode dauer formation has emerged as a major model to study chemical communication in development and evolution. Developmental arrest as nonfeeding and stress-resistant dauer larvae represents the major survival and dispersal strategy. Detailed studies in Caenorhabditis elegans and Pristionchus pacificus revealed that small-molecule communication changes rapidly in evolution resulting in extreme structural diversity of small-molecule compounds. In C. elegans, a blend of ascarosides constitutes the dauer pheromone, whereas the P. pacificus dauer pheromone includes additional paratosides and integrates building blocks from diverse primary metabolic pathways. Despite this complexity of small-molecule structures and functions, little is known about the biosynthesis of small molecules in nematodes outside C. elegans Here, we show that the genes encoding enzymes of the peroxisomal β-oxidation pathway involved in small-molecule biosynthesis evolve rapidly, including gene duplications and domain switching. The thiolase daf-22, the most downstream factor in C. elegans peroxisomal β-oxidation, has duplicated in P. pacificus, resulting in Ppa-daf-22.1, which still contains the sterol-carrier-protein (SCP) domain that was lost in C. elegans daf-22, and Ppa-daf-22.2. Using the CRISPR/Cas9 system, we induced mutations in both P. pacificus daf-22 genes and identified an unexpected complexity of functional conservation and divergence. Under well-fed conditions, ascaroside biosynthesis proceeds exclusively via Ppa-daf-22.1 In contrast, starvation conditions induce Ppa-daf-22.2 activity, resulting in the production of a specific subset of ascarosides. Gene expression studies indicate a reciprocal up-regulation of both Ppa-daf-22 genes, which is, however, independent of starvation. Thus, our study reveals an unexpected functional complexity of dauer development and evolution. © The Author 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Engineered M13 bacteriophage nanocarriers for intracellular delivery of exogenous proteins to human prostate cancer cells.

PubMed

DePorter, Sandra M; McNaughton, Brian R

2014-09-17

The size, well-defined structure, and relatively high folding energies of most proteins allow them to recognize disease-relevant receptors that present a challenge to small molecule reagents. While multiple challenges must be overcome in order to fully exploit the use of protein reagents in basic research and medicine, perhaps the greatest challenge is their intracellular delivery to a particular diseased cell. Here, we describe the genetic and enzymatic manipulation of prostate cancer cell-penetrating M13 bacteriophage to generate nanocarriers for the intracellular delivery of functional exogenous proteins to a human prostate cancer cell line.

Fundamentals and applications of SERS-based bioanalytical sensing

NASA Astrophysics Data System (ADS)

Kahraman, Mehmet; Mullen, Emma R.; Korkmaz, Aysun; Wachsmann-Hogiu, Sebastian

2017-03-01

Plasmonics is an emerging field that examines the interaction between light and metallic nanostructures at the metal-dielectric interface. Surface-enhanced Raman scattering (SERS) is a powerful analytical technique that uses plasmonics to obtain detailed chemical information of molecules or molecular assemblies adsorbed or attached to nanostructured metallic surfaces. For bioanalytical applications, these surfaces are engineered to optimize for high enhancement factors and molecular specificity. In this review we focus on the fabrication of SERS substrates and their use for bioanalytical applications. We review the fundamental mechanisms of SERS and parameters governing SERS enhancement. We also discuss developments in the field of novel SERS substrates. This includes the use of different materials, sizes, shapes, and architectures to achieve high sensitivity and specificity as well as tunability or flexibility. Different fundamental approaches are discussed, such as label-free and functional assays. In addition, we highlight recent relevant advances for bioanalytical SERS applied to small molecules, proteins, DNA, and biologically relevant nanoparticles. Subsequently, we discuss the importance of data analysis and signal detection schemes to achieve smaller instruments with low cost for SERS-based point-of-care technology developments. Finally, we review the main advantages and challenges of SERS-based biosensing and provide a brief outlook.

Exchange-biased quantum tunnelling in a supramolecular dimer of single-molecule magnets.

PubMed

Wernsdorfer, Wolfgang; Aliaga-Alcalde, Núria; Hendrickson, David N; Christou, George

2002-03-28

Various present and future specialized applications of magnets require monodisperse, small magnetic particles, and the discovery of molecules that can function as nanoscale magnets was an important development in this regard. These molecules act as single-domain magnetic particles that, below their blocking temperature, exhibit magnetization hysteresis, a classical property of macroscopic magnets. Such 'single-molecule magnets' (SMMs) straddle the interface between classical and quantum mechanical behaviour because they also display quantum tunnelling of magnetization and quantum phase interference. Quantum tunnelling of magnetization can be advantageous for some potential applications of SMMs, for example, in providing the quantum superposition of states required for quantum computing. However, it is a disadvantage in other applications, such as information storage, where it would lead to information loss. Thus it is important to both understand and control the quantum properties of SMMs. Here we report a supramolecular SMM dimer in which antiferromagnetic coupling between the two components results in quantum behaviour different from that of the individual SMMs. Our experimental observations and theoretical analysis suggest a means of tuning the quantum tunnelling of magnetization in SMMs. This system may also prove useful for studying quantum tunnelling of relevance to mesoscopic antiferromagnets.

Fisetin Acts on Multiple Pathways to Reduce the Impact of Age and Disease on CNS Function

PubMed Central

Maher, Pamela

2017-01-01

It is becoming increasingly clear that neurological diseases are multi-factorial involving disruptions in multiple cellular systems. Thus, while each disease has its own initiating mechanisms and pathologies, certain common pathways appear to be involved in most, if not all, neurological diseases described to date. Thus, it is unlikely that modulating only a single factor will be effective at either preventing disease development or slowing disease progression. A better approach is to identify small (< 900 daltons) molecules that have multiple biological activities relevant to the maintenance of brain function. Over the last few years, we have identified an orally active, novel neuroprotective and cognition-enhancing molecule, the flavonoid fisetin. Fisetin not only has direct antioxidant activity but it can also increase the intracellular levels of glutathione, the major intracellular antioxidant. Fisetin can also activate key neurotrophic factor signaling pathways. In addition, it has anti-inflammatory activity against microglial cells and inhibits the activity of lipoxygenases, thereby reducing the production of pro-inflammatory eicosanoids and their by-products. This wide range of actions suggests that fisetin has the ability to reduce the impact of age-related neurological diseases on brain function. PMID:25961687

Formylbenzene diazonium hexafluorophosphate reagent for tyrosine-selective modification of proteins and the introduction of a bioorthogonal aldehyde

PubMed Central

Gavrilyuk, Julia; Ban, Hitoshi; Nagano, Masanobu; Hakamata, Wataru; Barbas, Carlos F.

2012-01-01

4-Formylbenzene diazonium hexafluorophosphate (FBDP) is a novel bench-stable crystalline diazonium salt that reacts selectively with tyrosine to install a bioorthogonal aldehyde functionality. Model studies with N-acyl-tyrosine methylamide allowed us to identify conditions optimal for tyrosine ligation reactions with small peptides and proteins. FBDP-based conjugation was used for the facile introduction of small molecule tags, poly(ethylene) glycol chains (PEGylation), and functional small molecules onto model proteins and to label the surface of living cells. PMID:23181702

Formylbenzene diazonium hexafluorophosphate reagent for tyrosine-selective modification of proteins and the introduction of a bioorthogonal aldehyde.

PubMed

Gavrilyuk, Julia; Ban, Hitoshi; Nagano, Masanobu; Hakamata, Wataru; Barbas, Carlos F

2012-12-19

4-Formylbenzene diazonium hexafluorophosphate (FBDP) is a novel bench-stable crystalline diazonium salt that reacts selectively with tyrosine to install a bioorthogonal aldehyde functionality. Model studies with N-acyl-tyrosine methylamide allowed us to identify conditions optimal for tyrosine ligation reactions with small peptides and proteins. FBDP-based conjugation was used for the facile introduction of small molecule tags, poly(ethylene glycol) chains (PEGylation), and functional small molecules onto model proteins and to label the surface of living cells.

Small Molecule Signaling Agents: The Integrated Chemistry and Biochemistry of Nitrogen Oxides, Oxides of Carbon, Dioxygen, Hydrogen Sulfide, and Their Derived Species

PubMed Central

Fukuto, Jon M.; Carrington, Samantha J.; Tantillo, Dean J.; Harrison, Jason G.; Ignarro, Louis J.; Freeman, Bruce A.; Chen, Andrew; Wink, David A.

2014-01-01

Several small molecule species formally known primarily as toxic gases have, over the past 20 years, been shown to be endogenously generated signaling molecules. The biological signaling associated with the small molecules NO, CO, H2S (and the nonendogenously generated O2), and their derived species have become a topic of extreme interest. It has become increasingly clear that these small molecule signaling agents form an integrated signaling web that affects/regulates numerous physiological processes. The chemical interactions between these species and each other or biological targets is an important factor in their roles as signaling agents. Thus, a fundamental understanding of the chemistry of these molecules is essential to understanding their biological/physiological utility. This review focuses on this chemistry and attempts to establish the chemical basis for their signaling functions. PMID:22263838

A Small Molecule Inverse Agonist for the Human Thyroid-Stimulating Hormone Receptor

PubMed Central

Neumann, Susanne; Huang, Wenwei; Eliseeva, Elena; Titus, Steve; Thomas, Craig J.; Gershengorn, Marvin C.

2010-01-01

Small molecule inverse agonists for the TSH receptor (TSHR) may be used as probes of the role of basal (or agonist-independent or constitutive) signaling and may have therapeutic potential as orally active drugs to inhibit basal signaling in patients with thyroid cancer and in some patients with hyperthyroidism. We describe the first small-molecule ligand [1;2-(3-((2,6-dimethylphenoxy)methyl)-4-methoxyphenyl)-3-(furan-2-ylmethyl)-2,3-dihydroquinazolin-4(1H)-one] that exhibits inverse agonist properties at TSHR. 1 inhibits basal and TSH-stimulated signaling, measured as cAMP production, by TSHRs in HEK-EM 293 cells stably expressing wild-type TSHRs; the antagonism of TSH-mediated signaling is competitive. 1 also inhibits basal signaling by wild-type TSHRs, and four constitutively active mutants of TSHR expressed transiently in HEK-EM 293 cells. 1 was active under more physiologically relevant conditions in primary cultures of human thyrocytes expressing endogenous TSHRs where it inhibited basal levels of mRNA transcripts for thyroglobulin, thyroperoxidase, sodium iodide symporter, and TSHR. These data serve as proof of principle that small, drug-like molecules can inhibit basal signaling by TSHR. We suggest that this small molecule is a lead compound for the development of higher-potency inverse agonists that can be used as probes of TSHR biology with therapeutic potential. PMID:20427476

Cyclosporin derivatives inhibit hepatitis B virus entry without interfering with NTCP transporter activity.

PubMed

Shimura, Satomi; Watashi, Koichi; Fukano, Kento; Peel, Michael; Sluder, Ann; Kawai, Fumihiro; Iwamoto, Masashi; Tsukuda, Senko; Takeuchi, Junko S; Miyake, Takeshi; Sugiyama, Masaya; Ogasawara, Yuki; Park, Sam-Yong; Tanaka, Yasuhito; Kusuhara, Hiroyuki; Mizokami, Masashi; Sureau, Camille; Wakita, Takaji

2017-04-01

The sodium taurocholate co-transporting polypeptide (NTCP) is the main target of most hepatitis B virus (HBV) specific entry inhibitors. Unfortunately, these agents also block NTCP transport of bile acids into hepatocytes, and thus have the potential to cause adverse effects. We aimed to identify small molecules that inhibit HBV entry while maintaining NTCP transporter function. We characterized a series of cyclosporine (CsA) derivatives for their anti-HBV activity and NTCP binding specificity using HepG2 cells overexpressing NTCP and primary human hepatocytes. The four most potent derivatives were tested for their capacity to prevent HBV entry, but maintain NTCP transporter function. Their antiviral activity against different HBV genotypes was analysed. We identified several CsA derivatives that inhibited HBV infection with a sub-micromolar IC 50 . Among them, SCY446 and SCY450 showed low activity against calcineurin (CN) and cyclophilins (CyPs), two major CsA cellular targets. This suggested that instead, these compounds interacted directly with NTCP to inhibit viral attachment to host cells, and have no immunosuppressive function. Importantly, we found that SCY450 and SCY995 did not impair the NTCP-dependent uptake of bile acids, and inhibited multiple HBV genotypes including a clinically relevant nucleoside analog-resistant HBV isolate. This is the first example of small molecule selective inhibition of HBV entry with no decrease in NTCP transporter activity. It suggests that the anti-HBV activity can be functionally separated from bile acid transport. These broadly active anti-HBV molecules are potential candidates for developing new drugs with fewer adverse effects. In this study, we identified new compounds that selectively inhibited hepatitis B virus (HBV) entry, and did not impair bile acid uptake. Our evidence offers a new strategy for developing anti-HBV drugs with fewer side effects. Copyright © 2016 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Biological and functional relevance of CASP predictions

PubMed Central

Liu, Tianyun; Ish‐Shalom, Shirbi; Torng, Wen; Lafita, Aleix; Bock, Christian; Mort, Matthew; Cooper, David N; Bliven, Spencer; Capitani, Guido; Mooney, Sean D.

2017-01-01

Abstract Our goal is to answer the question: compared with experimental structures, how useful are predicted models for functional annotation? We assessed the functional utility of predicted models by comparing the performances of a suite of methods for functional characterization on the predictions and the experimental structures. We identified 28 sites in 25 protein targets to perform functional assessment. These 28 sites included nine sites with known ligand binding (holo‐sites), nine sites that are expected or suggested by experimental authors for small molecule binding (apo‐sites), and Ten sites containing important motifs, loops, or key residues with important disease‐associated mutations. We evaluated the utility of the predictions by comparing their microenvironments to the experimental structures. Overall structural quality correlates with functional utility. However, the best‐ranked predictions (global) may not have the best functional quality (local). Our assessment provides an ability to discriminate between predictions with high structural quality. When assessing ligand‐binding sites, most prediction methods have higher performance on apo‐sites than holo‐sites. Some servers show consistently high performance for certain types of functional sites. Finally, many functional sites are associated with protein‐protein interaction. We also analyzed biologically relevant features from the protein assemblies of two targets where the active site spanned the protein‐protein interface. For the assembly targets, we find that the features in the models are mainly determined by the choice of template. PMID:28975675

Hierarchical Co-Assembly Enhanced Direct Ink Writing.

PubMed

Li, Longyu; Zhang, Pengfei; Zhang, Zhiyun; Lin, Qianming; Wu, Yuyang; Cheng, Alexander; Lin, Yunxiao; Thompson, Christina M; Smaldone, Ronald A; Ke, Chenfeng

2018-04-23

Integrating intelligent molecular systems into 3D printing materials and transforming their molecular functions to the macroscale with controlled superstructures will unleash great potential for the development of smart materials. Compared to macromolecular 3D printing materials, self-assembled small-molecule-based 3D printing materials are very rare owing to the difficulties of facilitating 3D printability as well as preserving their molecular functions macroscopically. Herein, we report a general approach for the integration of functional small molecules into 3D printing materials for direct ink writing through the introduction of a supramolecular template. A variety of inorganic and organic small-molecule-based inks were 3D-printed, and their superstructures were refined by post-printing hierarchical co-assembly. Through spatial and temporal control of individual molecular events from the nano- to the macroscale, fine-tuned macroscale features were successfully installed in the monoliths. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Small-molecule kinase inhibitors provide insight into Mps1 cell cycle function.

PubMed

Kwiatkowski, Nicholas; Jelluma, Nannette; Filippakopoulos, Panagis; Soundararajan, Meera; Manak, Michael S; Kwon, Mijung; Choi, Hwan Geun; Sim, Taebo; Deveraux, Quinn L; Rottmann, Sabine; Pellman, David; Shah, Jagesh V; Kops, Geert J P L; Knapp, Stefan; Gray, Nathanael S

2010-05-01

Mps1, a dual-specificity kinase, is required for the proper functioning of the spindle assembly checkpoint and for the maintenance of chromosomal stability. As Mps1 function has been implicated in numerous phases of the cell cycle, the development of a potent, selective small-molecule inhibitor of Mps1 should facilitate dissection of Mps1-related biology. We describe the cellular effects and Mps1 cocrystal structures of new, selective small-molecule inhibitors of Mps1. Consistent with RNAi studies, chemical inhibition of Mps1 leads to defects in Mad1 and Mad2 establishment at unattached kinetochores, decreased Aurora B kinase activity, premature mitotic exit and gross aneuploidy, without any evidence of centrosome duplication defects. However, in U2OS cells having extra centrosomes (an abnormality found in some cancers), Mps1 inhibition increases the frequency of multipolar mitoses. Lastly, Mps1 inhibitor treatment resulted in a decrease in cancer cell viability.

Small mitochondria-targeting molecules as anti-cancer agents

PubMed Central

Wang, Feng; Ogasawara, Marcia A.; Huang, Peng

2009-01-01

Alterations in mitochondrial structure and functions have long been observed in cancer cells. Targeting mitochondria as a cancer therapeutic strategy has gained momentum in the recent years. The signaling pathways that govern mitochondrial function, apoptosis and molecules that affect mitochondrial integrity and cell viability have been important topics of the recent review in the literature. In this article, we first briefly summarize the rationale and biological basis for developing mitochondrial-targeted compounds as potential anticancer agents, and then provide key examples of small molecules that either directly impact mitochondria or functionally affect the metabolic alterations in cancer cells with mitochondrial dysfunction. The main focus is on the small molecular weight compounds with potential applications in cancer treatment. We also summarize information on the drug developmental stages of the key mitochondria-targeted compounds and their clinical trial status. The advantages and potential shortcomings of targeting the mitochondria for cancer treatment are also discussed. PMID:19995573

Multifunctional and biologically active matrices from multicomponent polymeric solutions

NASA Technical Reports Server (NTRS)

Kiick, Kristi L. (Inventor); Yamaguchi, Nori (Inventor); Rabolt, John (Inventor); Casper, Cheryl (Inventor)

2012-01-01

A functionalized electrospun matrix for the controlled-release of biologically active agents, such as growth factors, is presented. The functionalized matrix comprises a matrix polymer, a compatibilizing polymer and a biomolecule or other small functioning molecule. In certain aspects the electrospun polymer fibers comprise at least one biologically active molecule functionalized with low molecular weight heparin.

Connecting synthetic chemistry decisions to cell and genome biology using small-molecule phenotypic profiling

PubMed Central

Wagner, Bridget K.; Clemons, Paul A.

2009-01-01

Discovering small-molecule modulators for thousands of gene products requires multiple stages of biological testing, specificity evaluation, and chemical optimization. Many cellular profiling methods, including cellular sensitivity, gene-expression, and cellular imaging, have emerged as methods to assess the functional consequences of biological perturbations. Cellular profiling methods applied to small-molecule science provide opportunities to use complex phenotypic information to prioritize and optimize small-molecule structures simultaneously against multiple biological endpoints. As throughput increases and cost decreases for such technologies, we see an emerging paradigm of using more information earlier in probe- and drug-discovery efforts. Moreover, increasing access to public datasets makes possible the construction of “virtual” profiles of small-molecule performance, even when multiplexed measurements were not performed or when multidimensional profiling was not the original intent. We review some key conceptual advances in small-molecule phenotypic profiling, emphasizing connections to other information, such as protein-binding measurements, genetic perturbations, and cell states. We argue that to maximally leverage these measurements in probe and drug discovery requires a fundamental connection to synthetic chemistry, allowing the consequences of synthetic decisions to be described in terms of changes in small-molecule profiles. Mining such data in the context of chemical structure and synthesis strategies can inform decisions about chemistry procurement and library development, leading to optimal small-molecule screening collections. PMID:19825513

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

Aldegunde, Manuel, E-mail: M.A.Aldegunde-Rodriguez@warwick.ac.uk; Kermode, James R., E-mail: J.R.Kermode@warwick.ac.uk; Zabaras, Nicholas

This paper presents the development of a new exchange–correlation functional from the point of view of machine learning. Using atomization energies of solids and small molecules, we train a linear model for the exchange enhancement factor using a Bayesian approach which allows for the quantification of uncertainties in the predictions. A relevance vector machine is used to automatically select the most relevant terms of the model. We then test this model on atomization energies and also on bulk properties. The average model provides a mean absolute error of only 0.116 eV for the test points of the G2/97 set butmore » a larger 0.314 eV for the test solids. In terms of bulk properties, the prediction for transition metals and monovalent semiconductors has a very low test error. However, as expected, predictions for types of materials not represented in the training set such as ionic solids show much larger errors.« less

Potential Impact and Study Considerations of Metabolomics in Cardiovascular Health and Disease: A Scientific Statement From the American Heart Association.

PubMed

Cheng, Susan; Shah, Svati H; Corwin, Elizabeth J; Fiehn, Oliver; Fitzgerald, Robert L; Gerszten, Robert E; Illig, Thomas; Rhee, Eugene P; Srinivas, Pothur R; Wang, Thomas J; Jain, Mohit

2017-04-01

Through the measure of thousands of small-molecule metabolites in diverse biological systems, metabolomics now offers the potential for new insights into the factors that contribute to complex human diseases such as cardiovascular disease. Targeted metabolomics methods have already identified new molecular markers and metabolomic signatures of cardiovascular disease risk (including branched-chain amino acids, select unsaturated lipid species, and trimethylamine- N -oxide), thus in effect linking diverse exposures such as those from dietary intake and the microbiota with cardiometabolic traits. As technologies for metabolomics continue to evolve, the depth and breadth of small-molecule metabolite profiling in complex systems continue to advance rapidly, along with prospects for ongoing discovery. Current challenges facing the field of metabolomics include scaling throughput and technical capacity for metabolomics approaches, bioinformatic and chemoinformatic tools for handling large-scale metabolomics data, methods for elucidating the biochemical structure and function of novel metabolites, and strategies for determining the true clinical relevance of metabolites observed in association with cardiovascular disease outcomes. Progress made in addressing these challenges will allow metabolomics the potential to substantially affect diagnostics and therapeutics in cardiovascular medicine. © 2017 American Heart Association, Inc.

Natural products as an inspiration in the diversity-oriented synthesis of bioactive compound libraries

PubMed Central

Cordier, Christopher; Morton, Daniel; Murrison, Sarah; O'Leary-Steele, Catherine

2008-01-01

The purpose of diversity-oriented synthesis is to drive the discovery of small molecules with previously unknown biological functions. Natural products necessarily populate biologically relevant chemical space, since they bind both their biosynthetic enzymes and their target macromolecules. Natural product families are, therefore, libraries of pre-validated, functionally diverse structures in which individual compounds selectively modulate unrelated macromolecular targets. This review describes examples of diversity-oriented syntheses which have, to some extent, been inspired by the structures of natural products. Particular emphasis is placed on innovations that allow the synthesis of compound libraries that, like natural products, are skeletally diverse. Mimicking the broad structural features of natural products may allow the discovery of compounds that modulate the functions of macromolecules for which ligands are not known. The ability of innovations in diversity-oriented synthesis to deliver such compounds is critically assessed. PMID:18663392

Discovery of a small-molecule binder of the oncoprotein gankyrin that modulates gankyrin activity in the cell

NASA Astrophysics Data System (ADS)

Chattopadhyay, Anasuya; O'Connor, Cornelius J.; Zhang, Fengzhi; Galvagnion, Celine; Galloway, Warren R. J. D.; Tan, Yaw Sing; Stokes, Jamie E.; Rahman, Taufiq; Verma, Chandra; Spring, David R.; Itzhaki, Laura S.

2016-04-01

Gankyrin is an ankyrin-repeat oncoprotein whose overexpression has been implicated in the development of many cancer types. Elevated gankyrin levels are linked to aberrant cellular events including enhanced degradation of tumour suppressor protein p53, and inhibition of gankyrin activity has therefore been identified as an attractive anticancer strategy. Gankyrin interacts with several partner proteins, and a number of these protein-protein interactions (PPIs) are of relevance to cancer. Thus, molecules that bind the PPI interface of gankyrin and interrupt these interactions are of considerable interest. Herein, we report the discovery of a small molecule termed cjoc42 that is capable of binding to gankyrin. Cell-based experiments demonstrate that cjoc42 can inhibit gankyrin activity in a dose-dependent manner: cjoc42 prevents the decrease in p53 protein levels normally associated with high amounts of gankyrin, and it restores p53-dependent transcription and sensitivity to DNA damage. The results represent the first evidence that gankyrin is a “druggable” target with small molecules.

Supercritical-Multiple-Solvent Extraction From Coal

NASA Technical Reports Server (NTRS)

Corcoran, W.; Fong, W.; Pichaichanarong, P.; Chan, P.; Lawson, D.

1983-01-01

Large and small molecules dissolve different constituents. Experimental apparatus used to test supercritical extraction of hydrogen rich compounds from coal in various organic solvents. In decreasing order of importance, relevant process parameters were found to be temperature, solvent type, pressure, and residence time.

Combined small-molecule inhibition accelerates the derivation of functional, early-born, cortical neurons from human pluripotent stem cells

PubMed Central

Qi, Yuchen; Zhang, Xin-Jun; Renier, Nicolas; Wu, Zhuhao; Atkin, Talia; Sun, Ziyi; Ozair, M. Zeeshan; Tchieu, Jason; Zimmer, Bastian; Fattahi, Faranak; Ganat, Yosif; Azevedo, Ricardo; Zeltner, Nadja; Brivanlou, Ali H.; Karayiorgou, Maria; Gogos, Joseph; Tomishima, Mark; Tessier-Lavigne, Marc; Shi, Song-Hai; Studer, Lorenz

2017-01-01

Considerable progress has been made in converting human pluripotent stem cells (hPSCs) into functional neurons. However, the protracted timing of human neuron specification and functional maturation remains a key challenge that hampers the routine application of hPSC-derived lineages in disease modeling and regenerative medicine. Using a combinatorial small-molecule screen, we previously identified conditions for the rapid differentiation of hPSCs into peripheral sensory neurons. Here we generalize the approach to central nervous system (CNS) fates by developing a small-molecule approach for accelerated induction of early-born cortical neurons. Combinatorial application of 6 pathway inhibitors induces post-mitotic cortical neurons with functional electrophysiological properties by day 16 of differentiation, in the absence of glial cell co-culture. The resulting neurons, transplanted at 8 days of differentiation into the postnatal mouse cortex, are functional and establish long-distance projections, as shown using iDISCO whole brain imaging. Accelerated differentiation into cortical neuron fates should facilitate hPSC-based strategies for disease modeling and cell therapy in CNS disorders. PMID:28112759

An ultra-HTS process for the identification of small molecule modulators of orphan G-protein-coupled receptors.

PubMed

Cacace, Angela; Banks, Martyn; Spicer, Timothy; Civoli, Francesca; Watson, John

2003-09-01

G-protein-coupled receptors (GPCRs) are the most successful target proteins for drug discovery research to date. More than 150 orphan GPCRs of potential therapeutic interest have been identified for which no activating ligands or biological functions are known. One of the greatest challenges in the pharmaceutical industry is to link these orphan GPCRs with human diseases. Highly automated parallel approaches that integrate ultra-high throughput and focused screening can be used to identify small molecule modulators of orphan GPCRs. These small molecules can then be employed as pharmacological tools to explore the function of orphan receptors in models of human disease. In this review, we describe methods that utilize powerful ultra-high-throughput screening technologies to identify surrogate ligands of orphan GPCRs.

Preserving Charge and Oxidation State of Au(III) Ions in an Agent-Functionalized Nanocrystal Model System

PubMed Central

2011-01-01

Supporting functional molecules on crystal facets is an established technique in nanotechnology. To preserve the original activity of ionic metallorganic agents on a supporting template, conservation of the charge and oxidation state of the active center is indispensable. We present a model system of a metallorganic agent that, indeed, fulfills this design criterion on a technologically relevant metal support with potential impact on Au(III)-porphyrin-functionalized nanoparticles for an improved anticancer-drug delivery. Employing scanning tunneling microscopy and -spectroscopy in combination with photoemission spectroscopy, we clarify at the single-molecule level the underlying mechanisms of this exceptional adsorption mode. It is based on the balance between a high-energy oxidation state and an electrostatic screening-response of the surface (image charge). Modeling with first principles methods reveals submolecular details of the metal–ligand bonding interaction and completes the study by providing an illustrative electrostatic model relevant for ionic metalorganic agent molecules, in general. PMID:21736315

Development of a Unique Small Molecule Modulator of CXCR4

PubMed Central

Yoon, Younghyoun; Lin, Songbai; Sasaki, Maiko; Klapproth, Jan-Michael A.; Yang, Hua; Grossniklaus, Hans E.; Xu, Jianguo; Rojas, Mauricio; Voll, Ronald J.; Goodman, Mark M.; Arrendale, Richard F.; Liu, Jin; Yun, C. Chris; Snyder, James P.; Liotta, Dennis C.; Shim, Hyunsuk

2012-01-01

Background Metastasis, the spread and growth of tumor cells to distant organ sites, represents the most devastating attribute and plays a major role in the morbidity and mortality of cancer. Inflammation is crucial for malignant tumor transformation and survival. Thus, blocking inflammation is expected to serve as an effective cancer treatment. Among anti-inflammation therapies, chemokine modulation is now beginning to emerge from the pipeline. CXC chemokine receptor-4 (CXCR4) and its ligand stromal cell-derived factor-1 (CXCL12) interaction and the resulting cell signaling cascade have emerged as highly relevant targets since they play pleiotropic roles in metastatic progression. The unique function of CXCR4 is to promote the homing of tumor cells to their microenvironment at the distant organ sites. Methodology/Principal Findings We describe the actions of N,N′-(1,4-phenylenebis(methylene))dipyrimidin-2-amine (designated MSX-122), a novel small molecule and partial CXCR4 antagonist with properties quite unlike that of any other reported CXCR4 antagonists, which was prepared in a single chemical step using a reductive amination reaction. Its specificity toward CXCR4 was tested in a binding affinity assay and a ligand competition assay using 18F-labeled MSX-122. The potency of the compound was determined in two functional assays, Matrigel invasion assay and cAMP modulation. The therapeutic potential of MSX-122 was evaluated in three different murine models for inflammation including an experimental colitis, carrageenan induced paw edema, and bleomycin induced lung fibrosis and three different animal models for metastasis including breast cancer micrometastasis in lung, head and neck cancer metastasis in lung, and uveal melanoma micrometastasis in liver in which CXCR4 was reported to play crucial roles. Conclusions/Significance We developed a novel small molecule, MSX-122, that is a partial CXCR4 antagonist without mobilizing stem cells, which can be safer for long-term blockade of metastasis than other reported CXCR4 antagonists. PMID:22485156

Combinatorics of feedback in cellular uptake and metabolism of small molecules.

PubMed

Krishna, Sandeep; Semsey, Szabolcs; Sneppen, Kim

2007-12-26

We analyze the connection between structure and function for regulatory motifs associated with cellular uptake and usage of small molecules. Based on the boolean logic of the feedback we suggest four classes: the socialist, consumer, fashion, and collector motifs. We find that the socialist motif is good for homeostasis of a useful but potentially poisonous molecule, whereas the consumer motif is optimal for nutrition molecules. Accordingly, examples of these motifs are found in, respectively, the iron homeostasis system in various organisms and in the uptake of sugar molecules in bacteria. The remaining two motifs have no obvious analogs in small molecule regulation, but we illustrate their behavior using analogies to fashion and obesity. These extreme motifs could inspire construction of synthetic systems that exhibit bistable, history-dependent states, and homeostasis of flux (rather than concentration).

One and two hydrogen molecules in the large cage of the structure II clathrate hydrate: quantum translation-rotation dynamics close to the cage wall.

PubMed

Sebastianelli, Francesco; Xu, Minzhong; Kanan, Dalal K; Bacić, Zlatko

2007-07-19

We have performed a rigorous theoretical study of the quantum translation-rotation (T-R) dynamics of one and two H2 and D2 molecules confined inside the large hexakaidecahedral (5(12)6(4)) cage of the sII clathrate hydrate. For a single encapsulated H2 and D2 molecule, accurate quantum five-dimensional calculations of the T-R energy levels and wave functions are performed that include explicitly, as fully coupled, all three translational and the two rotational degrees of freedom of the hydrogen molecule, while the cage is taken to be rigid. In addition, the ground-state properties, energetics, and spatial distribution of one and two p-H2 and o-D2 molecules in the large cage are calculated rigorously using the diffusion Monte Carlo method. These calculations reveal that the low-energy T-R dynamics of hydrogen molecules in the large cage are qualitatively different from that inside the small cage, studied by us recently. This is caused by the following: (i) The large cage has a cavity whose diameter is about twice that of the small cage for the hydrogen molecule. (ii) In the small cage, the potential energy surface (PES) for H2 is essentially flat in the central region, while in the large cage the PES has a prominent maximum at the cage center, whose height exceeds the T-R zero-point energy of H2/D2. As a result, the guest molecule is excluded from the central part of the large cage, its wave function localized around the off-center global minimum. Peculiar quantum dynamics of the hydrogen molecule squeezed between the central maximum and the cage wall manifests in the excited T-R states whose energies and wave functions differ greatly from those for the small cage. Moreover, they are sensitive to the variations in the hydrogen-bonding topology, which modulate the corrugation of the cage wall.

Generalized extracellular molecule sensor platform for programming cellular behavior.

PubMed

Scheller, Leo; Strittmatter, Tobias; Fuchs, David; Bojar, Daniel; Fussenegger, Martin

2018-04-23

Strategies for expanding the sensor space of designer receptors are urgently needed to tailor cell-based therapies to respond to any type of medically relevant molecules. Here, we describe a universal approach to designing receptor scaffolds that enables antibody-specific molecular input to activate JAK/STAT, MAPK, PLCG or PI3K/Akt signaling rewired to transgene expression driven by synthetic promoters. To demonstrate its scope, we equipped the GEMS (generalized extracellular molecule sensor) platform with antibody fragments targeting a synthetic azo dye, nicotine, a peptide tag and the PSA (prostate-specific antigen) biomarker, thereby covering inputs ranging from small molecules to proteins. These four GEMS devices provided robust signaling and transgene expression with high signal-to-noise ratios in response to their specific ligands. The sensitivity of the nicotine- and PSA-specific GEMS devices matched the clinically relevant concentration ranges, and PSA-specific GEMS were able to detect pathological PSA levels in the serum of patients diagnosed with prostate cancer.

The fabrication of small molecule organic light-emitting diode pixels by laser-induced forward transfer

NASA Astrophysics Data System (ADS)

Shaw-Stewart, J. R. H.; Mattle, T.; Lippert, T. K.; Nagel, M.; Nüesch, F. A.; Wokaun, A.

2013-01-01

Laser-induced forward transfer (LIFT) is a versatile organic light-emitting diode (OLED) pixel deposition process, but has hitherto been applied exclusively to polymeric materials. Here, a modified LIFT process has been used to fabricate small molecule Alq3 organic light-emitting diodes (SMOLEDs). Small molecule thin films are considerably more mechanically brittle than polymeric thin films, which posed significant challenges for LIFT of these materials. The LIFT process presented here uses a polymeric dynamic release layer, a reduced environmental pressure, and a well-defined receiver-donor gap. The Alq3 pixels demonstrate good morphology and functionality, even when compared to conventionally fabricated OLEDs. The Alq3 SMOLED pixel performances show a significant amount of fluence dependence, not observed with polymerical OLED pixels made in previous studies. A layer of tetrabutyl ammonium hydroxide has been deposited on top of the aluminium cathode, as part of the donor substrate, to improve electron injection to the Alq3, by over 600%. These results demonstrate that this variant of LIFT is applicable for the deposition of functional small molecule OLEDs as well as polymeric OLEDs.

Design of a bioactive small molecule that targets r(AUUCU) repeats in spinocerebellar ataxia 10.

PubMed

Yang, Wang-Yong; Gao, Rui; Southern, Mark; Sarkar, Partha S; Disney, Matthew D

2016-06-01

RNA is an important target for chemical probes of function and lead therapeutics; however, it is difficult to target with small molecules. One approach to tackle this problem is to identify compounds that target RNA structures and utilize them to multivalently target RNA. Here we show that small molecules can be identified to selectively bind RNA base pairs by probing a library of RNA-focused small molecules. A small molecule that selectively binds AU base pairs informed design of a dimeric compound (2AU-2) that targets the pathogenic RNA, expanded r(AUUCU) repeats, that causes spinocerebellar ataxia type 10 (SCA10) in patient-derived cells. Indeed, 2AU-2 (50 nM) ameliorates various aspects of SCA10 pathology including improvement of mitochondrial dysfunction, reduced activation of caspase 3, and reduction of nuclear foci. These studies provide a first-in-class chemical probe to study SCA10 RNA toxicity and potentially define broadly applicable compounds targeting RNA AU base pairs in cells.

Small molecules enhance CRISPR genome editing in pluripotent stem cells.

PubMed

Yu, Chen; Liu, Yanxia; Ma, Tianhua; Liu, Kai; Xu, Shaohua; Zhang, Yu; Liu, Honglei; La Russa, Marie; Xie, Min; Ding, Sheng; Qi, Lei S

2015-02-05

The bacterial CRISPR-Cas9 system has emerged as an effective tool for sequence-specific gene knockout through non-homologous end joining (NHEJ), but it remains inefficient for precise editing of genome sequences. Here we develop a reporter-based screening approach for high-throughput identification of chemical compounds that can modulate precise genome editing through homology-directed repair (HDR). Using our screening method, we have identified small molecules that can enhance CRISPR-mediated HDR efficiency, 3-fold for large fragment insertions and 9-fold for point mutations. Interestingly, we have also observed that a small molecule that inhibits HDR can enhance frame shift insertion and deletion (indel) mutations mediated by NHEJ. The identified small molecules function robustly in diverse cell types with minimal toxicity. The use of small molecules provides a simple and effective strategy to enhance precise genome engineering applications and facilitates the study of DNA repair mechanisms in mammalian cells. Copyright © 2015 Elsevier Inc. All rights reserved.

Controlling the Morphology of BDTT-DPP-Based Small Molecules via End-Group Functionalization for Highly Efficient Single and Tandem Organic Photovoltaic Cells.

PubMed

Kim, Ji-Hoon; Park, Jong Baek; Yang, Hoichang; Jung, In Hwan; Yoon, Sung Cheol; Kim, Dongwook; Hwang, Do-Hoon

2015-11-04

A series of narrow-band gap, π-conjugated small molecules based on diketopyrrolopyrrole (DPP) electron acceptor units coupled with alkylthienyl-substituted-benzodithiophene (BDTT) electron donors were designed and synthesized for use as donor materials in solution-processed organic photovoltaic cells. In particular, by end-group functionalization of the small molecules with fluorine derivatives, the nanoscale morphologies of the photoactive layers of the photovoltaic cells were successfully controlled. The influences of different fluorine-based end-groups on the optoelectronic and morphological properties, carrier mobilities, and the photovoltaic performances of these materials were investigated. A high power conversion efficiency (PCE) of 6.00% under simulated solar light (AM 1.5G) illumination has been achieved for organic photovoltaic cells based on a small-molecule bulk heterojunction system consisting of a trifluoromethylbenzene (CF3) end-group-containing oligomer (BDTT-(DPP)2-CF3) as the donor and [6,6]-phenyl-C71-butyric acid methyl ester (PC71BM) as the acceptor. As a result, the introduction of CF3 end-groups has been found to enhance both the short circuit current density (JSC) and fill factor (FF). A tandem photovoltaic device comprising an inverted BDTT-(DPP)2-CF3:PC71BM cell and a poly(3-hexylthiophene) (P3HT):indene-C60-bisadduct (IC60BA)-based cell as the top and bottom cell components, respectively, showed a maximum PCE of 8.30%. These results provide valuable guidelines for the rational design of conjugated small molecules for applications in high-performance organic photovoltaic cells. Furthermore, to the best of our knowledge, this is the first report on the design of fluorine-functionalized BDTT-DPP-based small molecules, which have been shown to be a viable candidate for use in inverted tandem cells.

Computer Simulations of Small Molecules in Membranes: Insights from Computer Simulations into the Interactions of Small Molecules with Lipid Bilayers

NASA Technical Reports Server (NTRS)

Pohorille, Andrew; New, Michael H.; Schweighofer, Karl; Wilson, Michael A.; DeVincenzi, Donald L. (Technical Monitor)

2000-01-01

Two of Ernest Overton's lasting contributions to biology are the Meyer-Overton relationship between the potency of an anesthetic and its solubility in oil, and the Overton rule which relates the permeability of a membrane to the oil-water partition coefficient of the permeating molecule. A growing body of experimental evidence, however, cannot be reconciled with these theories. In particular, the molecular nature of membranes, unknown to Overton, needs to be included in any description of these phenomena. Computer simulations are ideally suited for providing atomic-level information about the behavior of small molecules in membranes. The authors discuss simulation studies relevant to Overton's ideas. Through simulations it was found that anesthetics tend to concentrate at interfaces and their anesthetic potency correlates better with solubility at the water-membrane interface than with solubility in oil. Simulation studies of membrane permeation revealed the anisotropic nature of the membranes, as evidenced, for example, by the highly nonuniform distribution of free volume in the bilayer. This, in turn, influences the diffusion rates of solutes, which increase with the depth in the membrane. Small solutes tend to move by hopping between voids in the bilayer, and this hopping motion may be responsible for the deviation from the Overton rule of the permeation rates of these molecules.

Chemical wiring and soldering toward all-molecule electronic circuitry.

PubMed

Okawa, Yuji; Mandal, Swapan K; Hu, Chunping; Tateyama, Yoshitaka; Goedecker, Stefan; Tsukamoto, Shigeru; Hasegawa, Tsuyoshi; Gimzewski, James K; Aono, Masakazu

2011-06-01

Key to single-molecule electronics is connecting functional molecules to each other using conductive nanowires. This involves two issues: how to create conductive nanowires at designated positions, and how to ensure chemical bonding between the nanowires and functional molecules. Here, we present a novel method that solves both issues. Relevant functional molecules are placed on a self-assembled monolayer of diacetylene compound. A probe tip of a scanning tunneling microscope is then positioned on the molecular row of the diacetylene compound to which the functional molecule is adsorbed, and a conductive polydiacetylene nanowire is fabricated by initiating chain polymerization by stimulation with the tip. Since the front edge of chain polymerization necessarily has a reactive chemical species, the created polymer nanowire forms chemical bonding with an encountered molecular element. We name this spontaneous reaction "chemical soldering". First-principles theoretical calculations are used to investigate the structures and electronic properties of the connection. We demonstrate that two conductive polymer nanowires are connected to a single phthalocyanine molecule. A resonant tunneling diode formed by this method is discussed. © 2011 American Chemical Society

Proteome-wide covalent ligand discovery in native biological systems

PubMed Central

Backus, Keriann M.; Correia, Bruno E.; Lum, Kenneth M.; Forli, Stefano; Horning, Benjamin D.; González-Páez, Gonzalo E.; Chatterjee, Sandip; Lanning, Bryan R.; Teijaro, John R.; Olson, Arthur J.; Wolan, Dennis W.; Cravatt, Benjamin F.

2016-01-01

Small molecules are powerful tools for investigating protein function and can serve as leads for new therapeutics. Most human proteins, however, lack small-molecule ligands, and entire protein classes are considered “undruggable” 1,2. Fragment-based ligand discovery (FBLD) can identify small-molecule probes for proteins that have proven difficult to target using high-throughput screening of complex compound libraries 1,3. Although reversibly binding ligands are commonly pursued, covalent fragments provide an alternative route to small-molecule probes 4–10, including those that can access regions of proteins that are difficult to access through binding affinity alone 5,10,11. In this manuscript, we report a quantitative analysis of cysteine-reactive small-molecule fragments screened against thousands of proteins. Covalent ligands were identified for >700 cysteines found in both druggable proteins and proteins deficient in chemical probes, including transcription factors, adaptor/scaffolding proteins, and uncharacterized proteins. Among the atypical ligand-protein interactions discovered were compounds that react preferentially with pro- (inactive) caspases. We used these ligands to distinguish extrinsic apoptosis pathways in human cell lines versus primary human T-cells, showing that the former is largely mediated by caspase-8 while the latter depends on both caspase-8 and −10. Fragment-based covalent ligand discovery provides a greatly expanded portrait of the ligandable proteome and furnishes compounds that can illuminate protein functions in native biological systems. PMID:27309814

Chemicals as the Sole Transformers of Cell Fate.

PubMed

Ebrahimi, Behnam

2016-05-30

Forced expression of lineage-specific transcription factors in somatic cells can result in the generation of different cell types in a process named direct reprogramming, bypassing the pluripotent state. However, the introduction of transgenes limits the therapeutic applications of the produced cells. Numerous small-molecules have been introduced in the field of stem cell biology capable of governing self-renewal, reprogramming, transdifferentiation and regeneration. These chemical compounds are versatile tools for cell fate conversion toward desired outcomes. Cell fate conversion using small-molecules alone (chemical reprogramming) has superiority over arduous traditional genetic techniques in several aspects. For instance, rapid, transient, and reversible effects in activation and inhibition of functions of specific proteins are of the profits of small-molecules. They are cost-effective, have a long half-life, diversity on structure and function, and allow for temporal and flexible regulation of signaling pathways. Additionally, their effects could be adjusted by fine-tuning concentrations and combinations of different small-molecules. Therefore, chemicals are powerful tools in cell fate conversion and study of stem cell and chemical biology in vitro and in vivo. Moreover, transgene-free and chemical-only transdifferentiation approaches provide alternative strategies for the generation of various cell types, disease modeling, drug screening, and regenerative medicine. The current review gives an overview of the recent findings concerning transdifferentiation by only small-molecules without the use of transgenes.

Second and third order nonlinear optical properties of conjugated molecules and polymers

NASA Technical Reports Server (NTRS)

Perry, Joseph W.; Stiegman, Albert E.; Marder, Seth R.; Coulter, Daniel R.; Beratan, David N.; Brinza, David E.

1988-01-01

Second- and third-order nonlinear optical properties of some newly synthesized organic molecules and polymers are reported. Powder second-harmonic-generation efficiencies of up to 200 times urea have been realized for asymmetric donor-acceptor acetylenes. Third harmonic generation chi(3)s have been determined for a series of small conjugated molecules in solution. THG chi(3)s have also been determined for a series of soluble conjugated copolymers prepared using ring-opening metathesis polymerization. The results are discussed in terms of relevant molecular and/or macroscopic structural features of these conjugated organic materials.

Allosteric Communication Disrupted by a Small Molecule Binding to the Imidazole Glycerol Phosphate Synthase Protein-Protein Interface.

PubMed

Rivalta, Ivan; Lisi, George P; Snoeberger, Ning-Shiuan; Manley, Gregory; Loria, J Patrick; Batista, Victor S

2016-11-29

Allosteric enzymes regulate a wide range of catalytic transformations, including biosynthetic mechanisms of important human pathogens, upon binding of substrate molecules to an orthosteric (or active) site and effector ligands at distant (allosteric) sites. We find that enzymatic activity can be impaired by small molecules that bind along the allosteric pathway connecting the orthosteric and allosteric sites, without competing with endogenous ligands. Noncompetitive allosteric inhibitors disrupted allostery in the imidazole glycerol phosphate synthase (IGPS) enzyme from Thermotoga maritima as evidenced by nuclear magnetic resonance, microsecond time-scale molecular dynamics simulations, isothermal titration calorimetry, and kinetic assays. The findings are particularly relevant for the development of allosteric antibiotics, herbicides, and antifungal compounds because IGPS is absent in mammals but provides an entry point to fundamental biosynthetic pathways in plants, fungi, and bacteria.

Aggregation-induced chemical reactions: acid dissociation in growing water clusters.

PubMed

Forbert, Harald; Masia, Marco; Kaczmarek-Kedziera, Anna; Nair, Nisanth N; Marx, Dominik

2011-03-23

Understanding chemical reactivity at ultracold conditions, thus enabling molecular syntheses via interstellar and atmospheric processes, is a key issue in cryochemistry. In particular, acid dissociation and proton transfer reactions are ubiquitous in aqueous microsolvation environments. Here, the full dissociation of a HCl molecule upon stepwise solvation by a small number of water molecules at low temperatures, as relevant to helium nanodroplet isolation (HENDI) spectroscopy, is analyzed in mechanistic detail. It is found that upon successive aggregation of HCl with H(2)O molecules, a series of cyclic heteromolecular structures, up to and including HCl(H(2)O)(3), are initially obtained before a precursor state for dissociation, HCl(H(2)O)(3)···H(2)O, is observed upon addition of a fourth water molecule. The latter partially aggregated structure can be viewed as an "activated species", which readily leads to dissociation of HCl and to the formation of a solvent-shared ion pair, H(3)O(+)(H(2)O)(3)Cl(-). Overall, the process is mostly downhill in potential energy, and, in addition, small remaining barriers are overcome by using kinetic energy released as a result of forming hydrogen bonds due to aggregation. The associated barrier is not ruled by thermal equilibrium but is generated by athermal non-equilibrium dynamics. These "aggregation-induced chemical reactions" are expected to be of broad relevance to chemistry at ultralow temperature much beyond HENDI spectroscopy.

Biological and functional relevance of CASP predictions.

PubMed

Liu, Tianyun; Ish-Shalom, Shirbi; Torng, Wen; Lafita, Aleix; Bock, Christian; Mort, Matthew; Cooper, David N; Bliven, Spencer; Capitani, Guido; Mooney, Sean D; Altman, Russ B

2018-03-01

Our goal is to answer the question: compared with experimental structures, how useful are predicted models for functional annotation? We assessed the functional utility of predicted models by comparing the performances of a suite of methods for functional characterization on the predictions and the experimental structures. We identified 28 sites in 25 protein targets to perform functional assessment. These 28 sites included nine sites with known ligand binding (holo-sites), nine sites that are expected or suggested by experimental authors for small molecule binding (apo-sites), and Ten sites containing important motifs, loops, or key residues with important disease-associated mutations. We evaluated the utility of the predictions by comparing their microenvironments to the experimental structures. Overall structural quality correlates with functional utility. However, the best-ranked predictions (global) may not have the best functional quality (local). Our assessment provides an ability to discriminate between predictions with high structural quality. When assessing ligand-binding sites, most prediction methods have higher performance on apo-sites than holo-sites. Some servers show consistently high performance for certain types of functional sites. Finally, many functional sites are associated with protein-protein interaction. We also analyzed biologically relevant features from the protein assemblies of two targets where the active site spanned the protein-protein interface. For the assembly targets, we find that the features in the models are mainly determined by the choice of template. © 2017 The Authors Proteins: Structure, Function and Bioinformatics Published by Wiley Periodicals, Inc.

Identification of Small Molecule Activators of Cryptochrome

PubMed Central

Hirota, Tsuyoshi; Lee, Jae Wook; St. John, Peter C.; Sawa, Mariko; Iwaisako, Keiko; Noguchi, Takako; Pongsawakul, Pagkapol Y.; Sonntag, Tim; Welsh, David K.; Brenner, David A.; Doyle, Francis J.; Schultz, Peter G.; Kay, Steve A.

2013-01-01

Impairment of the circadian clock has been associated with numerous disorders, including metabolic disease. Although small molecules that modulate clock function might offer therapeutic approaches to such diseases, only a few compound have been identified that selectively target core clock proteins. From an unbiased cell-based circadian screen, we identified KL001, a small molecule that specifically interacts with cryptochrome (CRY). KL001 prevented ubiquitin-dependent degradation of CRY, resulting in lengthening of the circadian period. In combination with mathematical modeling, KL001 revealed that CRY1 and CRY2 share a similar functional role in the period regulation. Furthermore, KL001- mediated CRY stabilization inhibited glucagon-induced gluconeogenesis in primary hepatocytes. KL001 thus provides a tool to study the regulation of CRY-dependent physiology and aid development of clock-based therapeutics of diabetes. PMID:22798407

Molecular trade-offs in RNA ligases affected the modular emergence of complex ribozymes at the origin of life

PubMed Central

Weinberg, Marc S.; Michod, Richard E.

2017-01-01

In the RNA world hypothesis complex, self-replicating ribozymes were essential. For the emergence of an RNA world, less is known about the early processes that accounted for the formation of complex, long catalysts from small passively formed molecules. The functional role of small sequences has not been fully explored and, here, a possible role for smaller ligases is demonstrated. An established RNA polymerase model, the R18, was truncated from the 3′ end to generate smaller molecules. All the molecules were investigated for self-ligation functions with a set of oligonucleotide substrates without predesigned base pairing. The smallest molecule that exhibited self-ligation activity was a 40-nucleotide RNA. It also demonstrated the greatest functional flexibility as it was more general in the kinds of substrates it ligated to itself although its catalytic efficiency was the lowest. The largest ribozyme (R18) ligated substrates more selectively and with greatest efficiency. With increase in size and predicted structural stability, self-ligation efficiency improved, while functional flexibility decreased. These findings reveal that molecular size could have increased from the activity of small ligases joining oligonucleotides to their own end. In addition, there is a size-associated molecular-level trade-off that could have impacted the evolution of RNA-based life. PMID:28989747

Molecular trade-offs in RNA ligases affected the modular emergence of complex ribozymes at the origin of life

NASA Astrophysics Data System (ADS)

Dhar, Nisha; Weinberg, Marc S.; Michod, Richard E.; Durand, Pierre M.

2017-09-01

In the RNA world hypothesis complex, self-replicating ribozymes were essential. For the emergence of an RNA world, less is known about the early processes that accounted for the formation of complex, long catalysts from small passively formed molecules. The functional role of small sequences has not been fully explored and, here, a possible role for smaller ligases is demonstrated. An established RNA polymerase model, the R18, was truncated from the 3' end to generate smaller molecules. All the molecules were investigated for self-ligation functions with a set of oligonucleotide substrates without predesigned base pairing. The smallest molecule that exhibited self-ligation activity was a 40-nucleotide RNA. It also demonstrated the greatest functional flexibility as it was more general in the kinds of substrates it ligated to itself although its catalytic efficiency was the lowest. The largest ribozyme (R18) ligated substrates more selectively and with greatest efficiency. With increase in size and predicted structural stability, self-ligation efficiency improved, while functional flexibility decreased. These findings reveal that molecular size could have increased from the activity of small ligases joining oligonucleotides to their own end. In addition, there is a size-associated molecular-level trade-off that could have impacted the evolution of RNA-based life.

Structures, electronic properties and reaction paths from Fe(CO)5 molecule to small Fe clusters

NASA Astrophysics Data System (ADS)

Li, Zhi; Zhao, Zhen

2018-04-01

The geometries, electrical characters and reaction paths from Fe(CO)5 molecule to small Fe clusters were investigated by using all-electron density functional theory. The results show that in the decomposition process of pentacarbonyl-iron, Fe(CO)5 molecule prefers to remove a carbon monoxide and adsorb another Fe(CO)5 molecule to produce nonacarbonyldiiron Fe2(CO)9 then Fe2(CO)9 gradually removes carbon monoxide to produce small Fe clusters. As It can be seen from the highest occupied molecule orbital-lowest unoccupied molecule orbital gap curves, the Fe(CO)n=3, and 5 and Fe2(CO)n=3, 7 and 9 intermediates have higher chemical stability than their neighbors. The local magnetic moment of the carbon monoxide is aligning anti-ferromagnetic. The effect of external magnetic field to the initial decomposition products of Fe(CO)5 can be ignored.

A Synthetic DNA-Binding Domain Guides Distinct Chromatin-Modifying Small Molecules to Activate an Identical Gene Network.

PubMed

Han, Le; Pandian, Ganesh N; Chandran, Anandhakumar; Sato, Shinsuke; Taniguchi, Junichi; Kashiwazaki, Gengo; Sawatani, Yoshito; Hashiya, Kaori; Bando, Toshikazu; Xu, Yufang; Qian, Xuhong; Sugiyama, Hiroshi

2015-07-20

Synthetic dual-function ligands targeting specific DNA sequences and histone-modifying enzymes were applied to achieve regulatory control over multi-gene networks in living cells. Unlike the broad array of targeting small molecules for histone deacetylases (HDACs), few modulators are known for histone acetyltransferases (HATs), which play a central role in transcriptional control. As a novel chemical approach to induce selective HAT-regulated genes, we conjugated a DNA-binding domain (DBD) "I" to N-(4-chloro-3-trifluoromethyl-phenyl)-2-ethoxy-benzamide (CTB), an artificial HAT activator. In vitro enzyme activity assays and microarray studies were used to demonstrate that distinct functional small molecules could be transformed to have identical bioactivity when conjugated with a targeting DBD. This proof-of-concept synthetic strategy validates the switchable functions of HDACs and HATs in gene regulation and provides a molecular basis for developing versatile bioactive ligands. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Evolution of the physicochemical properties of marketed drugs: can history foretell the future?

PubMed

Faller, Bernard; Ottaviani, Giorgio; Ertl, Peter; Berellini, Giuliano; Collis, Alan

2011-11-01

A set of diverse bioactive molecules, relevant from a medicinal chemistry viewpoint, was assembled and used to navigate the physicochemical property space of new and old, or traditional drugs against a larger set of 12,000 diverse bioactive small molecules. Most drugs on the market only occupy a fraction of the property space of the bioactive molecules, whereas new molecular entities (NMEs) approved since 2002 are moving away from this traditional drug space. In this new territory, semi-empirical rules derived from knowledge accumulated from historic, older molecules are not necessarily valid and different liabilities become more prominent. Copyright © 2011 Elsevier Ltd. All rights reserved.

Cellular manganese content is developmentally regulated in human dopaminergic neurons

NASA Astrophysics Data System (ADS)

Kumar, Kevin K.; Lowe, Edward W., Jr.; Aboud, Asad A.; Neely, M. Diana; Redha, Rey; Bauer, Joshua A.; Odak, Mihir; Weaver, C. David; Meiler, Jens; Aschner, Michael; Bowman, Aaron B.

2014-10-01

Manganese (Mn) is both an essential biological cofactor and neurotoxicant. Disruption of Mn biology in the basal ganglia has been implicated in the pathogenesis of neurodegenerative disorders, such as parkinsonism and Huntington's disease. Handling of other essential metals (e.g. iron and zinc) occurs via complex intracellular signaling networks that link metal detection and transport systems. However, beyond several non-selective transporters, little is known about the intracellular processes regulating neuronal Mn homeostasis. We hypothesized that small molecules that modulate intracellular Mn could provide insight into cell-level Mn regulatory mechanisms. We performed a high throughput screen of 40,167 small molecules for modifiers of cellular Mn content in a mouse striatal neuron cell line. Following stringent validation assays and chemical informatics, we obtained a chemical `toolbox' of 41 small molecules with diverse structure-activity relationships that can alter intracellular Mn levels under biologically relevant Mn exposures. We utilized this toolbox to test for differential regulation of Mn handling in human floor-plate lineage dopaminergic neurons, a lineage especially vulnerable to environmental Mn exposure. We report differential Mn accumulation between developmental stages and stage-specific differences in the Mn-altering activity of individual small molecules. This work demonstrates cell-level regulation of Mn content across neuronal differentiation.

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

Matz, Dallas L.; Schalnat, Matthew C.; Pemberton, Jeanne E.

The reaction between small organic molecules and low work function metals is of interest in organometallic, astronomical, and optoelectronic device chemistry. Here, thin, solid-state, amorphous benzene and pyridine films are reacted with Ca at 30 K under ultrahigh vacuum with the reaction progress monitored by Raman spectroscopy. Although both films react with Ca to produce product species identifiable by their vibrational spectroscopic signatures, benzene is less reactive with Ca than pyridine. Benzene reacts by electron transfer from Ca to benzene producing multiple species including the phenyl radical anion, the phenyl radical, and the benzyne diradical. Pyridine initially reacts along amore » similar electron transfer pathway as indicated by the presence of the corresponding pyridyl radical and pyridyne diradical species, but these pyridyl radicals are less stable and subject to further ring-opening reactions that lead to a complex array of smaller molecule reaction products and ultimately amorphous carbon. The elucidation of this reaction pathway provides insight into the reactions of aromatics with Ca that are relevant in the areas of catalysis, astrochemistry, and organic optoelectronics.« less

Chemical Modification of the Multi-Target Neuroprotective Compound Fisetin

PubMed Central

Chiruta, Chandramouli; Schubert, David; Dargusch, Richard; Maher, Pamela

2012-01-01

Many factors are implicated in age-related CNS disorders making it unlikely that modulating only a single factor will provide effective treatment. Perhaps a better approach is to identify small molecules that have multiple biological activities relevant to the maintenance of brain function. Recently, we identified an orally active, neuroprotective and cognition-enhancing molecule, the flavonoid fisetin, that is effective in several animal models of CNS disorders. Fisetin has direct antioxidant activity and can also increase the intracellular levels of glutathione (GSH), the major endogenous antioxidant. In addition, fisetin has both neurotrophic and anti-inflammatory activity. However, its relatively high EC50 in cell based assays, low lipophilicity, high tPSA and poor bioavailability suggest that there is room for medicinal chemical improvement. Here we describe a multi-tiered approach to screening that has allowed us to identify fisetin derivatives with significantly enhanced activity in an in vitro neuroprotection model while at the same time maintaining other key activities. PMID:22192055

Biased and unbiased strategies to identify biologically active small molecules.

PubMed

Abet, Valentina; Mariani, Angelica; Truscott, Fiona R; Britton, Sébastien; Rodriguez, Raphaël

2014-08-15

Small molecules are central players in chemical biology studies. They promote the perturbation of cellular processes underlying diseases and enable the identification of biological targets that can be validated for therapeutic intervention. Small molecules have been shown to accurately tune a single function of pluripotent proteins in a reversible manner with exceptional temporal resolution. The identification of molecular probes and drugs remains a worthy challenge that can be addressed by the use of biased and unbiased strategies. Hypothesis-driven methodologies employs a known biological target to synthesize complementary hits while discovery-driven strategies offer the additional means of identifying previously unanticipated biological targets. This review article provides a general overview of recent synthetic frameworks that gave rise to an impressive arsenal of biologically active small molecules with unprecedented cellular mechanisms. Copyright © 2014. Published by Elsevier Ltd.

Micro and Nano Material Carriers for Immunomodulation

PubMed Central

Bracho-Sanchez, Evelyn; Xia, Chang Qing; Clare-Salzler, Michael J.; Keselowsky, Benjamin G.

2016-01-01

Modulation of the immune system through the use of micro and nano carriers offers opportunities in transplant tolerance, autoimmunity, infectious disease and cancer. In particular, polymeric, lipid and inorganic materials have been used as carriers of proteins, nucleic acids, and small drug molecules to direct the immune system toward either suppressive or stimulatory states. Current technologies have focused on the use of particulates or scaffolds, the modulation of materials properties, and the delivery of biologics or small drug molecules to achieve a desired response. Discussed are relevant immunology concepts, the types of biomaterial-carriers used for immunomodulation highlighting their benefits and drawbacks, the material properties influencing immune responses, and recent examples in the field of transplant tolerance. PMID:27214679

Identification of C3b-Binding Small-Molecule Complement Inhibitors Using Cheminformatics.

PubMed

Garcia, Brandon L; Skaff, D Andrew; Chatterjee, Arindam; Hanning, Anders; Walker, John K; Wyckoff, Gerald J; Geisbrecht, Brian V

2017-05-01

The complement system is an elegantly regulated biochemical cascade formed by the collective molecular recognition properties and proteolytic activities of more than two dozen membrane-bound or serum proteins. Complement plays diverse roles in human physiology, such as acting as a sentry against invading microorganisms, priming of the adaptive immune response, and removal of immune complexes. However, dysregulation of complement can serve as a trigger for a wide range of human diseases, which include autoimmune, inflammatory, and degenerative conditions. Despite several potential advantages of modulating complement with small-molecule inhibitors, small-molecule drugs are highly underrepresented in the current complement-directed therapeutics pipeline. In this study, we have employed a cheminformatics drug discovery approach based on the extensive structural and functional knowledge available for the central proteolytic fragment of the cascade, C3b. Using parallel in silico screening methodologies, we identified 45 small molecules that putatively bind C3b near ligand-guided functional hot spots. Surface plasmon resonance experiments resulted in the validation of seven dose-dependent C3b-binding compounds. Competition-based biochemical assays demonstrated the ability of several C3b-binding compounds to interfere with binding of the original C3b ligand that guided their discovery. In vitro assays of complement function identified a single complement inhibitory compound, termed cmp-5, and mechanistic studies of the cmp-5 inhibitory mode revealed it acts at the level of C5 activation. This study has led to the identification of a promising new class of C3b-binding small-molecule complement inhibitors and, to our knowledge, provides the first demonstration of cheminformatics-based, complement-directed drug discovery. Copyright © 2017 by The American Association of Immunologists, Inc.

Identification of C3b-binding Small Molecule Complement Inhibitors Using Cheminformatics

PubMed Central

Garcia, Brandon L.; Skaff, D. Andrew; Chatterjee, Arindam; Hanning, Anders; Walker, John K.; Wyckoff, Gerald J.; Geisbrecht, Brian V.

2017-01-01

The complement system is an elegantly regulated biochemical cascade formed by the collective molecular recognition properties and proteolytic activities of over two dozen membrane-bound or serum proteins. Complement plays diverse roles in human physiology which include acting as a sentry against invading microorganisms, priming of the adaptive immune response, and removal of immune complexes. However, dysregulation of complement can serve as a trigger for a wide range of human diseases which include autoimmune, inflammatory, and degenerative conditions. Despite several potential advantages of modulating complement with small molecule inhibitors, small molecule drugs are highly underrepresented in the current complement-directed therapeutics pipeline. In this study we have employed a cheminformatics drug discovery approach based on the extensive structural and functional knowledge available for the central proteolytic fragment of the cascade, C3b. Using parallel in silico screening methodologies we identified 45 small molecules which putatively bind C3b near ligand-guided functional hot-spots. Surface plasmon resonance experiments resulted in the validation of seven dose-dependent C3b-binding compounds. Competition-based biochemical assays demonstrated the ability of several C3b-binding compounds to interfere with binding of the original C3b ligand which guided their discovery. In vitro assays of complement function identified a single complement inhibitory compound, termed cmp-5, and mechanistic studies of the cmp-5 inhibitory mode revealed it acts at the level of C5 activation. This study has led to the identification of a promising new class of C3b-binding small molecule complement inhibitors, and to our knowledge, provides the first demonstration of cheminformatics-based complement-directed drug discovery. PMID:28298523

Rapid and simultaneous detection of ricin, staphylococcal enterotoxin B and saxitoxin by chemiluminescence-based microarray immunoassay.

PubMed

Szkola, A; Linares, E M; Worbs, S; Dorner, B G; Dietrich, R; Märtlbauer, E; Niessner, R; Seidel, M

2014-11-21

Simultaneous detection of small and large molecules on microarray immunoassays is a challenge that limits some applications in multiplex analysis. This is the case for biosecurity, where fast, cheap and reliable simultaneous detection of proteotoxins and small toxins is needed. Two highly relevant proteotoxins, ricin (60 kDa) and bacterial toxin staphylococcal enterotoxin B (SEB, 30 kDa) and the small phycotoxin saxitoxin (STX, 0.3 kDa) are potential biological warfare agents and require an analytical tool for simultaneous detection. Proteotoxins are successfully detected by sandwich immunoassays, whereas competitive immunoassays are more suitable for small toxins (<1 kDa). Based on this need, this work provides a novel and efficient solution based on anti-idiotypic antibodies for small molecules to combine both assay principles on one microarray. The biotoxin measurements are performed on a flow-through chemiluminescence microarray platform MCR3 in 18 minutes. The chemiluminescence signal was amplified by using a poly-horseradish peroxidase complex (polyHRP), resulting in low detection limits: 2.9 ± 3.1 μg L(-1) for ricin, 0.1 ± 0.1 μg L(-1) for SEB and 2.3 ± 1.7 μg L(-1) for STX. The developed multiplex system for the three biotoxins is completely novel, relevant in the context of biosecurity and establishes the basis for research on anti-idiotypic antibodies for microarray immunoassays.

Expression of checkpoint molecules on myeloid-derived suppressor cells.

PubMed

Ballbach, Marlene; Dannert, Angelika; Singh, Anurag; Siegmund, Darina M; Handgretinger, Rupert; Piali, Luca; Rieber, Nikolaus; Hartl, Dominik

2017-12-01

Myeloid-derived suppressor cells (MDSCs) are a heterogeneous cell population expanded in cancer, infection and autoimmunity capable of suppressing T-cell functions. Checkpoint inhibitors have emerged as a key therapeutic strategy in immune-oncology. While checkpoint molecules were initially associated with T cell functions, recent evidence suggests a broader expression and function in innate myeloid cells. Previous studies provided first evidence for a potential role for checkpoints on MDSCs, yet the human relevance remained poorly understood. Therefore, we investigated the expression and functional relevance of checkpoint molecules in human MDSC-T-cell interactions. Our studies demonstrate that programmed death-ligand 1 (PD-L1) is expressed on granulocytic MDSCs upon co-culture with T cells. Transwell experiments showed that cell-to-cell contact was required for MDSC-T-cell interactions and antibody blocking studies showed that targeting PD-L1 partially impaired MDSC-mediated T-cell suppression. Collectively, these studies suggest a role for PD-L1 in human MDSC function and thereby expand the functionality of this checkpoint beyond T cells, which could pave the way for further understanding and therapeutic targeting of PD-1/PD-L1 in innate immune-mediated diseases. Copyright © 2017 European Federation of Immunological Societies. Published by Elsevier B.V. All rights reserved.

Emergence of complex chemistry on an organic monolayer.

PubMed

Prins, Leonard J

2015-07-21

In many origin-of-life scenarios, inorganic materials, such as FeS or mineral clays, play an important role owing to their ability to concentrate and select small organic molecules on their surface and facilitate their chemical transformations into new molecules. However, considering that life is made up of organic matter, at a certain stage during the evolution the role of the inorganic material must have been taken over by organic molecules. How this exactly happened is unclear, and, indeed, a big gap separates the rudimentary level of organization involving inorganic materials and the complex organization of cells, which are the building blocks of life. Over the past years, we have extensively studied the interaction of small molecules with monolayer-protected gold nanoparticles (Au NPs) for the purpose of developing innovative sensing and catalytic systems. During the course of these studies, we realized that the functional role of this system is very similar to that typically attributed to inorganic surfaces in the early stages of life, with the important being difference that the functional properties (molecular recognition, catalysis, signaling, adaptation) originate entirely from the organic monolayer rather than the inorganic support. This led us to the proposition that this system may serve as a model that illustrates how the important role of inorganic surfaces in dictating chemical processes in the early stages of life may have been taken over by organic matter. Here, we reframe our previously obtained results in the context of the origin-of-life question. The following functional roles of Au NPs will be discussed: the ability to concentrate small molecules and create different local populations, the ability to catalyze the chemical transformation of bound molecules, and, finally, the ability to install rudimentary signaling pathways and display primitive adaptive behavior. In particular, we will show that many of the functional properties of the system originate from two features: the presence of metal ions that are complexed in the organic monolayer and the multivalent nature of the system. Complexed metal ions play an important role in determining the affinity and selectivity of the interaction with small molecules, but serve also as regulatory elements for determining how many molecules are bound simultaneously. Importantly, neighboring metal ion complexes also create catalytic pockets in which two metal ions cooperatively catalyze the cleavage of an RNA-model compound. The multivalent nature of the system permits multiple noncovalent interactions with small molecules that enhances the affinity, but is also at the basis of simple signal transduction pathways and adaptive behavior.

Ion-momentum imaging of dissociative attachment of electrons to molecules

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

Slaughter, D. S.; Belkacem, A.; McCurdy, C. W.

Here, we present an overview of experiments and theory relevant to dissociative electron attachment studied by momentum imaging. We describe several key examples of characteristic transient anion dynamics in the form of small polyatomic electron-molecule systems. In each of these examples the so-called axial recoil approximation is found to break down due to correlation of the electronic and nuclear degrees of freedom of the transient anion. Guided by anion fragment momentum measurements and predictions of the electron scattering attachment probability in the molecular frame, we demonstrate that accurate predictions of the dissociation dynamics can be achieved without a detailed investigationmore » of the surface topology of the relevant electronic states or the fragment trajectories on those surfaces.« less

Ion-momentum imaging of dissociative attachment of electrons to molecules

DOE PAGES

Slaughter, D. S.; Belkacem, A.; McCurdy, C. W.; ...

2016-10-24

Here, we present an overview of experiments and theory relevant to dissociative electron attachment studied by momentum imaging. We describe several key examples of characteristic transient anion dynamics in the form of small polyatomic electron-molecule systems. In each of these examples the so-called axial recoil approximation is found to break down due to correlation of the electronic and nuclear degrees of freedom of the transient anion. Guided by anion fragment momentum measurements and predictions of the electron scattering attachment probability in the molecular frame, we demonstrate that accurate predictions of the dissociation dynamics can be achieved without a detailed investigationmore » of the surface topology of the relevant electronic states or the fragment trajectories on those surfaces.« less

Shift in the equilibrium between on and off states of the allosteric switch in Ras-GppNHp affected by small molecules and bulk solvent composition.

PubMed

Holzapfel, Genevieve; Buhrman, Greg; Mattos, Carla

2012-08-07

Ras GTPase cycles between its active GTP-bound form promoted by GEFs and its inactive GDP-bound form promoted by GAPs to affect the control of various cellular functions. It is becoming increasingly apparent that subtle regulation of the GTP-bound active state may occur through promotion of substates mediated by an allosteric switch mechanism that induces a disorder to order transition in switch II upon ligand binding at an allosteric site. We show with high-resolution structures that calcium acetate and either dithioerythritol (DTE) or dithiothreitol (DTT) soaked into H-Ras-GppNHp crystals in the presence of a moderate amount of poly(ethylene glycol) (PEG) can selectively shift the equilibrium to the "on" state, where the active site appears to be poised for catalysis (calcium acetate), or to what we call the "ordered off" state, which is associated with an anticatalytic conformation (DTE or DTT). We also show that the equilibrium is reversible in our crystals and dependent on the nature of the small molecule present. Calcium acetate binding in the allosteric site stabilizes the conformation observed in the H-Ras-GppNHp/NOR1A complex, and PEG, DTE, and DTT stabilize the anticatalytic conformation observed in the complex between the Ras homologue Ran and Importin-β. The small molecules are therefore selecting biologically relevant conformations in the crystal that are sampled by the disordered switch II in the uncomplexed GTP-bound form of H-Ras. In the presence of a large amount of PEG, the ordered off conformation predominates, whereas in solution, in the absence of PEG, switch regions appear to remain disordered in what we call the off state, unable to bind DTE.

Shift in the Equilibrium between On and Off States of the Allosteric Switch in Ras-GppNHp Affected by Small Molecules and Bulk Solvent Composition

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

Holzapfel, Genevieve; Buhrman, Greg; Mattos, Carla

2012-08-31

Ras GTPase cycles between its active GTP-bound form promoted by GEFs and its inactive GDP-bound form promoted by GAPs to affect the control of various cellular functions. It is becoming increasingly apparent that subtle regulation of the GTP-bound active state may occur through promotion of substates mediated by an allosteric switch mechanism that induces a disorder to order transition in switch II upon ligand binding at an allosteric site. We show with high-resolution structures that calcium acetate and either dithioerythritol (DTE) or dithiothreitol (DTT) soaked into H-Ras-GppNHp crystals in the presence of a moderate amount of poly(ethylene glycol) (PEG) canmore » selectively shift the equilibrium to the 'on' state, where the active site appears to be poised for catalysis (calcium acetate), or to what we call the 'ordered off' state, which is associated with an anticatalytic conformation (DTE or DTT). We also show that the equilibrium is reversible in our crystals and dependent on the nature of the small molecule present. Calcium acetate binding in the allosteric site stabilizes the conformation observed in the H-Ras-GppNHp/NOR1A complex, and PEG, DTE, and DTT stabilize the anticatalytic conformation observed in the complex between the Ras homologue Ran and Importin-{beta}. The small molecules are therefore selecting biologically relevant conformations in the crystal that are sampled by the disordered switch II in the uncomplexed GTP-bound form of H-Ras. In the presence of a large amount of PEG, the ordered off conformation predominates, whereas in solution, in the absence of PEG, switch regions appear to remain disordered in what we call the off state, unable to bind DTE.« less

Test measurement on ion-molecule reactions in a ringelectrode ion trap

NASA Astrophysics Data System (ADS)

Savic, I.; Lukic, S. R.; Guth, I.; Gerlich, D.

2006-05-01

Very recently a new experimental setup has been developed allowing studies of astrophysically relevant collisions between neutral atoms and small pure carbon molecules from one side and ions from the other side and first results are obtained (Savić et al., 2005). The ions are stored in a radio- frequency (rf) ring-electrode trap and during reaction time exposed to the effusive carbon beam. In this paper, one of the final tests of the experimental setup is presented.

Small-Molecule Hormones: Molecular Mechanisms of Action

PubMed Central

Budzińska, Monika

2013-01-01

Small-molecule hormones play crucial roles in the development and in the maintenance of an adult mammalian organism. On the molecular level, they regulate a plethora of biological pathways. Part of their actions depends on their transcription-regulating properties, exerted by highly specific nuclear receptors which are hormone-dependent transcription factors. Nuclear hormone receptors interact with coactivators, corepressors, basal transcription factors, and other transcription factors in order to modulate the activity of target genes in a manner that is dependent on tissue, age and developmental and pathophysiological states. The biological effect of this mechanism becomes apparent not earlier than 30–60 minutes after hormonal stimulus. In addition, small-molecule hormones modify the function of the cell by a number of nongenomic mechanisms, involving interaction with proteins localized in the plasma membrane, in the cytoplasm, as well as with proteins localized in other cellular membranes and in nonnuclear cellular compartments. The identity of such proteins is still under investigation; however, it seems that extranuclear fractions of nuclear hormone receptors commonly serve this function. A direct interaction of small-molecule hormones with membrane phospholipids and with mRNA is also postulated. In these mechanisms, the reaction to hormonal stimulus appears within seconds or minutes. PMID:23533406

N-terminal aliphatic residues dictate the structure, stability, assembly, and small molecule binding of the coiled-coil region of cartilage oligomeric matrix protein.

PubMed

Gunasekar, Susheel K; Asnani, Mukta; Limbad, Chandani; Haghpanah, Jennifer S; Hom, Wendy; Barra, Hanna; Nanda, Soumya; Lu, Min; Montclare, Jin Kim

2009-09-15

The coiled-coil domain of cartilage oligomeric matrix protein (COMPcc) assembles into a homopentamer that naturally recognizes the small molecule 1,25-dihydroxyvitamin D(3) (vit D). To identify the residues critical for the structure, stability, oligomerization, and binding to vit D as well as two other small molecules, all-trans-retinol (ATR) and curcumin (CCM), here we perform an alanine scanning mutagenesis study. Ten residues lining the hydrophobic pocket of COMPcc were mutated into alanine; of the mutated residues, the N-terminal aliphatic residues L37, L44, V47, and L51 are responsible for maintaining the structure and function. Furthermore, two polar residues, T40 and Q54, within the N-terminal region when converted into alanine improve the alpha-helical structure, stability, and self-assembly behavior. Helical stability, oligomerization, and binding appear to be linked in a manner in which mutations that abolish helical structure and assembly bind poorly to vit D, ATR, and CCM. These results provide not only insight into COMPcc and its functional role but also useful guidelines for the design of stable, pentameric coiled-coils capable of selectively storing and delivering various small molecules.

LigandRNA: computational predictor of RNA–ligand interactions

PubMed Central

Philips, Anna; Milanowska, Kaja; Łach, Grzegorz; Bujnicki, Janusz M.

2013-01-01

RNA molecules have recently become attractive as potential drug targets due to the increased awareness of their importance in key biological processes. The increase of the number of experimentally determined RNA 3D structures enabled structure-based searches for small molecules that can specifically bind to defined sites in RNA molecules, thereby blocking or otherwise modulating their function. However, as of yet, computational methods for structure-based docking of small molecule ligands to RNA molecules are not as well established as analogous methods for protein-ligand docking. This motivated us to create LigandRNA, a scoring function for the prediction of RNA–small molecule interactions. Our method employs a grid-based algorithm and a knowledge-based potential derived from ligand-binding sites in the experimentally solved RNA–ligand complexes. As an input, LigandRNA takes an RNA receptor file and a file with ligand poses. As an output, it returns a ranking of the poses according to their score. The predictive power of LigandRNA favorably compares to five other publicly available methods. We found that the combination of LigandRNA and Dock6 into a “meta-predictor” leads to further improvement in the identification of near-native ligand poses. The LigandRNA program is available free of charge as a web server at http://ligandrna.genesilico.pl. PMID:24145824

Application of chemical biology in target identification and drug discovery.

PubMed

Zhu, Yue; Xiao, Ting; Lei, Saifei; Zhou, Fulai; Wang, Ming-Wei

2015-09-01

Drug discovery and development is vital to the well-being of mankind and sustainability of the pharmaceutical industry. Using chemical biology approaches to discover drug leads has become a widely accepted path partially because of the completion of the Human Genome Project. Chemical biology mainly solves biological problems through searching previously unknown targets for pharmacologically active small molecules or finding ligands for well-defined drug targets. It is a powerful tool to study how these small molecules interact with their respective targets, as well as their roles in signal transduction, molecular recognition and cell functions. There have been an increasing number of new therapeutic targets being identified and subsequently validated as a result of advances in functional genomics, which in turn led to the discovery of numerous active small molecules via a variety of high-throughput screening initiatives. In this review, we highlight some applications of chemical biology in the context of drug discovery.

Mining for Micropeptides.

PubMed

Makarewich, Catherine A; Olson, Eric N

2017-09-01

Advances in computational biology and large-scale transcriptome analyses have revealed that a much larger portion of the genome is transcribed than was previously recognized, resulting in the production of a diverse population of RNA molecules with both protein-coding and noncoding potential. Emerging evidence indicates that several RNA molecules have been mis-annotated as noncoding and in fact harbor short open reading frames (sORFs) that encode functional peptides and that have evaded detection until now due to their small size. sORF-encoded peptides (SEPs), or micropeptides, have been shown to have important roles in fundamental biological processes and in the maintenance of cellular homeostasis. These small proteins can act independently, for example as ligands or signaling molecules, or they can exert their biological functions by engaging with and modulating larger regulatory proteins. Given their small size, micropeptides may be uniquely suited to fine-tune complex biological systems. Copyright © 2017 Elsevier Ltd. All rights reserved.

Charge transfer through amino groups-small molecules interface improving the performance of electroluminescent devices

NASA Astrophysics Data System (ADS)

Havare, Ali Kemal; Can, Mustafa; Tozlu, Cem; Kus, Mahmut; Okur, Salih; Demic, Şerafettin; Demirak, Kadir; Kurt, Mustafa; Icli, Sıddık

2016-05-01

A carboxylic group functioned charge transporting was synthesized and self-assembled on an indium tin oxide (ITO) anode. A typical electroluminescent device [modified ITO/TPD (50 nm)/Alq3 (60 nm)/LiF (2 nm)/(120 nm)] was fabricated to investigate the effect of the amino groups-small molecules interface on the characteristics of the device. The increase in the surface work function of ITO is expected to facilitate the hole injection from the ITO anode to the Hole Transport Layer (HTL) in electroluminescence. The modified electroluminescent device could endure a higher current and showed a much higher luminance than the nonmodified one. For the produced electroluminescent devices, the I-V characteristics, optical characterization and quantum yields were performed. The external quantum efficiency of the modified electroluminescent device is improved as the result of the presence of the amino groups-small molecules interface.

A size selective porous silicon grating-coupled Bloch surface and sub-surface wave biosensor.

PubMed

Rodriguez, Gilberto A; Ryckman, Judson D; Jiao, Yang; Weiss, Sharon M

2014-03-15

A porous silicon (PSi) grating-coupled Bloch surface and sub-surface wave (BSW/BSSW) biosensor is demonstrated to size selectively detect the presence of both large and small molecules. The BSW is used to sense large immobilized analytes at the surface of the structure while the BSSW that is confined inside but near the top of the structure is used to sensitively detect small molecules. Functionality of the BSW and BSSW modes is theoretically described by dispersion relations, field confinements, and simulated refractive index shifts within the structure. The theoretical results are experimentally verified by detecting two different small chemical molecules and one large 40 base DNA oligonucleotide. The PSi-BSW/BSSW structure is benchmarked against current porous silicon technology and is shown to have a 6-fold higher sensitivity in detecting large molecules and a 33% improvement in detecting small molecules. This is the first report of a grating-coupled BSW biosensor and the first report of a BSSW propagating mode. © 2013 Published by Elsevier B.V.

Design of small-molecule epigenetic modulators

PubMed Central

Pachaiyappan, Boobalan

2013-01-01

The field of epigenetics has expanded rapidly to reveal multiple new targets for drug discovery. The functional elements of the epigenomic machinery can be catagorized as writers, erasers and readers, and together these elements control cellular gene expression and homeostasis. It is increasingly clear that aberrations in the epigenome can underly a variety of diseases, and thus discovery of small molecules that modulate the epigenome in a specific manner is a viable approach to the discovery of new therapeutic agents. In this Digest, the components of epigenetic control of gene expression will be briefly summarized, and efforts to identify small molecules that modulate epigenetic processes will be described. PMID:24300735

Induction and reversal of myotonic dystrophy type 1 pre-mRNA splicing defects by small molecules.

PubMed

Childs-Disney, Jessica L; Stepniak-Konieczna, Ewa; Tran, Tuan; Yildirim, Ilyas; Park, HaJeung; Chen, Catherine Z; Hoskins, Jason; Southall, Noel; Marugan, Juan J; Patnaik, Samarjit; Zheng, Wei; Austin, Chris P; Schatz, George C; Sobczak, Krzysztof; Thornton, Charles A; Disney, Matthew D

2013-01-01

The ability to control pre-mRNA splicing with small molecules could facilitate the development of therapeutics or cell-based circuits that control gene function. Myotonic dystrophy type 1 is caused by the dysregulation of alternative pre-mRNA splicing due to sequestration of muscleblind-like 1 protein (MBNL1) by expanded, non-coding r(CUG) repeats (r(CUG)(exp)). Here we report two small molecules that induce or ameliorate alternative splicing dysregulation. A thiophene-containing small molecule (1) inhibits the interaction of MBNL1 with its natural pre-mRNA substrates. Compound (2), a substituted naphthyridine, binds r(CUG)(exp) and displaces MBNL1. Structural models show that 1 binds MBNL1 in the Zn-finger domain and that 2 interacts with UU loops in r(CUG)(exp). This study provides a structural framework for small molecules that target MBNL1 by mimicking r(CUG)(exp) and shows that targeting MBNL1 causes dysregulation of alternative splicing, suggesting that MBNL1 is thus not a suitable therapeutic target for the treatment of myotonic dystrophy type 1.

Induction and Reversal of Myotonic Dystrophy Type 1 Pre-mRNA Splicing Defects by Small Molecules

PubMed Central

Childs-Disney, Jessica L.; Stepniak-Konieczna, Ewa; Tran, Tuan; Yildirim, Ilyas; Park, HaJeung; Chen, Catherine Z.; Hoskins, Jason; Southall, Noel; Marugan, Juan J.; Patnaik, Samarjit; Zheng, Wei; Austin, Chris P.; Schatz, George C.; Sobczak, Krzysztof; Thornton, Charles A.; Disney, Matthew D.

2013-01-01

The ability to control pre-mRNA splicing with small molecules could facilitate the development of therapeutics or cell-based circuits that control gene function. Myotonic dystrophy type 1 (DM1) is caused by the dysregulation of alternative pre-mRNA splicing due to sequestration of muscleblind-like 1 protein (MBNL1) by expanded, non-coding r(CUG) repeats (r(CUG)exp). Here we report two small molecules that induce or ameliorate alternative splicing dysregulation. The thiophene-containing small molecule (1) inhibits the interaction of MBNL1 with its natural pre-mRNA substrates. Compound (2), a substituted naphthyridine, binds r(CUG)exp and displaces MBNL1. Structural models show that 1 binds MBNL1 in the Zn-finger domain and that 2 interacts with UU loops in r(CUG)exp. This study provides a structural framework for small molecules that target MBNL1 by mimicking r(CUG)exp and shows that targeting MBNL1 causes dysregulation of alternative splicing, suggesting that MBNL1 is thus not a suitable therapeutic target for the treatment of DM1. PMID:23806903

Re-education begins at home: an overview of the discovery of in vivo-active small molecule modulators of endogenous stem cells.

PubMed

Um, JungIn; Lee, Ji-Hyung; Jung, Da-Woon; Williams, Darren R

2018-04-01

Degenerative diseases, such as Alzheimer's disease, heart disease and arthritis cause great suffering and are major socioeconomic burdens. An attractive treatment approach is stem cell transplantation to regenerate damaged or destroyed tissues. However, this can be problematic. For example, donor cells may not functionally integrate into the host tissue. An alternative methodology is to deliver bioactive agents, such as small molecules, directly into the diseased tissue to enhance the regenerative potential of endogenous stem cells. Areas covered: In this review, the authors discuss the necessity of developing these small molecules to treat degenerative diseases and survey progress in their application as therapeutics. They describe both the successes and caveats of developing small molecules that target endogenous stem cells to induce tissue regeneration. This article is based on literature searches which encompass databases for biomedical research and clinical trials. These small molecules are also categorized per their target disease and mechanism of action. Expert opinion: The development of small molecules targeting endogenous stem cells is a high-profile research area. Some compounds have made the successful transition to the clinic. Novel approaches, such as modulating the stem cell niche or targeted delivery to disease sites, should increase the likelihood of future successes in this field.

Small Molecule Supplements Improve Cultured Megakaryocyte Polyploidization by Modulating Multiple Cell Cycle Regulators.

PubMed

Zou, Xiaojing; Qu, Mingyi; Fang, Fang; Fan, Zeng; Chen, Lin; Yue, Wen; Xie, Xiaoyan; Pei, Xuetao

2017-01-01

Platelets (PLTs) are produced by megakaryocytes (MKs) that completed differentiation and endomitosis. Endomitosis is an important process in which the cell replicates its DNA without cytokinesis and develops highly polyploid MK. In this study, to gain a better PLTs production, four small molecules (Rho-Rock inhibitor (RRI), nicotinamide (NIC), Src inhibitor (SI), and Aurora B inhibitor (ABI)) and their combinations were surveyed as MK culture supplements for promoting polyploidization. Three leukemia cell lines as well as primary mononuclear cells were chosen in the function and mechanism studies of the small molecules. In an optimal culture method, cells were treated with different small molecules and their combinations. The impact of the small molecules on megakaryocytic surface marker expression, polyploidy, proliferation, and apoptosis was examined for the best MK polyploidization supplement. The elaborate analysis confirmed that the combination of SI and RRI together with our MK induction system might result in efficient ploidy promotion. Our experiments demonstrated that, besides direct downregulation on the expression of cytoskeleton protein actin, SI and RRI could significantly enhance the level of cyclins through the suppression of p53 and p21. The verified small molecule combination might be further used in the in vitro PLT manufacture and clinical applications.

Small Molecule Supplements Improve Cultured Megakaryocyte Polyploidization by Modulating Multiple Cell Cycle Regulators

PubMed Central

Fang, Fang; Chen, Lin; Yue, Wen

2017-01-01

Platelets (PLTs) are produced by megakaryocytes (MKs) that completed differentiation and endomitosis. Endomitosis is an important process in which the cell replicates its DNA without cytokinesis and develops highly polyploid MK. In this study, to gain a better PLTs production, four small molecules (Rho-Rock inhibitor (RRI), nicotinamide (NIC), Src inhibitor (SI), and Aurora B inhibitor (ABI)) and their combinations were surveyed as MK culture supplements for promoting polyploidization. Three leukemia cell lines as well as primary mononuclear cells were chosen in the function and mechanism studies of the small molecules. In an optimal culture method, cells were treated with different small molecules and their combinations. The impact of the small molecules on megakaryocytic surface marker expression, polyploidy, proliferation, and apoptosis was examined for the best MK polyploidization supplement. The elaborate analysis confirmed that the combination of SI and RRI together with our MK induction system might result in efficient ploidy promotion. Our experiments demonstrated that, besides direct downregulation on the expression of cytoskeleton protein actin, SI and RRI could significantly enhance the level of cyclins through the suppression of p53 and p21. The verified small molecule combination might be further used in the in vitro PLT manufacture and clinical applications. PMID:29201898

Theoretical Investigation of Single-Molecule Sensing Using Nanotube-Enhanced Circular Dichroism.

PubMed

Silva, Jaime; Milne, Bruce F; Nogueira, Fernando

2018-06-19

First-principles calculations have been used to investigate the potential use of circular dichroism (CD) spectroscopy in single-molecule sensing. Using a real-space implementation of time-dependent density functional theory (TDDFT), several systems involving single-walled carbon nanotubes (SWCNT) and small molecules have been studied to evaluate their CD response. Large induced CD (ICD) effects, differing for each test molecule, were observed in all SWCNT-molecule complexes. As the SWCNT used in this study shows no intrinsic CD response, the ICD spectra are the result of interaction with the small molecules. This finding is general and independent of the (a)chiral nature of the adsorbed molecule. Our results indicate that it is possible to design a system that uses SWCNT for detection of molecules using the change in CD spectrum of the system induced by adsorption of the molecule onto the SWCNT surface.

Probing the catalytic functions of Bub1 kinase using the small molecule inhibitors BAY-320 and BAY-524

PubMed Central

Baron, Anna P; von Schubert, Conrad; Cubizolles, Fabien; Siemeister, Gerhard; Hitchcock, Marion; Mengel, Anne; Schröder, Jens; Fernández-Montalván, Amaury; von Nussbaum, Franz; Mumberg, Dominik; Nigg, Erich A

2016-01-01

The kinase Bub1 functions in the spindle assembly checkpoint (SAC) and in chromosome congression, but the role of its catalytic activity remains controversial. Here, we use two novel Bub1 inhibitors, BAY-320 and BAY-524, to demonstrate potent Bub1 kinase inhibition both in vitro and in intact cells. Then, we compared the cellular phenotypes of Bub1 kinase inhibition in HeLa and RPE1 cells with those of protein depletion, indicative of catalytic or scaffolding functions, respectively. Bub1 inhibition affected chromosome association of Shugoshin and the chromosomal passenger complex (CPC), without abolishing global Aurora B function. Consequently, inhibition of Bub1 kinase impaired chromosome arm resolution but exerted only minor effects on mitotic progression or SAC function. Importantly, BAY-320 and BAY-524 treatment sensitized cells to low doses of Paclitaxel, impairing both chromosome segregation and cell proliferation. These findings are relevant to our understanding of Bub1 kinase function and the prospects of targeting Bub1 for therapeutic applications. DOI: http://dx.doi.org/10.7554/eLife.12187.001 PMID:26885717

microRNA function in left-right neuronal asymmetry: perspectives from C. elegans.

PubMed

Alqadah, Amel; Hsieh, Yi-Wen; Chuang, Chiou-Fen

2013-09-23

Left-right asymmetry in anatomical structures and functions of the nervous system is present throughout the animal kingdom. For example, language centers are localized in the left side of the human brain, while spatial recognition functions are found in the right hemisphere in the majority of the population. Disruption of asymmetry in the nervous system is correlated with neurological disorders. Although anatomical and functional asymmetries are observed in mammalian nervous systems, it has been a challenge to identify the molecular basis of these asymmetries. C. elegans has emerged as a prime model organism to investigate molecular asymmetries in the nervous system, as it has been shown to display functional asymmetries clearly correlated to asymmetric distribution and regulation of biologically relevant molecules. Small non-coding RNAs have been recently implicated in various aspects of neural development. Here, we review cases in which microRNAs are crucial for establishing left-right asymmetries in the C. elegans nervous system. These studies may provide insight into how molecular and functional asymmetries are established in the human brain.

Side-chain Engineering of Benzo[1,2-b:4,5-b’]dithiophene Core-structured Small Molecules for High-Performance Organic Solar Cells

PubMed Central

Yin, Xinxing; An, Qiaoshi; Yu, Jiangsheng; Guo, Fengning; Geng, Yongliang; Bian, Linyi; Xu, Zhongsheng; Zhou, Baojing; Xie, Linghai; Zhang, Fujun; Tang, Weihua

2016-01-01

Three novel small molecules have been developed by side-chain engineering on benzo[1,2-b:4,5-b’]dithiophene (BDT) core. The typical acceptor-donor-acceptor (A-D-A) structure is adopted with 4,8-functionalized BDT moieties as core, dioctylterthiophene as π bridge and 3-ethylrhodanine as electron-withdrawing end group. Side-chain engineering on BDT core exhibits small but measurable effect on the optoelectronic properties of small molecules. Theoretical simulation and X-ray diffraction study reveal the subtle tuning of interchain distance between conjugated backbones has large effect on the charge transport and thus the photovoltaic performance of these molecules. Bulk-heterojunction solar cells fabricated with a configuration of ITO/PEDOT:PSS/SM:PC71BM/PFN/Al exhibit a highest power conversion efficiency (PCE) of 6.99% after solvent vapor annealing. PMID:27140224

Side-chain Engineering of Benzo[1,2-b:4,5-b']dithiophene Core-structured Small Molecules for High-Performance Organic Solar Cells.

PubMed

Yin, Xinxing; An, Qiaoshi; Yu, Jiangsheng; Guo, Fengning; Geng, Yongliang; Bian, Linyi; Xu, Zhongsheng; Zhou, Baojing; Xie, Linghai; Zhang, Fujun; Tang, Weihua

2016-05-03

Three novel small molecules have been developed by side-chain engineering on benzo[1,2-b:4,5-b']dithiophene (BDT) core. The typical acceptor-donor-acceptor (A-D-A) structure is adopted with 4,8-functionalized BDT moieties as core, dioctylterthiophene as π bridge and 3-ethylrhodanine as electron-withdrawing end group. Side-chain engineering on BDT core exhibits small but measurable effect on the optoelectronic properties of small molecules. Theoretical simulation and X-ray diffraction study reveal the subtle tuning of interchain distance between conjugated backbones has large effect on the charge transport and thus the photovoltaic performance of these molecules. Bulk-heterojunction solar cells fabricated with a configuration of ITO/PEDOT:PSS/SM:PC71BM/PFN/Al exhibit a highest power conversion efficiency (PCE) of 6.99% after solvent vapor annealing.

Defining scaffold geometries for interacting with proteins: geometrical classification of secondary structure linking regions.

PubMed

Tran, Tran T; Kulis, Christina; Long, Steven M; Bryant, Darryn; Adams, Peter; Smythe, Mark L

2010-11-01

Medicinal chemists synthesize arrays of molecules by attaching functional groups to scaffolds. There is evidence suggesting that some scaffolds yield biologically active molecules more than others, these are termed privileged substructures. One role of the scaffold is to present its side-chains for molecular recognition, and biologically relevant scaffolds may present side-chains in biologically relevant geometries or shapes. Since drug discovery is primarily focused on the discovery of compounds that bind to proteinaceous targets, we have been deciphering the scaffold shapes that are used for binding proteins as they reflect biologically relevant shapes. To decipher the scaffold architecture that is important for binding protein surfaces, we have analyzed the scaffold architecture of protein loops, which are defined in this context as continuous four residue segments of a protein chain that are not part of an α-helix or β-strand secondary structure. Loops are an important molecular recognition motif of proteins. We have found that 39 clusters reflect the scaffold architecture of 89% of the 23,331 loops in the dataset, with average intra-cluster and inter-cluster RMSD of 0.47 and 1.91, respectively. These protein loop scaffolds all have distinct shapes. We have used these 39 clusters that reflect the scaffold architecture of protein loops as biological descriptors. This involved generation of a small dataset of scaffold-based peptidomimetics. We found that peptidomimetic scaffolds with reported biological activities matched loop scaffold geometries and those peptidomimetic scaffolds with no reported biologically activities did not. This preliminary evidence suggests that organic scaffolds with tight matches to the preferred loop scaffolds of proteins, implies the likelihood of the scaffold to be biologically relevant.

Defining scaffold geometries for interacting with proteins: geometrical classification of secondary structure linking regions

NASA Astrophysics Data System (ADS)

Tran, Tran T.; Kulis, Christina; Long, Steven M.; Bryant, Darryn; Adams, Peter; Smythe, Mark L.

2010-11-01

Medicinal chemists synthesize arrays of molecules by attaching functional groups to scaffolds. There is evidence suggesting that some scaffolds yield biologically active molecules more than others, these are termed privileged substructures. One role of the scaffold is to present its side-chains for molecular recognition, and biologically relevant scaffolds may present side-chains in biologically relevant geometries or shapes. Since drug discovery is primarily focused on the discovery of compounds that bind to proteinaceous targets, we have been deciphering the scaffold shapes that are used for binding proteins as they reflect biologically relevant shapes. To decipher the scaffold architecture that is important for binding protein surfaces, we have analyzed the scaffold architecture of protein loops, which are defined in this context as continuous four residue segments of a protein chain that are not part of an α-helix or β-strand secondary structure. Loops are an important molecular recognition motif of proteins. We have found that 39 clusters reflect the scaffold architecture of 89% of the 23,331 loops in the dataset, with average intra-cluster and inter-cluster RMSD of 0.47 and 1.91, respectively. These protein loop scaffolds all have distinct shapes. We have used these 39 clusters that reflect the scaffold architecture of protein loops as biological descriptors. This involved generation of a small dataset of scaffold-based peptidomimetics. We found that peptidomimetic scaffolds with reported biological activities matched loop scaffold geometries and those peptidomimetic scaffolds with no reported biologically activities did not. This preliminary evidence suggests that organic scaffolds with tight matches to the preferred loop scaffolds of proteins, implies the likelihood of the scaffold to be biologically relevant.

Solving the Schrödinger equation of molecules by relaxing the antisymmetry rule: Inter-exchange theory.

PubMed

Nakatsuji, Hiroshi; Nakashima, Hiroyuki

2015-05-21

The Schrödinger equation (SE) and the antisymmetry principle constitute the governing principle of chemistry. A general method of solving the SE was presented before as the free complement (FC) theory, which gave highly accurate solutions for small atoms and molecules. We assume here to use the FC theory starting from the local valence bond wave function. When this theory is applied to larger molecules, antisymmetrizations of electronic wave functions become time-consuming and therefore, an additional breakthrough is necessary concerning the antisymmetry principle. Usually, in molecular calculations, we first construct the wave function to satisfy the antisymmetry rule, "electronic wave functions must be prescribed to be antisymmetric for all exchanges of electrons, otherwise bosonic interference may disturb the basis of the science." Starting from determinantal wave functions is typical. Here, we give an antisymmetrization theory, called inter-exchange (iExg) theory, by dividing molecular antisymmetrizations to those within atoms and between atoms. For the electrons belonging to distant atoms in a molecule, only partial antisymmetrizations or even no antisymmetrizations are necessary, depending on the distance between the atoms. So, the above antisymmetry rule is not necessarily followed strictly to get the results of a desired accuracy. For this and other reasons, the necessary parts of the antisymmetrization operations become very small as molecules become larger, leading finally to the operation counts of lower orders of N, the number of electrons. This theory creates a natural antisymmetrization method that is useful for large molecules.

Fission yeast tropomyosin specifies directed transport of myosin-V along actin cables

PubMed Central

Clayton, Joseph E.; Pollard, Luther W.; Sckolnick, Maria; Bookwalter, Carol S.; Hodges, Alex R.; Trybus, Kathleen M.; Lord, Matthew

2014-01-01

A hallmark of class-V myosins is their processivity—the ability to take multiple steps along actin filaments without dissociating. Our previous work suggested, however, that the fission yeast myosin-V (Myo52p) is a nonprocessive motor whose activity is enhanced by tropomyosin (Cdc8p). Here we investigate the molecular mechanism and physiological relevance of tropomyosin-mediated regulation of Myo52p transport, using a combination of in vitro and in vivo approaches. Single molecules of Myo52p, visualized by total internal reflection fluorescence microscopy, moved processively only when Cdc8p was present on actin filaments. Small ensembles of Myo52p bound to a quantum dot, mimicking the number of motors bound to physiological cargo, also required Cdc8p for continuous motion. Although a truncated form of Myo52p that lacked a cargo-binding domain failed to support function in vivo, it still underwent actin-dependent movement to polarized growth sites. This result suggests that truncated Myo52p lacking cargo, or single molecules of wild-type Myo52p with small cargoes, can undergo processive movement along actin-Cdc8p cables in vivo. Our findings outline a mechanism by which tropomyosin facilitates sorting of transport to specific actin tracks within the cell by switching on myosin processivity. PMID:24196839

Open Innovation Drug Discovery (OIDD): a potential path to novel therapeutic chemical space.

PubMed

Alvim-Gaston, Maria; Grese, Timothy; Mahoui, Abdelaziz; Palkowitz, Alan D; Pineiro-Nunez, Marta; Watson, Ian

2014-01-01

The continued development of computational and synthetic methods has enabled the enumeration or preparation of a nearly endless universe of chemical structures. Nevertheless, the ability of this chemical universe to deliver small molecules that can both modulate biological targets and have drug-like physicochemical properties continues to be a topic of interest to the pharmaceutical industry and academic researchers alike. The chemical space described by public, commercial, in-house and virtual compound collections has been interrogated by multiple approaches including biochemical, cellular and virtual screening, diversity analysis, and in-silico profiling. However, current drugs and known chemical probes derived from these efforts are contained within a remarkably small volume of the predicted chemical space. Access to more diverse classes of chemical scaffolds that maintain the properties relevant for drug discovery is certainly needed to meet the increasing demands for pharmaceutical innovation. The Lilly Open Innovation Drug Discovery platform (OIDD) was designed to tackle barriers to innovation through the identification of novel molecules active in relevant disease biology models. In this article we will discuss several computational approaches towards describing novel, biologically active, drug-like chemical space and illustrate how the OIDD program may facilitate access to previously untapped molecules that may aid in the search for innovative pharmaceuticals.

The interdependence between screening methods and screening libraries.

PubMed

Shelat, Anang A; Guy, R Kiplin

2007-06-01

The most common methods for discovery of chemical compounds capable of manipulating biological function involves some form of screening. The success of such screens is highly dependent on the chemical materials - commonly referred to as libraries - that are assayed. Classic methods for the design of screening libraries have depended on knowledge of target structure and relevant pharmacophores for target focus, and on simple count-based measures to assess other properties. The recent proliferation of two novel screening paradigms, structure-based screening and high-content screening, prompts a profound rethink about the ideal composition of small-molecule screening libraries. We suggest that currently utilized libraries are not optimal for addressing new targets by high-throughput screening, or complex phenotypes by high-content screening.

Molecular dynamics simulations of trihalomethanes removal from water using boron nitride nanosheets.

PubMed

Azamat, Jafar; Khataee, Alireza; Joo, Sang Woo

2016-04-01

Molecular dynamics simulations were performed to investigate the separation of trihalomethanes (THMs) from water using boron nitride nanosheets (BNNSs). The studied systems included THM molecules and a functionalized BNNS membrane immersed in an aqueous solution. An external pressure was applied to the z axis of the systems. Two functionalized BNNSs with large fluorinated-hydrogenated pore (F-H-pores) and small hydrogen-hydroxyl pore (H-OH-pores) were used. The pores of the BNNS membrane were obtained by passivating each nitrogen and boron atoms at the pore edges with fluorine and hydrogen atoms in the large pore or with hydroxyl and hydrogen atoms in the small pore. The results show that the BNNS with a small functionalized pore was impermeable to THM molecules, in contrast to the BNNS with a large functionalized pore. Using these membranes, water contaminants can be removed at lower cost.

The Endoplasmic Reticulum Membrane Is Permeable to Small Molecules

PubMed Central

Le Gall, Sylvie; Neuhof, Andrea; Rapoport, Tom

2004-01-01

The lumen of the endoplasmic reticulum (ER) differs from the cytosol in its content of ions and other small molecules, but it is unclear whether the ER membrane is as impermeable as other membranes in the cell. Here, we have tested the permeability of the ER membrane to small, nonphysiological molecules. We report that isolated ER vesicles allow different chemical modification reagents to pass from the outside into the lumen with little hindrance. In permeabilized cells, the ER membrane allows the passage of a small, charged modification reagent that is unable to cross the plasma membrane or the lysosomal and trans-Golgi membranes. A larger polar reagent of ∼5 kDa is unable to pass through the ER membrane. Permeation of the small molecules is passive because it occurs at low temperature in the absence of energy. These data indicate that the ER membrane is significantly more leaky than other cellular membranes, a property that may be required for protein folding and other functions of the ER. PMID:14617815

Computational multiscale modeling in protein--ligand docking.

PubMed

Taufer, Michela; Armen, Roger; Chen, Jianhan; Teller, Patricia; Brooks, Charles

2009-01-01

In biological systems, the binding of small molecule ligands to proteins is a crucial process for almost every aspect of biochemistry and molecular biology. Enzymes are proteins that function by catalyzing specific biochemical reactions that convert reactants into products. Complex organisms are typically composed of cells in which thousands of enzymes participate in complex and interconnected biochemical pathways. Some enzymes serve as sequential steps in specific pathways (such as energy metabolism), while others function to regulate entire pathways and cellular functions [1]. Small molecule ligands can be designed to bind to a specific enzyme and inhibit the biochemical reaction. Inhibiting the activity of key enzymes may result in the entire biochemical pathways being turned on or off [2], [3]. Many small molecule drugs marketed today function in this generic way as enzyme inhibitors. If research identifies a specific enzyme as being crucial to the progress of disease, then this enzyme may be targeted with an inhibitor, which may slow down or reverse the progress of disease. In this way, enzymes are targeted from specific pathogens (e.g., virus, bacteria, fungi) for infectious diseases [4], [5], and human enzymes are targeted for noninfectious diseases such as cardiovascular disease, cancer, diabetes, and neurodegenerative diseases [6].

The role of p53 in cancer drug resistance and targeted chemotherapy.

PubMed

Hientz, Karin; Mohr, André; Bhakta-Guha, Dipita; Efferth, Thomas

2017-01-31

Cancer has long been a grievous disease complicated by innumerable players aggravating its cure. Many clinical studies demonstrated the prognostic relevance of the tumor suppressor protein p53 for many human tumor types. Overexpression of mutated p53 with reduced or abolished function is often connected to resistance to standard medications, including cisplatin, alkylating agents (temozolomide), anthracyclines, (doxorubicin), antimetabolites (gemcitabine), antiestrogenes (tamoxifen) and EGFR-inhibitors (cetuximab). Such mutations in the TP53 gene are often accompanied by changes in the conformation of the p53 protein. Small molecules that restore the wild-type conformation of p53 and, consequently, rebuild its proper function have been identified. These promising agents include PRIMA-1, MIRA-1, and several derivatives of the thiosemicarbazone family. In addition to mutations in p53 itself, p53 activity may be also be impaired due to alterations in p53's regulating proteins such as MDM2. MDM2 functions as primary cellular p53 inhibitor and deregulation of the MDM2/p53-balance has serious consequences. MDM2 alterations often result in its overexpression and therefore promote inhibition of p53 activity. To deal with this problem, a judicious approach is to employ MDM2 inhibitors. Several promising MDM2 inhibitors have been described such as nutlins, benzodiazepinediones or spiro-oxindoles as well as novel compound classes such as xanthone derivatives and trisubstituted aminothiophenes. Furthermore, even naturally derived inhibitor compounds such as α-mangostin, gambogic acid and siladenoserinols have been discovered. In this review, we discuss in detail such small molecules that play a pertinent role in affecting the p53-MDM2 signaling axis and analyze their potential as cancer chemotherapeutics.

Glutamate system, amyloid β peptides and tau protein: functional interrelationships and relevance to Alzheimer disease pathology

PubMed Central

Revett, Timothy J.; Baker, Glen B.; Jhamandas, Jack; Kar, Satyabrata

2013-01-01

Alzheimer disease is the most prevalent form of dementia globally and is characterized premortem by a gradual memory loss and deterioration of higher cognitive functions and postmortem by neuritic plaques containing amyloid β peptide and neurofibrillary tangles containing phospho-tau protein. Glutamate is the most abundant neurotransmitter in the brain and is essential to memory formation through processes such as long-term potentiation and so might be pivotal to Alzheimer disease progression. This review discusses how the glutamatergic system is impaired in Alzheimer disease and how interactions of amyloid β and glutamate influence synaptic function, tau phosphorylation and neurodegeneration. Interestingly, glutamate not only influences amyloid β production, but also amyloid β can alter the levels of glutamate at the synapse, indicating that small changes in the concentrations of both molecules could influence Alzheimer disease progression. Finally, we describe how the glutamate receptor antagonist, memantine, has been used in the treatment of individuals with Alzheimer disease and discuss its effectiveness. PMID:22894822

Polyamines: naturally occurring small molecule modulators of electrostatic protein-protein interactions.

PubMed

Berwanger, Anja; Eyrisch, Susanne; Schuster, Inge; Helms, Volkhard; Bernhardt, Rita

2010-02-01

Modulations of protein-protein interactions are a key step in regulating protein function, especially in networks. Modulators of these interactions are supposed to be candidates for the development of novel drugs. Here, we describe the role of the small, polycationic and highly abundant natural polyamines that could efficiently bind to charged spots at protein interfaces as modulators of such protein-protein interactions. Using the mitochondrial cytochrome P45011A1 (CYP11A1) electron transfer system as a model, we have analyzed the capability of putrescine, spermidine, and spermine at physiologically relevant concentrations to affect the protein-protein interactions between adrenodoxin reductase (AdR), adrenodoxin (Adx), and CYP11A1. The actions of polyamines on the individual components, on their association/dissociation, on electron transfer, and on substrate conversion were examined. These studies revealed modulating effects of polyamines on distinct interactions and on the entire system in a complex way. Modulation via changed protein-protein interactions appeared plausible from docking experiments that suggested favourable high-affinity binding sites of polyamines (spermine>spermidine>putrescine) at the AdR-Adx interface. Our findings imply for the first time that small endogenous compounds are capable of interfering with distinct components of transient protein complexes and might control protein functions by modulating electrostatic protein-protein interactions.

Noncompetitive Homogeneous Detection of Small Molecules Using Synthetic Nanopeptamer-Based Luminescent Oxygen Channeling.

PubMed

Lassabe, Gabriel; Kramer, Karl; Hammock, Bruce D; González-Sapienza, Gualberto; González-Techera, Andrés

2018-05-15

Our group has previously developed immunoassays for noncompetitive detection of small molecules based on the use of phage borne anti-immunocomplex peptides. Recently, we substituted the phage particles by biotinylated synthetic anti-immunocomplex peptides complexed with streptavidin and named these constructs nanopeptamers. In this work, we report the results of combining AlphaLisa, a commercial luminescent oxygen channeling bead system, with nanopeptamers for the development of a noncompetitive homogeneous assay for the detection of small molecules. The signal generation of AlphaLisa assays relies on acceptor-donor bead proximity induced by the presence of the analyte (a macromolecule) simultaneously bound by antibodies immobilized on the surface of these beads. In the developed assay, termed as nanoAlphaLisa, bead proximity is sustained by the presence of a small model molecule (atrazine, MW = 215) using an antiatrazine antibody captured on the acceptor bead and an atrazine nanopeptamer on the donor bead. Atrazine is one of the most used pesticides worldwide, and its monitoring in water has relevant human health implications. NanoAlphaLisa allowed the homogeneous detection of atrazine down to 0.3 ng/mL in undiluted water samples in 1 h, which is 10-fold below the accepted limit in drinking water. NanoAlphaLisa has the intrinsic advantages for automation and high-throughput, simple, and fast homogeneous detection of target analytes that AlphaLisa assay provides.

Computational systems chemical biology.

PubMed

Oprea, Tudor I; May, Elebeoba E; Leitão, Andrei; Tropsha, Alexander

2011-01-01

There is a critical need for improving the level of chemistry awareness in systems biology. The data and information related to modulation of genes and proteins by small molecules continue to accumulate at the same time as simulation tools in systems biology and whole body physiologically based pharmacokinetics (PBPK) continue to evolve. We called this emerging area at the interface between chemical biology and systems biology systems chemical biology (SCB) (Nat Chem Biol 3: 447-450, 2007).The overarching goal of computational SCB is to develop tools for integrated chemical-biological data acquisition, filtering and processing, by taking into account relevant information related to interactions between proteins and small molecules, possible metabolic transformations of small molecules, as well as associated information related to genes, networks, small molecules, and, where applicable, mutants and variants of those proteins. There is yet an unmet need to develop an integrated in silico pharmacology/systems biology continuum that embeds drug-target-clinical outcome (DTCO) triplets, a capability that is vital to the future of chemical biology, pharmacology, and systems biology. Through the development of the SCB approach, scientists will be able to start addressing, in an integrated simulation environment, questions that make the best use of our ever-growing chemical and biological data repositories at the system-wide level. This chapter reviews some of the major research concepts and describes key components that constitute the emerging area of computational systems chemical biology.

Computational Systems Chemical Biology

PubMed Central

Oprea, Tudor I.; May, Elebeoba E.; Leitão, Andrei; Tropsha, Alexander

2013-01-01

There is a critical need for improving the level of chemistry awareness in systems biology. The data and information related to modulation of genes and proteins by small molecules continue to accumulate at the same time as simulation tools in systems biology and whole body physiologically-based pharmacokinetics (PBPK) continue to evolve. We called this emerging area at the interface between chemical biology and systems biology systems chemical biology, SCB (Oprea et al., 2007). The overarching goal of computational SCB is to develop tools for integrated chemical-biological data acquisition, filtering and processing, by taking into account relevant information related to interactions between proteins and small molecules, possible metabolic transformations of small molecules, as well as associated information related to genes, networks, small molecules and, where applicable, mutants and variants of those proteins. There is yet an unmet need to develop an integrated in silico pharmacology / systems biology continuum that embeds drug-target-clinical outcome (DTCO) triplets, a capability that is vital to the future of chemical biology, pharmacology and systems biology. Through the development of the SCB approach, scientists will be able to start addressing, in an integrated simulation environment, questions that make the best use of our ever-growing chemical and biological data repositories at the system-wide level. This chapter reviews some of the major research concepts and describes key components that constitute the emerging area of computational systems chemical biology. PMID:20838980

Quantifying small molecule phenotypic effects using mitochondrial morpho-functional fingerprinting and machine learning.

PubMed

Blanchet, Lionel; Smeitink, Jan A M; van Emst-de Vries, Sjenet E; Vogels, Caroline; Pellegrini, Mina; Jonckheere, An I; Rodenburg, Richard J T; Buydens, Lutgarde M C; Beyrath, Julien; Willems, Peter H G M; Koopman, Werner J H

2015-01-26

In primary fibroblasts from Leigh Syndrome (LS) patients, isolated mitochondrial complex I deficiency is associated with increased reactive oxygen species levels and mitochondrial morpho-functional changes. Empirical evidence suggests these aberrations constitute linked therapeutic targets for small chemical molecules. However, the latter generally induce multiple subtle effects, meaning that in vitro potency analysis or single-parameter high-throughput cell screening are of limited use to identify these molecules. We combine automated image quantification and artificial intelligence to discriminate between primary fibroblasts of a healthy individual and a LS patient based upon their mitochondrial morpho-functional phenotype. We then evaluate the effects of newly developed Trolox variants in LS patient cells. This revealed that Trolox ornithylamide hydrochloride best counterbalanced mitochondrial morpho-functional aberrations, effectively scavenged ROS and increased the maximal activity of mitochondrial complexes I, IV and citrate synthase. Our results suggest that Trolox-derived antioxidants are promising candidates in therapy development for human mitochondrial disorders.

Quantifying small molecule phenotypic effects using mitochondrial morpho-functional fingerprinting and machine learning

NASA Astrophysics Data System (ADS)

Blanchet, Lionel; Smeitink, Jan A. M.; van Emst-de Vries, Sjenet E.; Vogels, Caroline; Pellegrini, Mina; Jonckheere, An I.; Rodenburg, Richard J. T.; Buydens, Lutgarde M. C.; Beyrath, Julien; Willems, Peter H. G. M.; Koopman, Werner J. H.

2015-01-01

In primary fibroblasts from Leigh Syndrome (LS) patients, isolated mitochondrial complex I deficiency is associated with increased reactive oxygen species levels and mitochondrial morpho-functional changes. Empirical evidence suggests these aberrations constitute linked therapeutic targets for small chemical molecules. However, the latter generally induce multiple subtle effects, meaning that in vitro potency analysis or single-parameter high-throughput cell screening are of limited use to identify these molecules. We combine automated image quantification and artificial intelligence to discriminate between primary fibroblasts of a healthy individual and a LS patient based upon their mitochondrial morpho-functional phenotype. We then evaluate the effects of newly developed Trolox variants in LS patient cells. This revealed that Trolox ornithylamide hydrochloride best counterbalanced mitochondrial morpho-functional aberrations, effectively scavenged ROS and increased the maximal activity of mitochondrial complexes I, IV and citrate synthase. Our results suggest that Trolox-derived antioxidants are promising candidates in therapy development for human mitochondrial disorders.

Solid harmonic wavelet scattering for predictions of molecule properties

NASA Astrophysics Data System (ADS)

Eickenberg, Michael; Exarchakis, Georgios; Hirn, Matthew; Mallat, Stéphane; Thiry, Louis

2018-06-01

We present a machine learning algorithm for the prediction of molecule properties inspired by ideas from density functional theory (DFT). Using Gaussian-type orbital functions, we create surrogate electronic densities of the molecule from which we compute invariant "solid harmonic scattering coefficients" that account for different types of interactions at different scales. Multilinear regressions of various physical properties of molecules are computed from these invariant coefficients. Numerical experiments show that these regressions have near state-of-the-art performance, even with relatively few training examples. Predictions over small sets of scattering coefficients can reach a DFT precision while being interpretable.

Defining the RNA Internal Loops Preferred by Benzimidazole Derivatives via Two-Dimensional Combinatorial Screening and Computational Analysis

PubMed Central

Velagapudi, Sai Pradeep; Seedhouse, Steven J.; French, Jonathan

2011-01-01

RNA is an important therapeutic target, however, RNA targets are generally underexploited due to a lack of understanding of the small molecules that bind RNA and the RNA motifs that bind small molecules. Herein, we describe the identification of the RNA internal loops derived from a 4096-member 3×3 nucleotide loop library that are the most specific and highest affinity binders to a series of four designer, drug-like benzimidazoles. These studies establish a potentially general protocol to define the highest affinity and most specific RNA motif targets for heterocyclic small molecules. Such information could be used to target functionally important RNAs in genomic sequence. PMID:21604752

Approaches to Validate and Manipulate RNA Targets with Small Molecules in Cells.

PubMed

Childs-Disney, Jessica L; Disney, Matthew D

2016-01-01

RNA has become an increasingly important target for therapeutic interventions and for chemical probes that dissect and manipulate its cellular function. Emerging targets include human RNAs that have been shown to directly cause cancer, metabolic disorders, and genetic disease. In this review, we describe various routes to obtain bioactive compounds that target RNA, with a particular emphasis on the development of small molecules. We use these cases to describe approaches that are being developed for target validation, which include target-directed cleavage, classic pull-down experiments, and covalent cross-linking. Thus, tools are available to design small molecules to target RNA and to identify the cellular RNAs that are their targets.

Design of small molecule epigenetic modulators.

PubMed

Pachaiyappan, Boobalan; Woster, Patrick M

2014-01-01

The field of epigenetics has expanded rapidly to reveal multiple new targets for drug discovery. The functional elements of the epigenomic machinery can be categorized as writers, erasers and readers, and together these elements control cellular gene expression and homeostasis. It is increasingly clear that aberrations in the epigenome can underly a variety of diseases, and thus discovery of small molecules that modulate the epigenome in a specific manner is a viable approach to the discovery of new therapeutic agents. In this Digest, the components of epigenetic control of gene expression will be briefly summarized, and efforts to identify small molecules that modulate epigenetic processes will be described. Copyright © 2013 The Authors. Published by Elsevier Ltd.. All rights reserved.

Immunological detection of small organic molecules in the presence of perchlorates: relevance to the life marker chip and life detection on Mars.

PubMed

Rix, Catherine S; Sims, Mark R; Cullen, David C

2011-11-01

The proposed ExoMars mission, due to launch in 2018, aims to look for evidence of extant and extinct life in martian rocks and regolith. Previous attempts to detect organic molecules of biological or abiotic origin on Mars have been unsuccessful, which may be attributable to destruction of these molecules by perchlorate salts during pyrolysis sample extraction techniques. Organic molecules can also be extracted and measured with solvent-based systems. The ExoMars payload includes the Life Marker Chip (LMC) instrument, capable of detecting biomarker molecules of extant and extinct Earth-like life in liquid extracts of martian samples with an antibody microarray assay. The aim of the work reported here was to investigate whether the presence of perchlorate salts, at levels similar to those at the NASA Phoenix landing site, would compromise the LMC extraction and detection method. To test this, we implemented an LMC-representative sample extraction process with an LMC-representative antibody assay and used these to extract and analyze a model sample that consisted of a Mars analog sample matrix (JSC Mars-1) spiked with a representative organic molecular target (pyrene, an example of abiotic meteoritic infall targets) in the presence of perchlorate salts. We found no significant change in immunoassay function when using pyrene standards with added perchlorate salts. When model samples spiked with perchlorate salts were subjected to an LMC-representative liquid extraction, immunoassays functioned in a liquid extract and detected extracted pyrene. For the same model sample matrix without perchlorate salts, we observed anomalous assay signals that coincided with yellow coloration of the extracts. This unexpected observation is being studied further. This initial study indicates that the presence of perchlorate salts, at levels similar to those detected at the NASA Phoenix landing site, is unlikely to prevent the LMC from extracting and detecting organic molecules from martian samples.

Developmental origins of novel gut morphology in frogs

PubMed Central

Bloom, Stephanie; Ledon-Rettig, Cris; Infante, Carlos; Everly, Anne; Hanken, James; Nascone-Yoder, Nanette

2013-01-01

SUMMARY Phenotypic variation is a prerequisite for evolution by natural selection, yet the processes that give rise to the novel morphologies upon which selection acts are poorly understood. We employed a chemical genetic screen to identify developmental changes capable of generating ecologically relevant morphological variation as observed among extant species. Specifically, we assayed for exogenously applied small molecules capable of transforming the ancestral larval foregut of the herbivorous Xenopus laevis to resemble the derived larval foregut of the carnivorous Lepidobatrachus laevis. Appropriately, the small molecules that demonstrate this capacity modulate conserved morphogenetic pathways involved in gut development, including downregulation of retinoic acid (RA) signaling. Identical manipulation of RA signaling in a species that is more closely related to Lepidobatrachus, Ceratophrys cranwelli, yielded even more similar transformations, corroborating the relevance of RA signaling variation in interspecific morphological change. Finally, we were able to recover the ancestral gut phenotype in Lepidobatrachus by performing a reverse chemical manipulation to upregulate RA signaling, providing strong evidence that modifications to this specific pathway promoted the emergence of a lineage-specific phenotypic novelty. Interestingly, our screen also revealed pathways that have not yet been implicated in early gut morphogenesis, such as thyroid hormone signaling. In general, the chemical genetic screen may be a valuable tool for identifying developmental mechanisms that underlie ecologically and evolutionarily relevant phenotypic variation. PMID:23607305

Studies Relevent to Catalytic Activation Co & other small Molecules

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

Ford, Peter C

2005-02-22

Detailed annual and triannual reports describing the progress accomplished during the tenure of this grant were filed with the Program Manager for Catalysis at the Office of Basic Energy Sciences. To avoid unnecessary duplication, the present report will provide a brief overview of the research areas that were sponsored by this grant and list the resulting publications and theses based on this DOE supported research. The scientific personnel participating in (and trained by) this grant's research are also listed. Research carried out under this DOE grant was largely concerned with the mechanisms of the homogeneous catalytic and photocatalytic activation ofmore » small molecules such as carbon monoxide, dihydrogen and various hydrocarbons. Much of the more recent effort has focused on the dynamics and mechanisms of reactions relevant to substrate carbonylations by homogeneous organometallic catalysts. A wide range of modern investigative techniques were employed, including quantitative fast reaction methodologies such as time-resolved optical (TRO) and time-resolved infrared (TRIR) spectroscopy and stopped flow kinetics. Although somewhat diverse, this research falls within the scope of the long-term objective of applying quantitative techniques to elucidate the dynamics and understand the principles of mechanisms relevant to the selective and efficient catalytic conversions of fundamental feedstocks to higher value materials.« less

[Advances in the study of natural small molecular antibody].

PubMed

Zhu, Lei; Zhang, Da-peng

2012-10-01

Small molecule antibodies are naturally existed and well functioned but not structurally related to the conventional antibodies. They are only composed of heavy protein chains or light chains, much smaller than common antibody. The first small molecule antibody, called Nanobody was engineered from heavy-chain antibodies found in camelids. Cartilaginous fishes also have heavy-chain antibodies (IgNAR, "immunoglobulin new antigen receptor"), from which single-domain antibodies called Vnar fragments can be obtained. In addition, free light chain (FLC) antibodies in human bodies are being developed as therapeutic and diagnostic agents. Comparing to intact antibodies, common advantages of small molecule antibodies are with better solubility, tissue penetration, stability towards heat and enzymes, and comparatively low production costs. This article reviews the structural characteristics and mechanism of action of the Nanobody, IgNAR and FLC.

The tortuous road to an ideal CGRP function blocker for the treatment of migraine.

PubMed

Davis, Carl D; Xu, Cen

2008-01-01

The critical role of Calcitonin Gene-Related Peptide (CGRP) in migraine has been validated, with two small molecule CGRP antagonists BIBN4096BS and MK-0974 demonstrating efficacy in the reversal of acute migraine attack. Multiple approaches have been taken to find the ideal agent that most effectively inhibits CGRP's function. Here, we have summarized the progress made in recent years, including the identification and optimization of an orally bioavailable small molecule CGRP receptor antagonist. We also describe other interventions such as scavenging of CGRP itself. The advantages and disadvantages of these distinct approaches are discussed.

Small molecule-induced cellular fate reprogramming: promising road leading to Rome.

PubMed

Li, Xiang; Xu, Jun; Deng, Hongkui

2018-05-29

Cellular fate reprogramming holds great promise to generate functional cell types for replenishing new cells and restoring functional loss. Inspired by transcription factor-induced reprogramming, the field of cellular reprogramming has greatly advanced and developed into divergent streams of reprogramming approaches. Remarkably, increasing studies have shown the power and advantages of small molecule-based approaches for cellular fate reprogramming, which could overcome the limitations of conventional transgenic-based reprogramming. In this concise review, we discuss these findings and highlight the future potentiality with particular focus on this new trend of chemical reprogramming. Copyright © 2018 Elsevier Ltd. All rights reserved.

In Silico Identification and Experimental Validation of Novel Anti-Alzheimer's Multitargeted Ligands from a Marine Source Featuring a "2-Aminoimidazole plus Aromatic Group" Scaffold.

PubMed

Vitale, Rosa Maria; Rispoli, Vincenzo; Desiderio, Doriana; Sgammato, Roberta; Thellung, Stefano; Canale, Claudio; Vassalli, Massimo; Carbone, Marianna; Ciavatta, Maria Letizia; Mollo, Ernesto; Felicità, Vera; Arcone, Rosaria; Gavagnin Capoggiani, Margherita; Masullo, Mariorosario; Florio, Tullio; Amodeo, Pietro

2018-03-07

Multitargeting or polypharmacological approaches, looking for single chemical entities retaining the ability to bind two or more molecular targets, are a potentially powerful strategy to fight complex, multifactorial pathologies. Unfortunately, the search for multiligand agents is challenging because only a small subset of molecules contained in molecular databases are bioactive and even fewer are active on a preselected set of multiple targets. However, collections of natural compounds feature a significantly higher fraction of bioactive molecules than synthetic ones. In this view, we searched our library of 1175 natural compounds from marine sources for molecules including a 2-aminoimidazole+aromatic group motif, found in known compounds active on single relevant targets for Alzheimer's disease (AD). This identified two molecules, a pseudozoanthoxanthin (1) and a bromo-pyrrole alkaloid (2), which were predicted by a computational approach to possess interesting multitarget profiles on AD target proteins. Biochemical assays experimentally confirmed their biological activities. The two compounds inhibit acetylcholinesterase, butyrylcholinesterase, and β-secretase enzymes in high- to sub-micromolar range. They are also able to prevent and revert β-amyloid (Aβ) aggregation of both Aβ 1-40 and Aβ 1-42 peptides, with 1 being more active than 2. Preliminary in vivo studies suggest that compound 1 is able to restore cholinergic cortico-hippocampal functional connectivity.

Transformational Medical Technologies Initiative (TMTI)

DTIC Science & Technology

2007-01-01

Parker Development of monoclonal antibodies (Mab) protein based therapeutics against viral hemorrhagic fevers (VHF) CUBRC Prosetta, Inc Development of...2,028,389 $178,416 $186,000 $558,804 CUBRC Prosetta, Inc 2 (1 base, 1yr opt) $8,300,922 $4,017,013 $1,070,977 $3,212,932 Small Molecule/Drugs...Green II Genomic Identification USAMRIID - Hart Green Yellow CUBRC Prosetta, Inc Yellow Yellow Small Molecule/Drugs Functional Genetics, Inc Green

Natural small-molecule enhancers of autophagy induce autophagic cell death in apoptosis-defective cells.

PubMed

Law, Betty Yuen Kwan; Chan, Wai Kit; Xu, Su Wei; Wang, Jing Rong; Bai, Li Ping; Liu, Liang; Wong, Vincent Kam Wai

2014-07-01

Resistance of cancer cells to chemotherapy is a significant problem in oncology, and the development of sensitising agents or small-molecules with new mechanisms of action to kill these cells is needed. Autophagy is a cellular process responsible for the turnover of misfolded proteins or damaged organelles, and it also recycles nutrients to maintain energy levels for cell survival. In some apoptosis-resistant cancer cells, autophagy can also enhance the efficacy of anti-cancer drugs through autophagy-mediated mechanisms of cell death. Because the modulation of autophagic processes can be therapeutically useful to circumvent chemoresistance and enhance the effects of cancer treatment, the identification of novel autophagic enhancers for use in oncology is highly desirable. Many novel anti-cancer compounds have been isolated from natural products; therefore, we worked to discover natural, anti-cancer small-molecule enhancers of autophagy. Here, we have identified a group of natural alkaloid small-molecules that function as novel autophagic enhancers. These alkaloids, including liensinine, isoliensinine, dauricine and cepharanthine, stimulated AMPK-mTOR dependent induction of autophagy and autophagic cell death in a panel of apoptosis-resistant cells. Taken together, our work provides novel insights into the biological functions, mechanisms and potential therapeutic values of alkaloids for the induction of autophagy.

Small-molecule agonists for the thyrotropin receptor stimulate thyroid function in human thyrocytes and mice

PubMed Central

Neumann, Susanne; Huang, Wenwei; Titus, Steve; Krause, Gerd; Kleinau, Gunnar; Alberobello, Anna Teresa; Zheng, Wei; Southall, Noel T.; Inglese, James; Austin, Christopher P.; Celi, Francesco S.; Gavrilova, Oksana; Thomas, Craig J.; Raaka, Bruce M.; Gershengorn, Marvin C.

2009-01-01

Seven-transmembrane-spanning receptors (7TMRs) are prominent drug targets. However, small-molecule ligands for 7-transmembrane-spanning receptors for which the natural ligands are large, heterodimeric glycoprotein hormones, like thyroid-stimulating hormone (TSH; thyrotropin), have only recently been reported, and none are approved for human use. We have used quantitative high-throughput screening to identify a small-molecule TSH receptor (TSHR) agonist that was modified to produce a second agonist with increased potency. We show that these agonists are highly selective for human TSHR versus other glycoprotein hormone receptors and interact with the receptor's serpentine domain. A binding pocket within the transmembrane domain was defined by docking into a TSHR homology model and was supported by site-directed mutagenesis. In primary cultures of human thyrocytes, both TSH and the agonists increase mRNA levels for thyroglobulin, thyroperoxidase, sodium iodide symporter, and deiodinase type 2, and deiodinase type 2 enzyme activity. Moreover, oral administration of the agonist stimulated thyroid function in mice, resulting in increased serum thyroxine and thyroidal radioiodide uptake. Thus, we discovered a small molecule that activates human TSHR in vitro, is orally active in mice, and could be a lead for development of drugs to use in place of recombinant human TSH in patients with thyroid cancer. PMID:19592511

Chemical methodology as a source of small-molecule checkpoint inhibitors and heat shock protein 70 (Hsp70) modulators.

PubMed

Huryn, Donna M; Brodsky, Jeffrey L; Brummond, Kay M; Chambers, Peter G; Eyer, Benjamin; Ireland, Alex W; Kawasumi, Masaoki; Laporte, Matthew G; Lloyd, Kayla; Manteau, Baptiste; Nghiem, Paul; Quade, Bettina; Seguin, Sandlin P; Wipf, Peter

2011-04-26

Unique chemical methodology enables the synthesis of innovative and diverse scaffolds and chemotypes and allows access to previously unexplored "chemical space." Compound collections based on such new synthetic methods can provide small-molecule probes of proteins and/or pathways whose functions are not fully understood. We describe the identification, characterization, and evolution of two such probes. In one example, a pathway-based screen for DNA damage checkpoint inhibitors identified a compound, MARPIN (ATM and ATR pathway inhibitor) that sensitizes p53-deficient cells to DNA-damaging agents. Modification of the small molecule and generation of an immobilized probe were used to selectively bind putative protein target(s) responsible for the observed activity. The second example describes a focused library approach that relied on tandem multicomponent reaction methodologies to afford a series of modulators of the heat shock protein 70 (Hsp70) molecular chaperone. The synthesis of libraries based on the structure of MAL3-101 generated a collection of chemotypes, each modulating Hsp70 function, but exhibiting divergent pharmacological activities. For example, probes that compromise the replication of a disease-associated polyomavirus were identified. These projects highlight the importance of chemical methodology development as a source of small-molecule probes and as a drug discovery starting point.

Chemical methodology as a source of small-molecule checkpoint inhibitors and heat shock protein 70 (Hsp70) modulators

PubMed Central

Huryn, Donna M.; Brodsky, Jeffrey L.; Brummond, Kay M.; Chambers, Peter G.; Eyer, Benjamin; Ireland, Alex W.; Kawasumi, Masaoki; LaPorte, Matthew G.; Lloyd, Kayla; Manteau, Baptiste; Nghiem, Paul; Quade, Bettina; Seguin, Sandlin P.; Wipf, Peter

2011-01-01

Unique chemical methodology enables the synthesis of innovative and diverse scaffolds and chemotypes and allows access to previously unexplored “chemical space.” Compound collections based on such new synthetic methods can provide small-molecule probes of proteins and/or pathways whose functions are not fully understood. We describe the identification, characterization, and evolution of two such probes. In one example, a pathway-based screen for DNA damage checkpoint inhibitors identified a compound, MARPIN (ATM and ATR pathway inhibitor) that sensitizes p53-deficient cells to DNA-damaging agents. Modification of the small molecule and generation of an immobilized probe were used to selectively bind putative protein target(s) responsible for the observed activity. The second example describes a focused library approach that relied on tandem multicomponent reaction methodologies to afford a series of modulators of the heat shock protein 70 (Hsp70) molecular chaperone. The synthesis of libraries based on the structure of MAL3-101 generated a collection of chemotypes, each modulating Hsp70 function, but exhibiting divergent pharmacological activities. For example, probes that compromise the replication of a disease-associated polyomavirus were identified. These projects highlight the importance of chemical methodology development as a source of small-molecule probes and as a drug discovery starting point. PMID:21502524

Targeting Cullin–RING E3 ubiquitin ligases for drug discovery: structure, assembly and small-molecule modulation

PubMed Central

Bulatov, Emil; Ciulli, Alessio

2015-01-01

In the last decade, the ubiquitin–proteasome system has emerged as a valid target for the development of novel therapeutics. E3 ubiquitin ligases are particularly attractive targets because they confer substrate specificity on the ubiquitin system. CRLs [Cullin–RING (really interesting new gene) E3 ubiquitin ligases] draw particular attention, being the largest family of E3s. The CRLs assemble into functional multisubunit complexes using a repertoire of substrate receptors, adaptors, Cullin scaffolds and RING-box proteins. Drug discovery targeting CRLs is growing in importance due to mounting evidence pointing to significant roles of these enzymes in diverse biological processes and human diseases, including cancer, where CRLs and their substrates often function as tumour suppressors or oncogenes. In the present review, we provide an account of the assembly and structure of CRL complexes, and outline the current state of the field in terms of available knowledge of small-molecule inhibitors and modulators of CRL activity. A comprehensive overview of the reported crystal structures of CRL subunits, components and full-size complexes, alone or with bound small molecules and substrate peptides, is included. This information is providing increasing opportunities to aid the rational structure-based design of chemical probes and potential small-molecule therapeutics targeting CRLs. PMID:25886174

Small tyrosine kinase inhibitors interrupt EGFR signaling by interacting with erbB3 and erbB4 in glioblastoma cell lines

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

Carrasco-Garcia, Estefania; Saceda, Miguel; Unidad de Investigacion, Hospital General Universitario de Elche, 03203 Elche

Signaling through the epidermal growth factor receptor (EGFR) is relevant in glioblastoma. We have determined the effects of the EGFR inhibitor AG1478 in glioblastoma cell lines and found that U87 and LN-229 cells were very sensitive to this drug, since their proliferation diminished and underwent a marked G{sub 1} arrest. T98 cells were a little more refractory to growth inhibition and A172 cells did not undergo a G{sub 1} arrest. This G{sub 1} arrest was associated with up-regulation of p27{sup kip1}, whose protein turnover was stabilized. EGFR autophosphorylation was blocked with AG1478 to the same extent in all the cellmore » lines. Other small-molecule EGFR tyrosine kinase inhibitors employed in the clinic, such as gefitinib, erlotinib and lapatinib, were able to abrogate proliferation of glioblastoma cell lines, which underwent a G{sub 1} arrest. However, the EGFR monoclonal antibody, cetuximab had no effect on cell proliferation and consistently, had no effect on cell cycle either. Similarly, cetuximab did not inhibit proliferation of U87 {Delta}EGFR cells or primary glioblastoma cell cultures, whereas small-molecule EGFR inhibitors did. Activity of downstream signaling molecules of EGFR such as Akt and especially ERK1/2 was interrupted with EGFR tyrosine kinase inhibitors, whereas cetuximab treatment could not sustain this blockade over time. Small-molecule EGFR inhibitors were able to prevent phosphorylation of erbB3 and erbB4, whereas cetuximab only hindered EGFR phosphorylation, suggesting that EGFR tyrosine kinase inhibitors may mediate their anti-proliferative effects through other erbB family members. We can conclude that small-molecule EGFR inhibitors may be a therapeutic approach for the treatment of glioblastoma patients.« less

Effect of the collective motions of molecules inside a condensed phase on fluctuations in the density of small bodies

NASA Astrophysics Data System (ADS)

Tovbin, Yu. K.

2017-11-01

An approach to calculating the effects of fluctuations in density that considers the collective motions of molecules in small condensed phases (e.g., droplets, microcrystals, adsorption at microcrystal faces) is proposed. Statistical sums of the vibrational, rotational, and translational motions of molecules are of a collective character expressed in the dependences of these statistical sums on the local configurations of neighboring molecules. This changes their individual contributions to the free energy and modifies fluctuations in density in the inner homogeneous regions of small bodies. Interactions between nearest neighbors are considered in a quasi-chemical approximation that reflects the effects of short-range direct correlations. Expressions for isotherms relating the densities of mixture components to the chemical potentials in a thermostat are obtained, along with equations for pair distribution functions.

Nanowire Electron Scattering Spectroscopy

NASA Technical Reports Server (NTRS)

Hunt, Brian; Bronikowsky, Michael; Wong, Eric; VonAllmen, Paul; Oyafuso, Fablano

2009-01-01

Nanowire electron scattering spectroscopy (NESS) has been proposed as the basis of a class of ultra-small, ultralow-power sensors that could be used to detect and identify chemical compounds present in extremely small quantities. State-of-the-art nanowire chemical sensors have already been demonstrated to be capable of detecting a variety of compounds in femtomolar quantities. However, to date, chemically specific sensing of molecules using these sensors has required the use of chemically functionalized nanowires with receptors tailored to individual molecules of interest. While potentially effective, this functionalization requires labor-intensive treatment of many nanowires to sense a broad spectrum of molecules. In contrast, NESS would eliminate the need for chemical functionalization of nanowires and would enable the use of the same sensor to detect and identify multiple compounds. NESS is analogous to Raman spectroscopy, the main difference being that in NESS, one would utilize inelastic scattering of electrons instead of photons to determine molecular vibrational energy levels. More specifically, in NESS, one would exploit inelastic scattering of electrons by low-lying vibrational quantum states of molecules attached to a nanowire or nanotube.

Quantitative analyses of bifunctional molecules.

PubMed

Braun, Patrick D; Wandless, Thomas J

2004-05-11

Small molecules can be discovered or engineered to bind tightly to biologically relevant proteins, and these molecules have proven to be powerful tools for both basic research and therapeutic applications. In many cases, detailed biophysical analyses of the intermolecular binding events are essential for improving the activity of the small molecules. These interactions can often be characterized as straightforward bimolecular binding events, and a variety of experimental and analytical techniques have been developed and refined to facilitate these analyses. Several investigators have recently synthesized heterodimeric molecules that are designed to bind simultaneously with two different proteins to form ternary complexes. These heterodimeric molecules often display compelling biological activity; however, they are difficult to characterize. The bimolecular interaction between one protein and the heterodimeric ligand (primary dissociation constant) can be determined by a number of methods. However, the interaction between that protein-ligand complex and the second protein (secondary dissociation constant) is more difficult to measure due to the noncovalent nature of the original protein-ligand complex. Consequently, these heterodimeric compounds are often characterized in terms of their activity, which is an experimentally dependent metric. We have developed a general quantitative mathematical model that can be used to measure both the primary (protein + ligand) and secondary (protein-ligand + protein) dissociation constants for heterodimeric small molecules. These values are largely independent of the experimental technique used and furthermore provide a direct measure of the thermodynamic stability of the ternary complexes that are formed. Fluorescence polarization and this model were used to characterize the heterodimeric molecule, SLFpYEEI, which binds to both FKBP12 and the Fyn SH2 domain, demonstrating that the model is useful for both predictive as well as ex post facto analytical applications.

Incorporation of functionalized gold nanoparticles into nanofibers for enhanced attachment and differentiation of mammalian cells

PubMed Central

2012-01-01

Background Electrospun nanofibers have been widely used as substrata for mammalian cell culture owing to their structural similarity to natural extracellular matrices. Structurally consistent electrospun nanofibers can be produced with synthetic polymers but require chemical modification to graft cell-adhesive molecules to make the nanofibers functional. Development of a facile method of grafting functional molecules on the nanofibers will contribute to the production of diverse cell type-specific nanofiber substrata. Results Small molecules, peptides, and functionalized gold nanoparticles were successfully incorporated with polymethylglutarimide (PMGI) nanofibers through electrospinning. The PMGI nanofibers functionalized by the grafted AuNPs, which were labeled with cell-adhesive peptides, enhanced HeLa cell attachment and potentiated cardiomyocyte differentiation of human pluripotent stem cells. Conclusions PMGI nanofibers can be functionalized simply by co-electrospinning with the grafting materials. In addition, grafting functionalized AuNPs enable high-density localization of the cell-adhesive peptides on the nanofiber. The results of the present study suggest that more cell type-specific synthetic substrata can be fabricated with molecule-doped nanofibers, in which diverse functional molecules are grafted alone or in combination with other molecules at different concentrations. PMID:22686683

Mass amplifying probe for sensitive fluorescence anisotropy detection of small molecules in complex biological samples.

PubMed

Cui, Liang; Zou, Yuan; Lin, Ninghang; Zhu, Zhi; Jenkins, Gareth; Yang, Chaoyong James

2012-07-03

Fluorescence anisotropy (FA) is a reliable and excellent choice for fluorescence sensing. One of the key factors influencing the FA value for any molecule is the molar mass of the molecule being measured. As a result, the FA method with functional nucleic acid aptamers has been limited to macromolecules such as proteins and is generally not applicable for the analysis of small molecules because their molecular masses are relatively too small to produce observable FA value changes. We report here a molecular mass amplifying strategy to construct anisotropy aptamer probes for small molecules. The probe is designed in such a way that only when a target molecule binds to the probe does it activate its binding ability to an anisotropy amplifier (a high molecular mass molecule such as protein), thus significantly increasing the molecular mass and FA value of the probe/target complex. Specifically, a mass amplifying probe (MAP) consists of a targeting aptamer domain against a target molecule and molecular mass amplifying aptamer domain for the amplifier protein. The probe is initially rendered inactive by a small blocking strand partially complementary to both target aptamer and amplifier protein aptamer so that the mass amplifying aptamer domain would not bind to the amplifier protein unless the probe has been activated by the target. In this way, we prepared two probes that constitute a target (ATP and cocaine respectively) aptamer, a thrombin (as the mass amplifier) aptamer, and a fluorophore. Both probes worked well against their corresponding small molecule targets, and the detection limits for ATP and cocaine were 0.5 μM and 0.8 μM, respectively. More importantly, because FA is less affected by environmental interferences, ATP in cell media and cocaine in urine were directly detected without any tedious sample pretreatment. Our results established that our molecular mass amplifying strategy can be used to design aptamer probes for rapid, sensitive, and selective detection of small molecules by means of FA in complex biological samples.

Calcium phosphate nanoparticles as versatile carrier for small and large molecules across cell membranes

NASA Astrophysics Data System (ADS)

Sokolova, Viktoriya; Rotan, Olga; Klesing, Jan; Nalbant, Perihan; Buer, Jan; Knuschke, Torben; Westendorf, Astrid M.; Epple, Matthias

2012-06-01

The successful transport of molecules across the cell membrane is a key point in biology and medicine. In most cases, molecules alone cannot penetrate the cell membrane, therefore an efficient carrier is needed. Calcium phosphate nanoparticles (diameter: 100-250 nm, depending on the functionalization) were loaded with fluorescent oligonucleotides, peptide, proteins, antibodies, polymers or porphyrins and characterized by dynamic light scattering, nanoparticle tracking analysis and scanning electron microscopy. Any excess of molecules was removed by ultracentrifugation, and the dissolved molecules at the same concentration were used as control. The uptake of such fluorescence-labeled nanoparticles into HeLa cells was monitored by fluorescence microscopy and confocal laser scanning microscopy. Calcium phosphate nanoparticles were able to transport all molecules across the cell membrane, whereas the dissolved molecules alone were taken up only to a very small extent or even not at all.

The role of MicroRNA molecules and MicroRNA-regulating machinery in the pathogenesis and progression of epithelial ovarian cancer.

PubMed

Wang, Xiyin; Ivan, Mircea; Hawkins, Shannon M

2017-11-01

MicroRNA molecules are small, single-stranded RNA molecules that function to regulate networks of genes. They play important roles in normal female reproductive tract biology, as well as in the pathogenesis and progression of epithelial ovarian cancer. DROSHA, DICER, and Argonaute proteins are components of the microRNA-regulatory machinery and mediate microRNA production and function. This review discusses aberrant expression of microRNA molecules and microRNA-regulating machinery associated with clinical features of epithelial ovarian cancer. Understanding the regulation of microRNA molecule production and function may facilitate the development of novel diagnostic and therapeutic strategies to improve the prognosis of women with epithelial ovarian cancer. Additionally, understanding microRNA molecules and microRNA-regulatory machinery associations with clinical features may influence prevention and early detection efforts. Copyright © 2017 Elsevier Inc. All rights reserved.

Single-Molecule Localization Microscopy allows for the analysis of cancer metastasis-specific miRNA distribution on the nanoscale

PubMed Central

Tezcan, Kerem Can; Schaufler, Wladimir; Bestvater, Felix; Patil, Nitin; Birk, Udo; Hafner, Mathias; Altevogt, Peter; Cremer, Christoph; Allgayer, Heike

2015-01-01

We describe a novel approach for the detection of small non-coding RNAs in single cells by Single-Molecule Localization Microscopy (SMLM). We used a modified SMLM–setup and applied this instrument in a first proof-of-principle concept to human cancer cell lines. Our method is able to visualize single microRNA (miR)-molecules in fixed cells with a localization accuracy of 10–15 nm, and is able to quantify and analyse clustering and localization in particular subcellular sites, including exosomes. We compared the metastasis-site derived (SW620) and primary site derived (SW480) human colorectal cancer (CRC) cell lines, and (as a proof of principle) evaluated the metastasis relevant miR-31 as a first example. We observed that the subcellular distribution of miR-31 molecules in both cell lines was very heterogeneous with the largest subpopulation of optically acquired weakly metastatic cells characterized by a low number of miR-31 molecules, as opposed to a significantly higher number in the majority of the highly metastatic cells. Furthermore, the highly metastatic cells had significantly more miR-31-molecules in the extracellular space, which were visualized to co-localize with exosomes in significantly higher numbers. From this study, we conclude that miRs are not only aberrantly expressed and regulated, but also differentially compartmentalized in cells with different metastatic potential. Taken together, this novel approach, by providing single molecule images of miRNAs in cellulo can be used as a powerful supplementary tool in the analysis of miRNA function and behaviour and has far reaching potential in defining metastasis-critical subpopulations within a given heterogeneous cancer cell population. PMID:26561203

Magnetoassociation of KRb Feshbach molecules

NASA Astrophysics Data System (ADS)

Cumby, Tyler; Perreault, John; Shewmon, Ruth; Jin, Deborah

2010-03-01

I will discuss experiments in which we study the creation of ^40K^87Rb Feshbach molecules via magnetoassociation. We measure the molecule number as a function of the magnetic-field sweep rate through the interspecies Feshbach resonance and explore the dependence of association on the initial atom gas conditions. This study of the Feshbach molecule creation process may be relevant to the production of ultracold polar molecules, where magnetoassociated Feshbach molecules can be a crucial first step [1].[4pt] [1] K.-K. Ni, S. Ospelkaus, M. H. G. de Miranda, A. Peer, B. Neyenhuis, J. J. Zirbel, S. Kotochigova, P. S. Julienne, D. S. Jin, and J. Ye, Science, 2008, 322, 231- 235.

Magnetoassociation of KRb Feshbach molecules

NASA Astrophysics Data System (ADS)

Cumby, Tyler; Perreault, John; Shewmon, Ruth; Jin, Deborah

2010-03-01

I will discuss experiments in which we study the creation of ^40K^87Rb Feshbach molecules via magnetoassociation. We measure the molecule number as a function of the magnetic-field sweep rate through the interspecies Feshbach resonance and explore the dependence of association on the initial atom gas conditions. This study of the Feshbach molecule creation process may be relevant to the production of ultracold polar molecules, where magnetoassociated Feshbach molecules can be a crucial first step [1].[4pt] [1] K.-K. Ni, S. Ospelkaus, M. H. G. de Miranda, A. Peer, B. Neyenhuis, J. J. Zirbel, S. Kotochigova, P. S. Julienne, D. S. Jin, and J. Ye, Science, 2008, 322, 231-235.

Experimental Evidence for Quantum Interference and Vibrationally Induced Decoherence in Single-Molecule Junctions

NASA Astrophysics Data System (ADS)

Ballmann, Stefan; Härtle, Rainer; Coto, Pedro B.; Elbing, Mark; Mayor, Marcel; Bryce, Martin R.; Thoss, Michael; Weber, Heiko B.

2012-08-01

We analyze quantum interference and decoherence effects in single-molecule junctions both experimentally and theoretically by means of the mechanically controlled break junction technique and density-functional theory. We consider the case where interference is provided by overlapping quasidegenerate states. Decoherence mechanisms arising from electronic-vibrational coupling strongly affect the electrical current flowing through a single-molecule contact and can be controlled by temperature variation. Our findings underline the universal relevance of vibrations for understanding charge transport through molecular junctions.

Experimental evidence for quantum interference and vibrationally induced decoherence in single-molecule junctions.

PubMed

Ballmann, Stefan; Härtle, Rainer; Coto, Pedro B; Elbing, Mark; Mayor, Marcel; Bryce, Martin R; Thoss, Michael; Weber, Heiko B

2012-08-03

We analyze quantum interference and decoherence effects in single-molecule junctions both experimentally and theoretically by means of the mechanically controlled break junction technique and density-functional theory. We consider the case where interference is provided by overlapping quasidegenerate states. Decoherence mechanisms arising from electronic-vibrational coupling strongly affect the electrical current flowing through a single-molecule contact and can be controlled by temperature variation. Our findings underline the universal relevance of vibrations for understanding charge transport through molecular junctions.

A small molecule screen identifies a novel compound that induces a homeotic transformation in Hydra

PubMed Central

Glauber, Kristine M.; Dana, Catherine E.; Park, Steve S.; Colby, David A.; Noro, Yukihiko; Fujisawa, Toshitaka; Chamberlin, A. Richard; Steele, Robert E.

2013-01-01

Developmental processes such as morphogenesis, patterning and differentiation are continuously active in the adult Hydra polyp. We carried out a small molecule screen to identify compounds that affect patterning in Hydra. We identified a novel molecule, DAC-2-25, that causes a homeotic transformation of body column into tentacle zone. This transformation occurs in a progressive and polar fashion, beginning at the oral end of the animal. We have identified several strains that respond to DAC-2-25 and one that does not, and we used chimeras from these strains to identify the ectoderm as the target tissue for DAC-2-25. Using transgenic Hydra that express green fluorescent protein under the control of relevant promoters, we examined how DAC-2-25 affects tentacle patterning. Genes whose expression is associated with the tentacle zone are ectopically expressed upon exposure to DAC-2-25, whereas those associated with body column tissue are turned off as the tentacle zone expands. The expression patterns of the organizer-associated gene HyWnt3 and the hypostome-specific gene HyBra2 are unchanged. Structure-activity relationship studies have identified features of DAC-2-25 that are required for activity and potency. This study shows that small molecule screens in Hydra can be used to dissect patterning processes. PMID:24255098

A small molecule screen identifies a novel compound that induces a homeotic transformation in Hydra.

PubMed

Glauber, Kristine M; Dana, Catherine E; Park, Steve S; Colby, David A; Noro, Yukihiko; Fujisawa, Toshitaka; Chamberlin, A Richard; Steele, Robert E

2013-12-01

Developmental processes such as morphogenesis, patterning and differentiation are continuously active in the adult Hydra polyp. We carried out a small molecule screen to identify compounds that affect patterning in Hydra. We identified a novel molecule, DAC-2-25, that causes a homeotic transformation of body column into tentacle zone. This transformation occurs in a progressive and polar fashion, beginning at the oral end of the animal. We have identified several strains that respond to DAC-2-25 and one that does not, and we used chimeras from these strains to identify the ectoderm as the target tissue for DAC-2-25. Using transgenic Hydra that express green fluorescent protein under the control of relevant promoters, we examined how DAC-2-25 affects tentacle patterning. Genes whose expression is associated with the tentacle zone are ectopically expressed upon exposure to DAC-2-25, whereas those associated with body column tissue are turned off as the tentacle zone expands. The expression patterns of the organizer-associated gene HyWnt3 and the hypostome-specific gene HyBra2 are unchanged. Structure-activity relationship studies have identified features of DAC-2-25 that are required for activity and potency. This study shows that small molecule screens in Hydra can be used to dissect patterning processes.

Small-Molecule Fluorescent Sensors for Investigating Zinc Metalloneurochemistry

PubMed Central

Nolan, Elizabeth M.; Lippard, Stephen J.

2008-01-01

Conspectus Metal ions are involved in many neurobiological processes relevant to human health and disease. The metalloneurochemistry of Zn(II) is of substantial current interest. Zinc is the second most abundant d-block metal ion in the human brain and its distribution varies, with relatively high concentrations found in the hippocampus. Brain zinc is generally divided into two categories: protein-bound and loosely-bound. The latter pool is also referred to as histochemically observable, chelatable, labile, or mobile zinc. The neurophysiological and neuropathological significance of such mobile Zn(II) remains enigmatic. Studies of Zn(II) distribution, translocation, and function in vivo require tools for its detection. Because Zn(II) has a closed-shell d10 configuration and no convenient spectroscopic signature, fluorescence is a suitable method for monitoring Zn(II) in biological contexts. This Account summarizes work by our laboratory addressing the design, preparation, characterization, and use of small-molecule fluorescent sensors for imaging mobile Zn(II) in living cells and samples of brain tissue. These sensors provide “turn-on” or ratiometric Zn(II) detection in aqueous solution at neutral pH. By making alterations to the Zn(II)-binding unit and fluorophore platform, we have devised sensors with varied photophysical and metal-binding properties. We used several of these probes to image Zn(II) distribution, uptake, and mobilization in a variety of cell types, including neuronal cultures. Goals for the future include developing strategies for multi-color imaging, further defining the quenching and turn-on mechanisms of the sensors, and employing the probes to elucidate the functional significance of Zn(II) in neurobiology. PMID:18989940

Mechanisms of Aquaporin-Facilitated Cancer Invasion and Metastasis

NASA Astrophysics Data System (ADS)

De Ieso, Michael L.; Yool, Andrea J.

2018-04-01

Cancer is a leading cause of death worldwide, and its incidence is rising with numbers expected to increase 70% in the next two decades. The fact that current mainline treatments for cancer patients are accompanied by debilitating side effects prompts a growing demand for new therapies that not only inhibit growth and proliferation of cancer cells, but also control invasion and metastasis. One class of targets gaining international attention is the aquaporins, a family of membrane-spanning water channels with diverse physiological functions and extensive tissue-specific distributions in humans. Aquaporins -1, -2, -3, -4, -5, -8, and -9 have been linked to roles in cancer proliferation, invasion and metastasis, but their mechanisms of action remain to be fully defined. Aquaporins are implicated in the metastatic cascade in processes of angiogenesis, cellular dissociation, migration and invasion. Cancer invasion and metastasis are proposed to be potentiated by aquaporins in boosting tumor angiogenesis, enhancing cell volume regulation, regulating cell-cell and cell-matrix adhesions, interacting with actin cytoskeleton, regulating proteases and extracellular-matrix degrading molecules, contributing to the regulation of epithelial-mesenchymal transitions, and interacting with signaling pathways enabling motility and invasion. Pharmacological modulators of aquaporin channels are being identified and tested for therapeutic potential, including compounds derived from loop diuretics, metal-containing organic compounds, plant natural products, and other small molecules. Further studies on aquaporin-dependent functions in cancer metastasis are needed to define the differential contributions of different classes of aquaporin channels to regulation of fluid balance, cell volume, small solute transport, signal transduction, their possible relevance as rate limiting steps, and potential values as therapeutic targets for proliferation and invasion.

KRAS: Reasons for optimism in lung cancer.

PubMed

Lindsay, C R; Jamal-Hanjani, M; Forster, M; Blackhall, F

2018-06-09

Despite being the most frequent gain-of-function genetic alteration in human cancer, KRAS mutation has to date offered only limited potential as a prognostic and predictive biomarker. Results from the phase III SELECT-1 trial in non-small cell lung cancer (NSCLC) recently added to a number of historical and more contemporary disappointments in targeting KRAS mutant disease, including farnesyl transferase inhibition and synthetic lethality partners such as STK33. This narrative review uses the context of these previous failures to demonstrate how the knowledge gained from these experiences can be used as a platform for exciting advances in NSCLC on the horizon. It now seems clear that mutational subtype (most commonly G12C) of individual mutations is of greater relevance than the categorical evaluation of KRAS mutation presence or otherwise. A number of direct small molecules targeted to these subtypes are in development and have shown promising biological activity, with some in the late stages of preclinical validation. Copyright © 2018 Elsevier Ltd. All rights reserved.

Helix-Grafted Pleckstrin Homology Domains Suppress HIV-1 Infection of CD4-Positive Cells.

PubMed

Tennyson, Rachel L; Walker, Susanne N; Ikeda, Terumasa; Harris, Reuben S; Kennan, Alan J; McNaughton, Brian R

2016-10-17

The size, functional group diversity and three-dimensional structure of proteins often allow these biomolecules to bind disease-relevant structures that challenge or evade small-molecule discovery. Additionally, folded proteins are often much more stable in biologically relevant environments compared to their peptide counterparts. We recently showed that helix-grafted display-extensive resurfacing and elongation of an existing solvent-exposed helix in a pleckstrin homology (PH) domain-led to a new protein that binds a surrogate of HIV-1 gp41, a validated target for inhibition of HIV-1 entry. Expanding on this work, we prepared a number of human-derived helix-grafted-display PH domains of varied helix length and measured properties relevant to therapeutic and basic research applications. In particular, we showed that some of these new reagents expressed well as recombinant proteins in Escherichia coli, were relatively stable in human serum, bound a mimic of pre-fusogenic HIV-1 gp41 in vitro and in complex biological environments, and significantly lowered the incidence of HIV-1 infection of CD4-positive cells. © 2016 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Aldolase-catalysed stereoselective synthesis of fluorinated small molecules.

PubMed

Windle, Claire L; Berry, Alan; Nelson, Adam

2017-04-01

The introduction of fluorine has been widely exploited to tune the biological functions of small molecules. Indeed, around 20% of leading drugs contain at least one fluorine atom. Yet, despite profound effects of fluorination on conformation, there is only a limited toolkit of reactions that enable stereoselective synthesis of fluorinated compounds. Aldolases are useful catalysts for the stereoselective synthesis of bioactive small molecules; however, despite fluoropyruvate being a viable nucleophile for some aldolases, the potential of aldolases to control the formation of fluorine-bearing stereocentres has largely been untapped. Very recently, it has been shown that aldolase-catalysed stereoselective carboncarbon bond formation with fluoropyruvate as nucleophile enable the synthesis of many α-fluoro β-hydroxy carboxyl derivatives. Furthermore, an understanding of the structural basis for the stereocontrol observed in these reactions is beginning to emerge. Here, we review the application of aldolase catalysis in the stereocontrolled synthesis of chiral fluorinated small molecules, and highlight likely areas for future developments. Copyright © 2017 Elsevier Ltd. All rights reserved.

Elementary Reactions and Their Role in Gas-Phase Prebiotic Chemistry

PubMed Central

Balucani, Nadia

2009-01-01

The formation of complex organic molecules in a reactor filled with gaseous mixtures possibly reproducing the primitive terrestrial atmosphere and ocean demonstrated more than 50 years ago that inorganic synthesis of prebiotic molecules is possible, provided that some form of energy is provided to the system. After that groundbreaking experiment, gas-phase prebiotic molecules have been observed in a wide variety of extraterrestrial objects (including interstellar clouds, comets and planetary atmospheres) where the physical conditions vary widely. A thorough characterization of the chemical evolution of those objects relies on a multi-disciplinary approach: 1) observations allow us to identify the molecules and their number densities as they are nowadays; 2) the chemistry which lies behind their formation starting from atoms and simple molecules is accounted for by complex reaction networks; 3) for a realistic modeling of such networks, a number of experimental parameters are needed and, therefore, the relevant molecular processes should be fully characterized in laboratory experiments. A survey of the available literature reveals, however, that much information is still lacking if it is true that only a small percentage of the elementary reactions considered in the models have been characterized in laboratory experiments. New experimental approaches to characterize the relevant elementary reactions in laboratory are presented and the implications of the results are discussed. PMID:19564951

A transgenic resource for conditional competitive inhibition of conserved Drosophila microRNAs

PubMed Central

Fulga, Tudor A.; McNeill, Elizabeth M.; Binari, Richard; Yelick, Julia; Blanche, Alexandra; Booker, Matthew; Steinkraus, Bruno R.; Schnall-Levin, Michael; Zhao, Yong; DeLuca, Todd; Bejarano, Fernando; Han, Zhe; Lai, Eric C.; Wall, Dennis P.; Perrimon, Norbert; Van Vactor, David

2015-01-01

Although the impact of microRNAs (miRNAs) in development and disease is well established, understanding the function of individual miRNAs remains challenging. Development of competitive inhibitor molecules such as miRNA sponges has allowed the community to address individual miRNA function in vivo. However, the application of these loss-of-function strategies has been limited. Here we offer a comprehensive library of 141 conditional miRNA sponges targeting well-conserved miRNAs in Drosophila. Ubiquitous miRNA sponge delivery and consequent systemic miRNA inhibition uncovers a relatively small number of miRNA families underlying viability and gross morphogenesis, with false discovery rates in the 4–8% range. In contrast, tissue-specific silencing of muscle-enriched miRNAs reveals a surprisingly large number of novel miRNA contributions to the maintenance of adult indirect flight muscle structure and function. A strong correlation between miRNA abundance and physiological relevance is not observed, underscoring the importance of unbiased screens when assessing the contributions of miRNAs to complex biological processes. PMID:26081261

A framework for multi-scale simulation of crystal growth in the presence of polymers.

PubMed

Mandal, Taraknath; Huang, Wenjun; Mecca, Jodi M; Getchell, Ashley; Porter, William W; Larson, Ronald G

2017-03-01

We present a multi-scale simulation method for modeling crystal growth in the presence of polymer excipients. The method includes a coarse-grained (CG) model for small molecules of known crystal structure whose force field is obtained using structural properties from atomistic simulations. This CG model is capable of stabilizing the molecular crystal structure and capturing the crystal growth from the melt for a wide range of small organic molecules, as demonstrated by application of our method to the molecules isoniazid, urea, sulfamethoxazole, prilocaine, oxcarbazepine, and phenytoin. This CG model can also be used to study the effect of additives, such as polymers, on the inhibition of crystal growth by polymers, as exemplified by our simulation of suppression of the rate of crystal growth of phenytoin, an active pharmaceutical ingredient (API), by a cellulose excipient, functionalized with acetate (Ac), hydroxy-propyl (Hp) and succinate (Su) groups. We show that the efficacy of the cellulosic polymers in slowing crystal growth of small molecules strongly depends on the functional group substitution on the cellulose backbone, with the acetate substituent group slowing crystal growth more than does the deprotonated succinate group, which we confirm by experimental drug supersaturation studies.

Small molecules targeting heterotrimeric G proteins.

PubMed

Ayoub, Mohammed Akli

2018-05-05

G protein-coupled receptors (GPCRs) represent the largest family of cell surface receptors regulating many human and animal physiological functions. Their implication in human pathophysiology is obvious with almost 30-40% medical drugs commercialized today directly targeting GPCRs as molecular entities. However, upon ligand binding GPCRs signal inside the cell through many key signaling, adaptor and regulatory proteins, including various classes of heterotrimeric G proteins. Therefore, G proteins are considered interesting targets for the development of pharmacological tools that are able to modulate their interaction with the receptors, as well as their activation/deactivation processes. In this review, old attempts and recent advances in the development of small molecules that directly target G proteins will be described with an emphasis on their utilization as pharmacological tools to dissect the mechanisms of activation of GPCR-G protein complexes. These molecules constitute a further asset for research in the "hot" areas of GPCR biology, areas such as multiple G protein coupling/signaling, GPCR-G protein preassembly, and GPCR functional selectivity or bias. Moreover, this review gives a particular focus on studies in vitro and in vivo supporting the potential applications of such small molecules in various GPCR/G protein-related diseases. Copyright © 2018 Elsevier B.V. All rights reserved.

Small-Molecule Modulators of Methyl-Lysine Binding for the CBX7 Chromodomain

DOE PAGES

Ren, Chunyan; Morohashi, Keita; Plotnikov, Alexander N.; ...

2015-02-05

Chromobox homolog 7 (CBX7) plays an important role in gene transcription in a wide array of cellular processes, ranging from stem cell self-renewal and differentiation to tumor progression. CBX7 functions through its N-terminal chromodomain (ChD), which recognizes tri-methylated lysine 27 of histone 3 (H3K27me3), a conserved epigenetic mark that signifies gene transcriptional repression. Here in this study, we report discovery of small molecules that inhibit CBX7ChD binding to H3K27me3. Our crystal structures reveal the binding modes of these molecules that compete against H3K27me3 binding through interactions with key residues in the methyl-lysine binding pocket of CBX7ChD. We further show thatmore » a lead compound MS37452, derepresses transcription of Polycomb repressive complex target gene p16/CDKN2A by displacing CBX7 binding to the INK4A/ARF locus in prostate cancer cells. Ultimately, these small molecules have the potential to be developed into high-potency chemical modulators that target CBX7 functions in gene transcription in different disease pathways.« less

Excellent Long-Term Stability of Power Conversion Efficiency in Non-Fullerene-Based Polymer Solar Cells Bearing Tricyanovinylene-Functionalized n-Type Small Molecules.

PubMed

Ko, Eun Yi; Park, Gi Eun; Lee, Ji Hyung; Kim, Hyung Jong; Lee, Dae Hee; Ahn, Hyungju; Uddin, Mohammad Afsar; Woo, Han Young; Cho, Min Ju; Choi, Dong Hoon

2017-03-15

New small molecules having modified acceptor strength and π-conjugation length and containing dicyanovinylene (DCV) and tricyanovinylene (TCV) as a strongly electron-accepting unit with indacenodithiophene, IDT(DCV) 2 , IDT(TCV) 2 , and IDTT(TCV) 2 , were synthesized and studied in terms of their applicability to polymer solar cells with PTB7-Th as an electron-donating polymer. Intriguingly, the blended films containing IDT(TCV) 2 and IDTT(TCV) 2 exhibited superior shelf life stabilities of more than 1000 h without any reduction in the initial power conversion efficiency. The low-lying lowest unoccupied molecular orbital energy levels and robust internal morphologies of small TCV-containing molecules could afford excellent shelf life stability.

Coevolution of T-cell receptors with MHC and non-MHC ligands

PubMed Central

Castro, Caitlin C.; Luoma, Adrienne M.; Adams, Erin J.

2015-01-01

Summary The structure and amino acid diversity of the T-cell receptor (TCR), similar in nature to that of Fab portions of antibodies, would suggest these proteins have a nearly infinite capacity to recognize antigen. Yet all currently defined native T cells expressing an α and β chain in their TCR can only sense antigen when presented in the context of a major histocompatibility complex (MHC) molecule. This MHC molecule can be one of many that exist in vertebrates, presenting small peptide fragments, lipid molecules, or small molecule metabolites. Here we review the pattern of TCR recognition of MHC molecules throughout a broad sampling of species and T-cell lineages and also touch upon T cells that do not appear to require MHC presentation for their surveillance function. We review the diversity of MHC molecules and information on the corresponding T-cell lineages identified in divergent species. We also discuss TCRs with structural domains unlike that of conventional TCRs of mouse and human. By presenting this broad view of TCR sequence, structure, domain organization, and function, we seek to explore how this receptor has evolved across time and been selected for alternative antigen-recognition capabilities in divergent lineages. PMID:26284470

Sustained release carrier for adenosine triphosphate as signaling molecule.

PubMed

Wischke, Christian; Weigel, Judith; Bulavina, Larisa; Lendlein, Andreas

2014-12-10

Adenosine triphosphate (ATP) is a molecule with a fascinating variety of intracellular and extracellular biological functions that go far beyond energy metabolism. Due to its limited passive diffusion through biological membranes, controlled release systems may allow to interact with ATP-mediated extracellular processes. In this study, two release systems were explored to evaluate the capacity for either long-term or short-term release: (i) Poly[(rac-lactide)-co-glycolide] (PLGA) implant rods were capable of ATP release over days to weeks, depending on the PLGA molecular weight and end-group capping, but were also associated with partial hydrolytic degradation of ATP to ADP and AMP, but not adenosine. (ii) Thermosensitive methylcellulose hydrogels with a gelation occurring at body temperature allowed combining adjustable loading levels and the capacity for injection, with injection forces less than 50N even for small 27G needles. Finally, a first in vitro study illustrated purinergic-triggered response of primary murine microglia to ATP released from hydrogels, demonstrating the potential relevance for biomedical applications. Copyright © 2014 Elsevier B.V. All rights reserved.

Conformal Electroplating of Azobenzene-Based Solar Thermal Fuels onto Large-Area and Fiber Geometries.

PubMed

Zhitomirsky, David; Grossman, Jeffrey C

2016-10-05

There is tremendous growth in fields where small functional molecules and colloidal nanomaterials are integrated into thin films for solid-state device applications. Many of these materials are synthesized in solution and there often exists a significant barrier to transition them into the solid state in an efficient manner. Here, we develop a methodology employing an electrodepositable copolymer consisting of small functional molecules for applications in solar energy harvesting and storage. We employ azobenzene solar thermal fuel polymers and functionalize them to enable deposition from low concentration solutions in methanol, resulting in uniform and large-area thin films. This approach enables conformal deposition on a variety of conducting substrates that can be either flat or structured depending on the application. Our approach further enables control over film growth via electrodepsition conditions and results in highly uniform films of hundreds of nanometers to microns in thickness. We demonstrate that this method enables superior retention of solar thermal fuel properties, with energy densities of ∼90 J/g, chargeability in the solid state, and exceptional materials utilization compared to other solid-state processing approaches. This novel approach is applicable to systems such as photon upconversion, photovoltaics, photosensing, light emission, and beyond, where small functional molecules enable solid-state applications.

Shaping Small Bioactive Molecules to Untangle Their Biological Function: A Focus on Fluorescent Plant Hormones.

PubMed

Lace, Beatrice; Prandi, Cristina

2016-08-01

Modern biology overlaps with chemistry in explaining the structure and function of all cellular processes at the molecular level. Plant hormone research is perfectly located at the interface between these two disciplines, taking advantage of synthetic and computational chemistry as a tool to decipher the complex biological mechanisms regulating the action of plant hormones. These small signaling molecules regulate a wide range of developmental processes, adapting plant growth to ever changing environmental conditions. The synthesis of small bioactive molecules mimicking the activity of endogenous hormones allows us to unveil many molecular features of their functioning, giving rise to a new field, plant chemical biology. In this framework, fluorescence labeling of plant hormones is emerging as a successful strategy to track the fate of these challenging molecules inside living organisms. Thanks to the increasing availability of new fluorescent probes as well as advanced and innovative imaging technologies, we are now in a position to investigate many of the dynamic mechanisms through which plant hormones exert their action. Such a deep and detailed comprehension is mandatory for the development of new green technologies for practical applications. In this review, we summarize the results obtained so far concerning the fluorescent labeling of plant hormones, highlighting the basic steps leading to the design and synthesis of these compelling molecular tools and their applications. Copyright © 2016 The Author. Published by Elsevier Inc. All rights reserved.

Small-molecule pheromones that control dauer development in Caenorhabditis elegans.

PubMed

Butcher, Rebecca A; Fujita, Masaki; Schroeder, Frank C; Clardy, Jon

2007-07-01

In response to high population density or low food supply, the nematode Caenorhabditis elegans enters an alternative larval stage, known as the dauer, that can withstand adverse conditions for prolonged periods. C. elegans senses its population density through a small-molecule signal, traditionally called the dauer pheromone, that it secretes into its surroundings. Here we show that the dauer pheromone consists of several structurally related ascarosides-derivatives of the dideoxysugar ascarylose-and that two of these ascarosides (1 and 2) are roughly two orders of magnitude more potent at inducing dauer formation than a previously reported dauer pheromone component (3) and constitute a physiologically relevant signal. The identification of dauer pheromone components 1 and 2 will facilitate the identification of target receptors and downstream signaling proteins.

Small-molecule modulators of PXR and CAR

PubMed Central

Chai, Sergio C.; Cherian, Milu T.; Wang, Yue-Ming; Chen, Taosheng

2016-01-01

Two nuclear receptors, the pregnane X receptor (PXR) and the constitutive androstane receptor (CAR), participate in the xenobiotic detoxification system by regulating the expression of drug-metabolizing enzymes and transporters in order to degrade and excrete foreign chemicals or endogenous metabolites. This review aims to expand the perceived relevance of PXR and CAR beyond their established role as master xenosensors to disease-oriented areas, emphasizing their modulation by small molecules. Structural studies of these receptors have provided much-needed insight into the nature of their binding promiscuity and the important elements that lead to ligand binding. Reports of species- and isoform-selective activation highlight the need for further scrutiny when extrapolating from animal data to humans, as animal models are at the forefront of early drug discovery. PMID:26921498

Micro and Nano Material Carriers for Immunomodulation.

PubMed

Bracho-Sanchez, E; Xia, C Q; Clare-Salzler, M J; Keselowsky, B G

2016-12-01

Modulation of the immune system through the use of micro and nano carriers offers opportunities in transplant tolerance, autoimmunity, infectious disease, and cancer. In particular, polymeric, lipid, and inorganic materials have been used as carriers of proteins, nucleic acids, and small drug molecules to direct the immune system toward either suppressive or stimulatory states. Current technologies have focused on the use of particulates or scaffolds, the modulation of materials properties, and the delivery of biologics or small drug molecules to achieve a desired response. Discussed are relevant immunology concepts, the types of biomaterial carriers used for immunomodulation highlighting their benefits and drawbacks, the material properties influencing immune responses, and recent examples in the field of transplant tolerance. © Copyright 2016 The American Society of Transplantation and the American Society of Transplant Surgeons.

Design and synthesis of type-III mimetics of ShK toxin

NASA Astrophysics Data System (ADS)

Baell, Jonathan B.; Harvey, Andrew J.; Norton, Raymond S.

2002-04-01

ShK toxin is a structurally defined, 35-residue polypeptide which blocks the voltage-gated Kv1.3 potassium channel in T-lymphocytes and has been identified as a possible immunosuppressant. Our interest lies in the rational design and synthesis of type-III mimetics of protein and polypeptide structure and function. ShK toxin is a challenging target for mimetic design as its binding epitope consists of relatively weakly binding residues, some of which are discontinuous. We discuss here our investigations into the design and synthesis of 1st generation, small molecule mimetics of ShK toxin and highlight any principles relevant to the generic design of type-III mimetics of continuous and discontinuous binding epitopes. We complement our approach with attempted pharmacophore-based database mining.

Development and maintenance of intestinal regulatory T cells.

PubMed

Tanoue, Takeshi; Atarashi, Koji; Honda, Kenya

2016-05-01

Gut-resident forkhead box P3 (FOXP3)(+)CD4(+) regulatory T cells (Treg cells) are distinct from those in other organs and have gut-specific phenotypes and functions. Whereas Treg cells in other organs have T cell receptors (TCRs) specific for self antigens, intestinal Treg cells have a distinct set of TCRs that are specific for intestinal antigens, and these cells have pivotal roles in the suppression of immune responses against harmless dietary antigens and commensal microorganisms. The differentiation, migration and maintenance of intestinal Treg cells are controlled by specific signals from the local environment. In particular, certain members of the microbiota continuously provide antigens and immunoregulatory small molecules that modulate intestinal Treg cells. Understanding the development and the maintenance of intestinal Treg cells provides important insights into disease-relevant host-microorganism interactions.

Structural Design Principle of Small-Molecule Organic Semiconductors for Metal-Free, Visible-Light-Promoted Photocatalysis.

PubMed

Wang, Lei; Huang, Wei; Li, Run; Gehrig, Dominik; Blom, Paul W M; Landfester, Katharina; Zhang, Kai A I

2016-08-08

Herein, we report on the structural design principle of small-molecule organic semiconductors as metal-free, pure organic and visible light-active photocatalysts. Two series of electron-donor and acceptor-type organic semiconductor molecules were synthesized to meet crucial requirements, such as 1) absorption range in the visible region, 2) sufficient photoredox potential, and 3) long lifetime of photogenerated excitons. The photocatalytic activity was demonstrated in the intermolecular C-H functionalization of electron-rich heteroaromates with malonate derivatives. A mechanistic study of the light-induced electron transport between the organic photocatalyst, substrate, and the sacrificial agent are described. With their tunable absorption range and defined energy-band structure, the small-molecule organic semiconductors could offer a new class of metal-free and visible light-active photocatalysts for chemical reactions. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Multifunctional and biologically active matrices from multicomponent polymeric solutions

NASA Technical Reports Server (NTRS)

Kiick, Kristi L. (Inventor); Yamaguchi, Nori (Inventor)

2010-01-01

The present invention relates to a biologically active functionalized electrospun matrix to permit immobilization and long-term delivery of biologically active agents. In particular the invention relates to a functionalized polymer matrix comprising a matrix polymer, a compatibilizing polymer and a biomolecule or other small functioning molecule. In certain aspects the electrospun polymer fibers comprise at least one biologically active molecule functionalized with low molecular weight heparin. Examples of active molecules that may be used with the multicomponent polymer of the invention include, for example, a drug, a biopolymer, for example a growth factor, a protein, a peptide, a nucleotide, a polysaccharide, a biological macromolecule or the like. The invention is further directed to the formation of functionalized crosslinked matrices, such as hydrogels, that include at least one functionalized compatibilizing polymer capable of assembly.

Update in TSH Receptor Agonists and Antagonists

PubMed Central

Neumann, Susanne

2012-01-01

The physiological role of the TSH receptor (TSHR) as a major regulator of thyroid function is well understood, but TSHRs are also expressed in multiple normal extrathyroidal tissues, and the physiological roles of TSHRs in these tissues are unclear. Moreover, TSHRs play a major role in several pathological conditions including hyperthyroidism, hypothyroidism, and thyroid tumors. Small molecule, “drug-like” TSHR agonists, neutral antagonists, and inverse agonists may be useful as probes of TSHR function in extrathyroidal tissues and as leads to develop drugs for several diseases of the thyroid. In this Update, we review the most recent findings regarding the development and use of these small molecule TSHR ligands. PMID:23019348

Elucidating compound mechanism of action by network perturbation analysis | Office of Cancer Genomics

Cancer.gov

Genome-wide identification of the mechanism of action (MoA) of small-molecule compounds characterizing their targets, effectors, and activity modulators represents a highly relevant yet elusive goal, with critical implications for assessment of compound efficacy and toxicity. Current approaches are labor intensive and mostly limited to elucidating high-affinity binding target proteins.

Features of Modularly Assembled Compounds That Impart Bioactivity Against an RNA Target

PubMed Central

Rzuczek, Suzanne G.; Gao, Yu; Tang, Zhen-Zhi; Thornton, Charles A.; Kodadek, Thomas; Disney, Matthew D.

2013-01-01

Transcriptomes provide a myriad of potential RNAs that could be the targets of therapeutics or chemical genetic probes of function. Cell permeable small molecules, however, generally do not exploit these targets, owing to the difficulty in the design of high affinity, specific small molecules targeting RNA. As part of a general program to study RNA function using small molecules, we designed bioactive, modularly assembled small molecules that target the non-coding expanded RNA repeat that causes myotonic dystrophy type 1 (DM1), r(CUG)exp. Herein, we present a rigorous study to elucidate features in modularly assembled compounds that afford bioactivity. Different modular assembly scaffolds were investigated including polyamines, α-peptides, β-peptides, and peptide tertiary amides (PTAs). Based on activity as assessed by improvement of DM1-associated defects, stability against proteases, cellular permeability, and toxicity, we discovered that constrained backbones, namely PTAs, are optimal. Notably, we determined that r(CUG)exp is the target of the optimal PTA in cellular models and that the optimal PTA improves DM1-associated defects in a mouse model. Biophysical analyses were employed to investigate potential sources of bioactivity. These investigations show that modularly assembled compounds have increased residence times on their targets and faster on rates than the RNA-binding modules from which they were derived and faster on rates than the protein that binds r(CUG)exp, the inactivation of which gives rise to DM1-associated defects. These studies provide information about features of small molecules that are programmable for targeting RNA, allowing for the facile optimization of therapeutics or chemical probes against other cellular RNA targets. PMID:24032410

Features of modularly assembled compounds that impart bioactivity against an RNA target.

PubMed

Rzuczek, Suzanne G; Gao, Yu; Tang, Zhen-Zhi; Thornton, Charles A; Kodadek, Thomas; Disney, Matthew D

2013-10-18

Transcriptomes provide a myriad of potential RNAs that could be the targets of therapeutics or chemical genetic probes of function. Cell-permeable small molecules, however, generally do not exploit these targets, owing to the difficulty in the design of high affinity, specific small molecules targeting RNA. As part of a general program to study RNA function using small molecules, we designed bioactive, modularly assembled small molecules that target the noncoding expanded RNA repeat that causes myotonic dystrophy type 1 (DM1), r(CUG)(exp). Herein, we present a rigorous study to elucidate features in modularly assembled compounds that afford bioactivity. Different modular assembly scaffolds were investigated, including polyamines, α-peptides, β-peptides, and peptide tertiary amides (PTAs). On the basis of activity as assessed by improvement of DM1-associated defects, stability against proteases, cellular permeability, and toxicity, we discovered that constrained backbones, namely, PTAs, are optimal. Notably, we determined that r(CUG)(exp) is the target of the optimal PTA in cellular models and that the optimal PTA improves DM1-associated defects in a mouse model. Biophysical analyses were employed to investigate potential sources of bioactivity. These investigations show that modularly assembled compounds have increased residence times on their targets and faster on rates than the RNA-binding modules from which they were derived. Moreover, they have faster on rates than the protein that binds r(CUG)(exp), the inactivation of which gives rise to DM1-associated defects. These studies provide information about features of small molecules that are programmable for targeting RNA, allowing for the facile optimization of therapeutics or chemical probes against other cellular RNA targets.

Substrates for Neuronal Cotransmission With Neuropeptides and Small Molecule Neurotransmitters in Drosophila

PubMed Central

Nässel, Dick R.

2018-01-01

It has been known for more than 40 years that individual neurons can produce more than one neurotransmitter and that neuropeptides often are colocalized with small molecule neurotransmitters (SMNs). Over the years much progress has been made in understanding the functional consequences of cotransmission in the nervous system of mammals. There are also some excellent invertebrate models that have revealed roles of coexpressed neuropeptides and SMNs in increasing complexity, flexibility, and dynamics in neuronal signaling. However, for the fly Drosophila there are surprisingly few functional studies on cotransmission, although there is ample evidence for colocalization of neuroactive compounds in neurons of the CNS, based both on traditional techniques and novel single cell transcriptome analysis. With the hope to trigger interest in initiating cotransmission studies, this review summarizes what is known about Drosophila neurons and neuronal circuits where different neuropeptides and SMNs are colocalized. Coexistence of neuroactive substances has been recorded in different neuron types such as neuroendocrine cells, interneurons, sensory cells and motor neurons. Some of the circuits highlighted here are well established in the analysis of learning and memory, circadian clock networks regulating rhythmic activity and sleep, as well as neurons and neuroendocrine cells regulating olfaction, nociception, feeding, metabolic homeostasis, diuretic functions, reproduction, and developmental processes. One emerging trait is the broad role of short neuropeptide F in cotransmission and presynaptic facilitation in a number of different neuronal circuits. This review also discusses the functional relevance of coexisting peptides in the intestine. Based on recent single cell transcriptomics data, it is likely that the neuronal systems discussed in this review are just a fraction of the total set of circuits where cotransmission occurs in Drosophila. Thus, a systematic search for colocalized neuroactive compounds in further neurons in anatomically defined circuits is of interest for the near future. PMID:29651236

Demonstration of sub-femtomole sensitivity for small molecules with microsphere ring resonator sensors

NASA Astrophysics Data System (ADS)

White, Ian M.; Oveys, Hesam; Fan, Xudong

2006-02-01

Optical microsphere resonators can function as highly sensitive bio/chemical sensors due to the large Q-factor, which leads to high light-matter interaction. The whispering gallery modes (WGM) arise at the surface of the microsphere, creating a highly enhanced optical field that interacts with matter on or near the microsphere surface. As a result, the spectral position of the WGM is extremely sensitive to refractive index changes near the surface, such as when bio/chemical molecules bind to the sphere. We show the potential feasibility of a microsphere ring resonator as a sensor for small molecules by demonstrating detection of sub-femtomole changes in SiO II molecules at the surface of the microsphere. In this experiment, the silica molecules act as an excellent model for small molecule analytes because of their 60 Dalton molecular weight, and because we know nearly the exact quantity of molecules at the surface, which enables a sensitivity characterization. We measure the spectral shifts in the WGMs when low concentrations of hydrofluoric acid (HF) are added to a solution that is being probed by the microsphere. As the HF molecules break apart the SiO II molecules at the sphere surface, the WGMs shift due to the sub-nano-scale decrease in the size of the microsphere. These calculations show that the sensitivity of this microsphere resonator is on the order of 500 attomoles. Our results will lead to the utilization of optical microspheres for detection of trace quantities of small molecules for such applications as drug discovery, environmental monitoring, and enzyme detection using peptide cleavage.

Small Molecule Fluoride Toxicity Agonists

PubMed Central

Nelson1, James W.; Plummer, Mark S.; Blount, Kenneth F.; Ames, Tyler D.; Breaker, Ronald R.

2015-01-01

SUMMARY Fluoride is a ubiquitous anion that inhibits a wide variety of metabolic processes. Here we report the identification of a series of compounds that enhance fluoride toxicity in Escherichia coli and Streptococcus mutans. These molecules were isolated by using a high-throughput screen (HTS) for compounds that increase intracellular fluoride levels as determined via a fluoride riboswitch-reporter fusion construct. A series of derivatives were synthesized to examine structure-activity relationships, leading to the identification of compounds with improved activity. Thus, we demonstrate that small molecule fluoride toxicity agonists can be identified by HTS from existing chemical libraries by exploiting a natural fluoride riboswitch. In addition, our findings suggest that some molecules might be further optimized to function as binary antibacterial agents when combined with fluoride. PMID:25910244

Evolution and Protein Packaging of Small Molecule RNA Aptamers

PubMed Central

Lau, Jolene L.; Baksh, Michael M.; Fiedler, Jason D.; Brown, Steven D.; Kussrow, Amanda; Bornhop, Darryl J.; Ordoukhanian, Phillip

2011-01-01

A high-affinity RNA aptamer (Kd = 50 nM) was efficiently identified by SELEX against a heteroaryl dihydropyrimidine structure, chosen as a representative drug-like molecule with no cross reactivity with mammalian or bacterial cells. This aptamer, its weaker-binding variants, and a known aptamer against theophylline were each embedded in a longer RNA sequence that was encapsidated inside a virus-like particle by a convenient expression technique. These nucleoprotein particles were shown by backscattering interferometry to bind to the small-molecule ligands with affinities similar to those of the free (non-encapsidated) aptamers. The system therefore comprises a general approach to the production and sequestration of functional RNA molecules, characterized by a convenient label-free analytical technique. PMID:21899290

High-throughput spectral and lifetime-based FRET screening in living cells to identify small-molecule effectors of SERCA

PubMed Central

Schaaf, Tory M.; Peterson, Kurt C.; Grant, Benjamin D.; Bawaskar, Prachi; Yuen, Samantha; Li, Ji; Muretta, Joseph M.; Gillispie, Gregory D.; Thomas, David D.

2017-01-01

A robust high-throughput screening (HTS) strategy has been developed to discover small-molecule effectors targeting the sarco/endoplasmic reticulum calcium ATPase (SERCA), based on a fluorescence microplate reader that records both the nanosecond decay waveform (lifetime mode) and the complete emission spectrum (spectral mode), with high precision and speed. This spectral unmixing plate reader (SUPR) was used to screen libraries of small molecules with a fluorescence resonance energy transfer (FRET) biosensor expressed in living cells. Ligand binding was detected by FRET associated with structural rearrangements of green (GFP, donor) and red (RFP, acceptor) fluorescent proteins fused to the cardiac-specific SERCA2a isoform. The results demonstrate accurate quantitation of FRET along with high precision of hit identification. Fluorescence lifetime analysis resolved SERCA’s distinct structural states, providing a method to classify small-molecule chemotypes on the basis of their structural effect on the target. The spectral analysis was also applied to flag interference by fluorescent compounds. FRET hits were further evaluated for functional effects on SERCA’s ATPase activity via both a coupled-enzyme assay and a FRET-based calcium sensor. Concentration-response curves indicated excellent correlation between FRET and function. These complementary spectral and lifetime FRET detection methods offer an attractive combination of precision, speed, and resolution for HTS. PMID:27899691

The methyltransferase SMYD3 mediates the recruitment of transcriptional cofactors at the myostatin and c-Met genes and regulates skeletal muscle atrophy

PubMed Central

Proserpio, Valentina; Fittipaldi, Raffaella; Ryall, James G.; Sartorelli, Vittorio; Caretti, Giuseppina

2013-01-01

Elucidating the epigenetic mechanisms underlying muscle mass determination and skeletal muscle wasting holds the potential of identifying molecular pathways that constitute possible drug targets. Here, we report that the methyltransferase SMYD3 modulates myostatin and c-Met transcription in primary skeletal muscle cells and C2C12 myogenic cells. SMYD3 targets the myostatin and c-Met genes and participates in the recruitment of the bromodomain protein BRD4 to their regulatory regions through protein–protein interaction. By recruiting BRD4, SMYD3 favors chromatin engagement of the pause–release factor p-TEFb (positive transcription elongation factor) and elongation of Ser2-phosphorylated RNA polymerase II (PolIISer2P). Reducing SMYD3 decreases myostatin and c-Met transcription, thus protecting from glucocorticoid-induced myotube atrophy. Supporting functional relevance of the SMYD3/BRD4 interaction, BRD4 pharmacological blockade by the small molecule JQ1 prevents dexamethasone-induced myostatin and atrogene up-regulation and spares myotube atrophy. Importantly, in a mouse model of dexamethasone-induced skeletal muscle atrophy, SMYD3 depletion prevents muscle loss and fiber size decrease. These findings reveal a mechanistic link between SMYD3/BRD4-dependent transcriptional regulation, muscle mass determination, and skeletal muscle atrophy and further encourage testing of small molecules targeting specific epigenetic regulators in animal models of muscle wasting. PMID:23752591

Discovery of a general method of solving the Schrödinger and dirac equations that opens a way to accurately predictive quantum chemistry.

PubMed

Nakatsuji, Hiroshi

2012-09-18

Just as Newtonian law governs classical physics, the Schrödinger equation (SE) and the relativistic Dirac equation (DE) rule the world of chemistry. So, if we can solve these equations accurately, we can use computation to predict chemistry precisely. However, for approximately 80 years after the discovery of these equations, chemists believed that they could not solve SE and DE for atoms and molecules that included many electrons. This Account reviews ideas developed over the past decade to further the goal of predictive quantum chemistry. Between 2000 and 2005, I discovered a general method of solving the SE and DE accurately. As a first inspiration, I formulated the structure of the exact wave function of the SE in a compact mathematical form. The explicit inclusion of the exact wave function's structure within the variational space allows for the calculation of the exact wave function as a solution of the variational method. Although this process sounds almost impossible, it is indeed possible, and I have published several formulations and applied them to solve the full configuration interaction (CI) with a very small number of variables. However, when I examined analytical solutions for atoms and molecules, the Hamiltonian integrals in their secular equations diverged. This singularity problem occurred in all atoms and molecules because it originates from the singularity of the Coulomb potential in their Hamiltonians. To overcome this problem, I first introduced the inverse SE and then the scaled SE. The latter simpler idea led to immediate and surprisingly accurate solution for the SEs of the hydrogen atom, helium atom, and hydrogen molecule. The free complement (FC) method, also called the free iterative CI (free ICI) method, was efficient for solving the SEs. In the FC method, the basis functions that span the exact wave function are produced by the Hamiltonian of the system and the zeroth-order wave function. These basis functions are called complement functions because they are the elements of the complete functions for the system under consideration. We extended this idea to solve the relativistic DE and applied it to the hydrogen and helium atoms, without observing any problems such as variational collapse. Thereafter, we obtained very accurate solutions of the SE for the ground and excited states of the Born-Oppenheimer (BO) and non-BO states of very small systems like He, H(2)(+), H(2), and their analogues. For larger systems, however, the overlap and Hamiltonian integrals over the complement functions are not always known mathematically (integration difficulty); therefore we formulated the local SE (LSE) method as an integral-free method. Without any integration, the LSE method gave fairly accurate energies and wave functions for small atoms and molecules. We also calculated continuous potential curves of the ground and excited states of small diatomic molecules by introducing the transferable local sampling method. Although the FC-LSE method is simple, the achievement of chemical accuracy in the absolute energy of larger systems remains time-consuming. The development of more efficient methods for the calculations of ordinary molecules would allow researchers to make these calculations more easily.

A Method for Extracting the Free Energy Surface and Conformational Dynamics of Fast-Folding Proteins from Single Molecule Photon Trajectories

PubMed Central

2015-01-01

Single molecule fluorescence spectroscopy holds the promise of providing direct measurements of protein folding free energy landscapes and conformational motions. However, fulfilling this promise has been prevented by technical limitations, most notably, the difficulty in analyzing the small packets of photons per millisecond that are typically recorded from individual biomolecules. Such limitation impairs the ability to accurately determine conformational distributions and resolve sub-millisecond processes. Here we develop an analytical procedure for extracting the conformational distribution and dynamics of fast-folding proteins directly from time-stamped photon arrival trajectories produced by single molecule FRET experiments. Our procedure combines the maximum likelihood analysis originally developed by Gopich and Szabo with a statistical mechanical model that describes protein folding as diffusion on a one-dimensional free energy surface. Using stochastic kinetic simulations, we thoroughly tested the performance of the method in identifying diverse fast-folding scenarios, ranging from two-state to one-state downhill folding, as a function of relevant experimental variables such as photon count rate, amount of input data, and background noise. The tests demonstrate that the analysis can accurately retrieve the original one-dimensional free energy surface and microsecond folding dynamics in spite of the sub-megahertz photon count rates and significant background noise levels of current single molecule fluorescence experiments. Therefore, our approach provides a powerful tool for the quantitative analysis of single molecule FRET experiments of fast protein folding that is also potentially extensible to the analysis of any other biomolecular process governed by sub-millisecond conformational dynamics. PMID:25988351

A Method for Extracting the Free Energy Surface and Conformational Dynamics of Fast-Folding Proteins from Single Molecule Photon Trajectories.

PubMed

Ramanathan, Ravishankar; Muñoz, Victor

2015-06-25

Single molecule fluorescence spectroscopy holds the promise of providing direct measurements of protein folding free energy landscapes and conformational motions. However, fulfilling this promise has been prevented by technical limitations, most notably, the difficulty in analyzing the small packets of photons per millisecond that are typically recorded from individual biomolecules. Such limitation impairs the ability to accurately determine conformational distributions and resolve sub-millisecond processes. Here we develop an analytical procedure for extracting the conformational distribution and dynamics of fast-folding proteins directly from time-stamped photon arrival trajectories produced by single molecule FRET experiments. Our procedure combines the maximum likelihood analysis originally developed by Gopich and Szabo with a statistical mechanical model that describes protein folding as diffusion on a one-dimensional free energy surface. Using stochastic kinetic simulations, we thoroughly tested the performance of the method in identifying diverse fast-folding scenarios, ranging from two-state to one-state downhill folding, as a function of relevant experimental variables such as photon count rate, amount of input data, and background noise. The tests demonstrate that the analysis can accurately retrieve the original one-dimensional free energy surface and microsecond folding dynamics in spite of the sub-megahertz photon count rates and significant background noise levels of current single molecule fluorescence experiments. Therefore, our approach provides a powerful tool for the quantitative analysis of single molecule FRET experiments of fast protein folding that is also potentially extensible to the analysis of any other biomolecular process governed by sub-millisecond conformational dynamics.

BiNChE: a web tool and library for chemical enrichment analysis based on the ChEBI ontology.

PubMed

Moreno, Pablo; Beisken, Stephan; Harsha, Bhavana; Muthukrishnan, Venkatesh; Tudose, Ilinca; Dekker, Adriano; Dornfeldt, Stefanie; Taruttis, Franziska; Grosse, Ivo; Hastings, Janna; Neumann, Steffen; Steinbeck, Christoph

2015-02-21

Ontology-based enrichment analysis aids in the interpretation and understanding of large-scale biological data. Ontologies are hierarchies of biologically relevant groupings. Using ontology annotations, which link ontology classes to biological entities, enrichment analysis methods assess whether there is a significant over or under representation of entities for ontology classes. While many tools exist that run enrichment analysis for protein sets annotated with the Gene Ontology, there are only a few that can be used for small molecules enrichment analysis. We describe BiNChE, an enrichment analysis tool for small molecules based on the ChEBI Ontology. BiNChE displays an interactive graph that can be exported as a high-resolution image or in network formats. The tool provides plain, weighted and fragment analysis based on either the ChEBI Role Ontology or the ChEBI Structural Ontology. BiNChE aids in the exploration of large sets of small molecules produced within Metabolomics or other Systems Biology research contexts. The open-source tool provides easy and highly interactive web access to enrichment analysis with the ChEBI ontology tool and is additionally available as a standalone library.

Developmental origins of a novel gut morphology in frogs.

PubMed

Bloom, Stephanie; Ledon-Rettig, Cris; Infante, Carlos; Everly, Anne; Hanken, James; Nascone-Yoder, Nanette

2013-05-01

Phenotypic variation is a prerequisite for evolution by natural selection, yet the processes that give rise to the novel morphologies upon which selection acts are poorly understood. We employed a chemical genetic screen to identify developmental changes capable of generating ecologically relevant morphological variation as observed among extant species. Specifically, we assayed for exogenously applied small molecules capable of transforming the ancestral larval foregut of the herbivorous Xenopus laevis to resemble the derived larval foregut of the carnivorous Lepidobatrachus laevis. Appropriately, the small molecules that demonstrate this capacity modulate conserved morphogenetic pathways involved in gut development, including downregulation of retinoic acid (RA) signaling. Identical manipulation of RA signaling in a species that is more closely related to Lepidobatrachus, Ceratophrys cranwelli, yielded even more similar transformations, corroborating the relevance of RA signaling variation in interspecific morphological change. Finally, we were able to recover the ancestral gut phenotype in Lepidobatrachus by performing a reverse chemical manipulation to upregulate RA signaling, providing strong evidence that modifications to this specific pathway promoted the emergence of a lineage-specific phenotypic novelty. Interestingly, our screen also revealed pathways that have not yet been implicated in early gut morphogenesis, such as thyroid hormone signaling. In general, the chemical genetic screen may be a valuable tool for identifying developmental mechanisms that underlie ecologically and evolutionarily relevant phenotypic variation. © 2013 Wiley Periodicals, Inc.

Growth of metal phthalocyanine on deactivated semiconducting surfaces steered by selective orbital coupling

DOE PAGES

Wagner, Sean R.; Feng, Jiagui; Yoon, Mina; ...

2015-08-25

Using scanning tunneling microscopy and density functional theory, we show that the molecular ordering and orientation of metal phthalocyanine molecules on the deactivated Si surface display a strong dependency on the central transition-metal ion, driven by the degree of orbital hybridization at the heterointerface via selective p – d orbital coupling. As a result, this Letter identifies a selective mechanism for modifying the molecule-substrate interaction which impacts the growth behavior of transition-metal-incorporated organic molecules on a technologically relevant substrate for silicon-based devices.

Monoclonal antibodies directed against surface molecules of multicell spheroids

NASA Technical Reports Server (NTRS)

Martinez, Andrew O.

1993-01-01

The objective of this project is to generate a library of monoclonal antibodies (MAb's) to surface molecules involved in the cell-cell interactions of mammalian cells grown as multicell spheroids (MCS). MCS are highly organized 3-dimensional multicellular structures which exhibit many characteristics in vivo tissues not found in conventional monolayer or suspension culture. They also provide a functional assay for surface adhesion molecules. In brief, MCS combine the relevance of organized tissues with the accuracy of in vitro methodology. Further, one can manipulate these MCS experimentally to discern important information about their biology.

E-Mail Molecules—Individualizing the Large Lecture Class

NASA Astrophysics Data System (ADS)

Wamser, Carl C.

2003-11-01

All students in the organic chemistry class are assigned a unique set of nine molecules to report on as optional extra credit assignments. The molecules are taken from a list containing over 200 molecules on the class Web site; they represent an assortment of biologically relevant compounds, from acetaminophen to yohimbine. Once a week, students may submit information about one of the molecules for two points extra credit (where the course includes a total of over 600 points from traditional quizzes and exams). The information requested about the molecules varies slightly each term as student expertise grows, for example, molecular formula, hybridizations, functional groups, or number of stereocenters, but always includes biological relevance and sources of information. Initially students submitted data directly to the instructor by e-mail, but submissions now are handled by a Web-based course management system (WebCT). The goal is to give students individualized assignments that are relatively realistic in light of their future careers in health sciences. Nearly all of the students do some of the molecules, and many students do all of them. About 30 40% of the students who do the assignments regularly gain a grade benefit. Student responses to the exercise have been positive.

Cholesterol - a biological compound as a building block in bionanotechnology

NASA Astrophysics Data System (ADS)

Hosta-Rigau, Leticia; Zhang, Yan; Teo, Boon M.; Postma, Almar; Städler, Brigitte

2012-12-01

Cholesterol is a molecule with many tasks in nature but also a long history in science. This feature article highlights the contribution of this small compound to bionanotechnology. We discuss relevant chemical aspects in this context followed by an overview of its self-assembly capabilities both as a free molecule and when conjugated to a polymer. Further, cholesterol in the context of liposomes is reviewed and its impact ranging from biosensing to drug delivery is outlined. Cholesterol is and will be an indispensable player in bionanotechnology, contributing to the progress of this potent field of research.

Thermoelectric effect and its dependence on molecular length and sequence in single DNA molecules.

PubMed

Li, Yueqi; Xiang, Limin; Palma, Julio L; Asai, Yoshihiro; Tao, Nongjian

2016-04-15

Studying the thermoelectric effect in DNA is important for unravelling charge transport mechanisms and for developing relevant applications of DNA molecules. Here we report a study of the thermoelectric effect in single DNA molecules. By varying the molecular length and sequence, we tune the charge transport in DNA to either a hopping- or tunnelling-dominated regimes. The thermoelectric effect is small and insensitive to the molecular length in the hopping regime. In contrast, the thermoelectric effect is large and sensitive to the length in the tunnelling regime. These findings indicate that one may control the thermoelectric effect in DNA by varying its sequence and length. We describe the experimental results in terms of hopping and tunnelling charge transport models.

Thermoelectric effect and its dependence on molecular length and sequence in single DNA molecules

PubMed Central

Li, Yueqi; Xiang, Limin; Palma, Julio L.; Asai, Yoshihiro; Tao, Nongjian

2016-01-01

Studying the thermoelectric effect in DNA is important for unravelling charge transport mechanisms and for developing relevant applications of DNA molecules. Here we report a study of the thermoelectric effect in single DNA molecules. By varying the molecular length and sequence, we tune the charge transport in DNA to either a hopping- or tunnelling-dominated regimes. The thermoelectric effect is small and insensitive to the molecular length in the hopping regime. In contrast, the thermoelectric effect is large and sensitive to the length in the tunnelling regime. These findings indicate that one may control the thermoelectric effect in DNA by varying its sequence and length. We describe the experimental results in terms of hopping and tunnelling charge transport models. PMID:27079152

The dynamics, structure, and conformational free energy of proline-containing antifreeze glycoprotein.

PubMed Central

Nguyen, Dat H; Colvin, Michael E; Yeh, Yin; Feeney, Robert E; Fink, William H

2002-01-01

Recent NMR studies of the solution structure of the 14-amino acid antifreeze glycoprotein AFGP-8 have concluded that the molecule lacks long-range order. The implication that an apparently unstructured molecule can still have a very precise function as a freezing inhibitor seems startling at first consideration. To gain insight into the nature of conformations and motions in AFGP-8, we have undertaken molecular dynamics simulations augmented with free energy calculations using a continuum solvation model. Starting from 10 different NMR structures, 20 ns of dynamics of AFGP were explored. The dynamics show that AFGP structure is composed of four segments, joined by very flexible pivots positioned at alanine 5, 8, and 11. The dynamics also show that the presence of prolines in this small AFGP structure facilitates the adoption of the poly-proline II structure as its overall conformation, although AFGP does adopt other conformations during the course of dynamics as well. The free energies calculated using a continuum solvation model show that the lowest free energy conformations, while being energetically equal, are drastically different in conformations. In other words, this AFGP molecule has many structurally distinct and energetically equal minima in its energy landscape. In addition, conformational, energetic, and hydrogen bond analyses suggest that the intramolecular hydrogen bonds between the N-acetyl group and the protein backbone are an important integral part of the overall stability of the AFGP molecule. The relevance of these findings to the mechanism of freezing inhibition is discussed. PMID:12023212

The dynamics, structure, and conformational free energy of proline-containing antifreeze glycoprotein.

PubMed

Nguyen, Dat H; Colvin, Michael E; Yeh, Yin; Feeney, Robert E; Fink, William H

2002-06-01

Recent NMR studies of the solution structure of the 14-amino acid antifreeze glycoprotein AFGP-8 have concluded that the molecule lacks long-range order. The implication that an apparently unstructured molecule can still have a very precise function as a freezing inhibitor seems startling at first consideration. To gain insight into the nature of conformations and motions in AFGP-8, we have undertaken molecular dynamics simulations augmented with free energy calculations using a continuum solvation model. Starting from 10 different NMR structures, 20 ns of dynamics of AFGP were explored. The dynamics show that AFGP structure is composed of four segments, joined by very flexible pivots positioned at alanine 5, 8, and 11. The dynamics also show that the presence of prolines in this small AFGP structure facilitates the adoption of the poly-proline II structure as its overall conformation, although AFGP does adopt other conformations during the course of dynamics as well. The free energies calculated using a continuum solvation model show that the lowest free energy conformations, while being energetically equal, are drastically different in conformations. In other words, this AFGP molecule has many structurally distinct and energetically equal minima in its energy landscape. In addition, conformational, energetic, and hydrogen bond analyses suggest that the intramolecular hydrogen bonds between the N-acetyl group and the protein backbone are an important integral part of the overall stability of the AFGP molecule. The relevance of these findings to the mechanism of freezing inhibition is discussed.

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

Nakatsuji, Hiroshi, E-mail: h.nakatsuji@qcri.or.jp; Nakashima, Hiroyuki

The Schrödinger equation (SE) and the antisymmetry principle constitute the governing principle of chemistry. A general method of solving the SE was presented before as the free complement (FC) theory, which gave highly accurate solutions for small atoms and molecules. We assume here to use the FC theory starting from the local valence bond wave function. When this theory is applied to larger molecules, antisymmetrizations of electronic wave functions become time-consuming and therefore, an additional breakthrough is necessary concerning the antisymmetry principle. Usually, in molecular calculations, we first construct the wave function to satisfy the antisymmetry rule, “electronic wave functionsmore » must be prescribed to be antisymmetric for all exchanges of electrons, otherwise bosonic interference may disturb the basis of the science.” Starting from determinantal wave functions is typical. Here, we give an antisymmetrization theory, called inter-exchange (iExg) theory, by dividing molecular antisymmetrizations to those within atoms and between atoms. For the electrons belonging to distant atoms in a molecule, only partial antisymmetrizations or even no antisymmetrizations are necessary, depending on the distance between the atoms. So, the above antisymmetry rule is not necessarily followed strictly to get the results of a desired accuracy. For this and other reasons, the necessary parts of the antisymmetrization operations become very small as molecules become larger, leading finally to the operation counts of lower orders of N, the number of electrons. This theory creates a natural antisymmetrization method that is useful for large molecules.« less

A Pilot Study of Ion - Molecule Reactions at Temperatures Relevant to the Atmosphere of Titan.

PubMed

Zymak, Illia; Žabka, Ján; Polášek, Miroslav; Španěl, Patrik; Smith, David

2016-11-01

Reliable theoretical models of the chemical kinetics of the ionosphere of Saturn's moon, Titan, is highly dependent on the precision of the rates of the reactions of ambient ions with hydrocarbon molecules at relevant temperatures. A Variable Temperature Selected Ions Flow Tube technique, which has been developed primarily to study these reactions at temperatures within the range of 200-330 K, is briefly described. The flow tube temperature regulation system and the thermalisation of ions are also discussed. Preliminary studies of two reactions have been carried out to check the reliability and efficacy of kinetics measurements: (i) Rate constants of the reaction of CH 3 + ions with molecular oxygen were measured at different temperatures, which indicate values in agreement with previous ion cyclotron resonance measurements ostensibly made at 300 K. (ii) Formation of CH 3 + ions in the reaction of N 2 + ions with CH 4 molecules were studied at temperatures within the range 240-310 K which showed a small but statistically significant decrease of the ratio of product CH 3 + ions to reactant N 2 + ions with reaction temperature.

Star PolyMOCs with Diverse Structures, Dynamics, and Functions by Three-Component Assembly

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

Wang, Yufeng; Gu, Yuwei; Keeler, Eric G.

2016-12-05

We report star polymer metal–organic cage (polyMOC) materials whose structures, mechanical properties, functionalities, and dynamics can all be precisely tailored through a simple three-component assembly strategy. The star polyMOC network is composed of tetra-arm star polymers functionalized with ligands on the chain ends, small molecule ligands, and palladium ions; polyMOCs are formed via metal–ligand coordination and thermal annealing. The ratio of small molecule ligands to polymer-bound ligands determines the connectivity of the MOC junctions and the network structure. The use of large M12L24 MOCs enables great flexibility in tuning this ratio, which provides access to a rich spectrum of materialmore » properties including tunable moduli and relaxation dynamics.« less

Nanoparticle-based delivery of small interfering RNA: challenges for cancer therapy

PubMed Central

Miele, Evelina; Spinelli, Gian Paolo; Miele, Ermanno; Di Fabrizio, Enzo; Ferretti, Elisabetta; Tomao, Silverio; Gulino, Alberto

2012-01-01

During recent decades there have been remarkable advances and profound changes in cancer therapy. Many therapeutic strategies learned at the bench, including monoclonal antibodies and small molecule inhibitors, have been used at the bedside, leading to important successes. One of the most important advances in biology has been the discovery that small interfering RNA (siRNA) is able to regulate the expression of genes, by a phenomenon known as RNA interference (RNAi). RNAi is one of the most rapidly growing fields of research in biology and therapeutics. Much research effort has gone into the application of this new discovery in the treatment of various diseases, including cancer. However, even though these molecules may have potential and strong utility, some limitations make their clinical application difficult, including delivery problems, side effects due to off-target actions, disturbance of physiological functions of the cellular machinery involved in gene silencing, and induction of the innate immune response. Many researchers have attempted to overcome these limitations and to improve the safety of potential RNAi-based therapeutics. Nanoparticles, which are nanostructured entities with tunable size, shape, and surface, as well as biological behavior, provide an ideal opportunity to modify current treatment regimens in a substantial way. These nanoparticles could be designed to surmount one or more of the barriers encountered by siRNA. Nanoparticle drug formulations afford the chance to improve drug bioavailability, exploiting superior tissue permeability, payload protection, and the “stealth” features of these entities. The main aims of this review are: to explain the siRNA mechanism with regard to potential applications in siRNA-based cancer therapy; to discuss the possible usefulness of nanoparticle-based delivery of certain molecules for overcoming present therapeutic limitations; to review the ongoing relevant clinical research with its pitfalls and promises; and to evaluate critically future perspectives and challenges in siRNA-based cancer therapy. PMID:22915840

FlexAID: Revisiting Docking on Non-Native-Complex Structures.

PubMed

Gaudreault, Francis; Najmanovich, Rafael J

2015-07-27

Small-molecule protein docking is an essential tool in drug design and to understand molecular recognition. In the present work we introduce FlexAID, a small-molecule docking algorithm that accounts for target side-chain flexibility and utilizes a soft scoring function, i.e. one that is not highly dependent on specific geometric criteria, based on surface complementarity. The pairwise energy parameters were derived from a large dataset of true positive poses and negative decoys from the PDBbind database through an iterative process using Monte Carlo simulations. The prediction of binding poses is tested using the widely used Astex dataset as well as the HAP2 dataset, while performance in virtual screening is evaluated using a subset of the DUD dataset. We compare FlexAID to AutoDock Vina, FlexX, and rDock in an extensive number of scenarios to understand the strengths and limitations of the different programs as well as to reported results for Glide, GOLD, and DOCK6 where applicable. The most relevant among these scenarios is that of docking on flexible non-native-complex structures where as is the case in reality, the target conformation in the bound form is not known a priori. We demonstrate that FlexAID, unlike other programs, is robust against increasing structural variability. FlexAID obtains equivalent sampling success as GOLD and performs better than AutoDock Vina or FlexX in all scenarios against non-native-complex structures. FlexAID is better than rDock when there is at least one critical side-chain movement required upon ligand binding. In virtual screening, FlexAID results are lower on average than those of AutoDock Vina and rDock. The higher accuracy in flexible targets where critical movements are required, intuitive PyMOL-integrated graphical user interface and free source code as well as precompiled executables for Windows, Linux, and Mac OS make FlexAID a welcome addition to the arsenal of existing small-molecule protein docking methods.

Bruton's Tyrosine Kinase Small Molecule Inhibitors Induce a Distinct Pancreatic Toxicity in Rats.

PubMed

Erickson, Rebecca I; Schutt, Leah K; Tarrant, Jacqueline M; McDowell, Michelle; Liu, Lichuan; Johnson, Adam R; Lewin-Koh, Sock-Cheng; Hedehus, Maj; Ross, Jed; Carano, Richard A D; Staflin, Karin; Zhong, Fiona; Crawford, James J; Zhong, Shelly; Reif, Karin; Katewa, Arna; Wong, Harvey; Young, Wendy B; Dambach, Donna M; Misner, Dinah L

2017-01-01

Bruton's tyrosine kinase (BTK) is a member of the Tec family of cytoplasmic tyrosine kinases involved in B-cell and myeloid cell signaling. Small molecule inhibitors of BTK are being investigated for treatment of several hematologic cancers and autoimmune diseases. GDC-0853 ((S)-2-(3'-(hydroxymethyl)-1-methyl-5-((5-(2-methyl-4-(oxetan-3-yl)piperazin-1-yl)pyridin-2-yl)amino)-6-oxo-1,6-dihydro-[3,4'-bipyridin]-2'-yl)-7,7-dimethyl-3,4,7,8-tetrahydro-2H-cyclopenta[4,5]pyrrolo[1,2-a]pyrazin-1(6H)-one) is a selective and reversible oral small-molecule BTK inhibitor in development for the treatment of rheumatoid arthritis and systemic lupus erythematosus. In Sprague-Dawley (SD) rats, administration of GDC-0853 and other structurally diverse BTK inhibitors for 7 days or longer caused pancreatic lesions consisting of multifocal islet-centered hemorrhage, inflammation, fibrosis, and pigment-laden macrophages with adjacent lobular exocrine acinar cell atrophy, degeneration, and inflammation. Similar findings were not observed in mice or dogs at much higher exposures. Hemorrhage in the peri-islet vasculature emerged between four and seven daily doses of GDC-0853 and was histologically similar to spontaneously occurring changes in aging SD rats. This suggests that GDC-0853 could exacerbate a background finding in younger animals. Glucose homeostasis was dysregulated following a glucose challenge; however, this occurred only after 28 days of administration and was not directly associated with onset or severity of pancreatic lesions. There were no changes in other common serum biomarkers assessing endocrine and exocrine pancreatic function. Additionally, these lesions were not readily detectable via Doppler ultrasound, computed tomography, or magnetic resonance imaging. Our results indicate that pancreatic lesions in rats are likely a class effect of BTK inhibitors, which may exacerbate an islet-centered pathology that is unlikely to be relevant to humans. Copyright © 2016 by The American Society for Pharmacology and Experimental Therapeutics.

Mitochondrial Targets for Pharmacological Intervention in Human Disease

PubMed Central

2015-01-01

Over the past several years, mitochondrial dysfunction has been linked to an increasing number of human illnesses, making mitochondrial proteins (MPs) an ever more appealing target for therapeutic intervention. With 20% of the mitochondrial proteome (312 of an estimated 1500 MPs) having known interactions with small molecules, MPs appear to be highly targetable. Yet, despite these targeted proteins functioning in a range of biological processes (including induction of apoptosis, calcium homeostasis, and metabolism), very few of the compounds targeting MPs find clinical use. Recent work has greatly expanded the number of proteins known to localize to the mitochondria and has generated a considerable increase in MP 3D structures available in public databases, allowing experimental screening and in silico prediction of mitochondrial drug targets on an unprecedented scale. Here, we summarize the current literature on clinically active drugs that target MPs, with a focus on how existing drug targets are distributed across biochemical pathways and organelle substructures. Also, we examine current strategies for mitochondrial drug discovery, focusing on genetic, proteomic, and chemogenomic assays, and relevant model systems. As cell models and screening techniques improve, MPs appear poised to emerge as relevant targets for a wide range of complex human diseases, an eventuality that can be expedited through systematic analysis of MP function. PMID:25367773

Low-temperature surface formation of NH3 and HNCO: hydrogenation of nitrogen atoms in CO-rich interstellar ice analogues

NASA Astrophysics Data System (ADS)

Fedoseev, G.; Ioppolo, S.; Zhao, D.; Lamberts, T.; Linnartz, H.

2015-01-01

Solid-state astrochemical reaction pathways have the potential to link the formation of small nitrogen-bearing species, like NH3 and HNCO, and prebiotic molecules, specifically amino acids. To date, the chemical origin of such small nitrogen-containing species is still not well understood, despite the fact that ammonia is an abundant constituent of interstellar ices towards young stellar objects and quiescent molecular clouds. This is mainly because of the lack of dedicated laboratory studies. The aim of this work is to experimentally investigate the formation routes of NH3 and HNCO through non-energetic surface reactions in interstellar ice analogues under fully controlled laboratory conditions and at astrochemically relevant temperatures. This study focuses on the formation of NH3 and HNCO in CO-rich (non-polar) interstellar ices that simulate the CO freeze-out stage in dark interstellar cloud regions, well before thermal and energetic processing start to become relevant. We demonstrate and discuss the surface formation of solid HNCO through the interaction of CO molecules with NH radicals - one of the intermediates in the formation of solid NH3 upon sequential hydrogenation of N atoms. The importance of HNCO for astrobiology is discussed.

Integrating computational and chemical biology tools in the discovery of antiangiogenic small molecule ligands of FGF2 derived from endogenous inhibitors

PubMed Central

Foglieni, Chiara; Pagano, Katiuscia; Lessi, Marco; Bugatti, Antonella; Moroni, Elisabetta; Pinessi, Denise; Resovi, Andrea; Ribatti, Domenico; Bertini, Sabrina; Ragona, Laura; Bellina, Fabio; Rusnati, Marco; Colombo, Giorgio; Taraboletti, Giulia

2016-01-01

The FGFs/FGFRs system is a recognized actionable target for therapeutic approaches aimed at inhibiting tumor growth, angiogenesis, metastasis, and resistance to therapy. We previously identified a non-peptidic compound (SM27) that retains the structural and functional properties of the FGF2-binding sequence of thrombospondin-1 (TSP-1), a major endogenous inhibitor of angiogenesis. Here we identified new small molecule inhibitors of FGF2 based on the initial lead. A similarity-based screening of small molecule libraries, followed by docking calculations and experimental studies, allowed selecting 7 bi-naphthalenic compounds that bound FGF2 inhibiting its binding to both heparan sulfate proteoglycans and FGFR-1. The compounds inhibit FGF2 activity in in vitro and ex vivo models of angiogenesis, with improved potency over SM27. Comparative analysis of the selected hits, complemented by NMR and biochemical analysis of 4 newly synthesized functionalized phenylamino-substituted naphthalenes, allowed identifying the minimal stereochemical requirements to improve the design of naphthalene sulfonates as FGF2 inhibitors. PMID:27000667

Heterogeneous Biomedical Database Integration Using a Hybrid Strategy: A p53 Cantcer Research Database

PubMed Central

Bichutskiy, Vadim Y.; Colman, Richard; Brachmann, Rainer K.; Lathrop, Richard H.

2006-01-01

Complex problems in life science research give rise to multidisciplinary collaboration, and hence, to the need for heterogeneous database integration. The tumor suppressor p53 is mutated in close to 50% of human cancers, and a small drug-like molecule with the ability to restore native function to cancerous p53 mutants is a long-held medical goal of cancer treatment. The Cancer Research DataBase (CRDB) was designed in support of a project to find such small molecules. As a cancer informatics project, the CRDB involved small molecule data, computational docking results, functional assays, and protein structure data. As an example of the hybrid strategy for data integration, it combined the mediation and data warehousing approaches. This paper uses the CRDB to illustrate the hybrid strategy as a viable approach to heterogeneous data integration in biomedicine, and provides a design method for those considering similar systems. More efficient data sharing implies increased productivity, and, hopefully, improved chances of success in cancer research. (Code and database schemas are freely downloadable, http://www.igb.uci.edu/research/research.html.) PMID:19458771

miRNAs and ovarian cancer: An overview.

PubMed

Deb, Bornali; Uddin, Arif; Chakraborty, Supriyo

2018-05-01

Ovarian cancer (OC) is the sixth most common cancer in women globally. However, even with the advances in detection and therapeutics it still represents the most dangerous gynecologic malignancy in women of the industrialized countries. The discovery of micro-RNAs (miRNA), a small noncoding RNA molecule targeting multiple mRNAs and regulation of gene expression by triggering translation repression and/or RNA degradation, has revealed the existence of a new array for regulation of genes involved in cancer. This review summarizes the current knowledge regarding the role of miRNAs expression in OC. It also provides information about potential clinical relevance of circulating miRNAs for OC diagnosis, prognosis, and therapeutics. The identification of functional targets for miRNAs represents a major obstacle in our understanding of microRNA function in OC, but significant progress is being made. The better understanding of the role of microRNA expression in ovarian cancer may provide new array for the detection, diagnosis, and therapy of the OC. © 2017 Wiley Periodicals, Inc.

Aquaporins in Clinical Medicine

PubMed Central

Verkman, A.S.

2012-01-01

The aquaporins are a family of membrane water channels, some of which also transport glycerol. They are involved in a wide range of physiological functions (including water/salt homeostasis, exocrine fluid secretion, and epidermal hydration) and human diseases (including glaucoma, cancer, epilepsy, and obesity). At the cellular level, aquaporin-mediated osmotic water transport across cell plasma membranes facilitates transepithelial fluid transport, cell migration, and neuroexcitation; aquaporin-mediated glycerol transport regulates cell proliferation, adipocyte metabolism, and epidermal water retention. Genetic diseases caused by loss-of-function mutations in aquaporins include nephrogenic diabetes insipidus and congenital cataracts. The neuroinflammatory demyelinating disease neuromyelitis optica is marked by pathogenic autoantibodies against astrocyte water channel aquaporin-4. There remain broad opportunities for the development of aquaporin-based diagnostics and therapeutics. Disease-relevant aquaporin polymorphisms are beginning to be explored. There is great promise in the development of small-molecule aquaporin modulators for therapy of some types of refractory edema, brain swelling, neuroinflammation, glaucoma, epilepsy, cancer, pain, and obesity. PMID:22248325

Homogeneous time-resolved G protein-coupled receptor-ligand binding assay based on fluorescence cross-correlation spectroscopy.

PubMed

Antoine, Thomas; Ott, David; Ebell, Katharina; Hansen, Kerrin; Henry, Luc; Becker, Frank; Hannus, Stefan

2016-06-01

G protein-coupled receptors (GPCRs) mediate many important physiological functions and are considered as one of the most successful therapeutic target classes for a wide spectrum of diseases. Drug discovery projects generally benefit from a broad range of experimental approaches for screening compound libraries and for the characterization of binding modes of drug candidates. Owing to the difficulties in solubilizing and purifying GPCRs, assay formats have been so far mainly limited to cell-based functional assays and radioligand binding assays. In this study, we used fluorescence cross-correlation spectroscopy (FCCS) to analyze the interaction of detergent-solubilized receptors to various types of GPCR ligands: endogenous peptides, small molecules, and a large surrogate antagonist represented by a blocking monoclonal antibody. Our work demonstrates the suitability of the homogeneous and time-resolved FCCS assay format for a robust, high-throughput determination of receptor-ligand binding affinities and kinetic rate constants for various therapeutically relevant GPCRs. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Facilitated transport of small molecules and ions for energy-efficient membranes.

PubMed

Li, Yifan; Wang, Shaofei; He, Guangwei; Wu, Hong; Pan, Fusheng; Jiang, Zhongyi

2015-01-07

In nature, the biological membrane can selectively transport essential small molecules/ions through facilitated diffusion via carrier proteins. Intrigued by this phenomenon and principle, membrane researchers have successfully employed synthetic carriers and carrier-mediated reversible reactions to enhance the separation performance of synthetic membranes. However, the existing facilitated transport membranes as well as the relevant facilitated transport theories have scarcely been comprehensively reviewed in the literature. This tutorial review primarily covers the two aspects of facilitated transport theories: carrier-mediated transport mechanisms and facilitated transport chemistries, including the design and fabrication of facilitated transport membranes. The applications of facilitated transport membranes in energy-intensive membrane processes (gas separation, pervaporation, and proton exchange membrane fuel cells) have also been discussed. Hopefully, this review will provide guidelines for the future research and development of facilitated transport membranes with high energy efficiency.

Large-Scale Validation of Mixed-Solvent Simulations to Assess Hotspots at Protein-Protein Interaction Interfaces.

PubMed

Ghanakota, Phani; van Vlijmen, Herman; Sherman, Woody; Beuming, Thijs

2018-04-23

The ability to target protein-protein interactions (PPIs) with small molecule inhibitors offers great promise in expanding the druggable target space and addressing a broad range of untreated diseases. However, due to their nature and function of interacting with protein partners, PPI interfaces tend to extend over large surfaces without the typical pockets of enzymes and receptors. These features present unique challenges for small molecule inhibitor design. As such, determining whether a particular PPI of interest could be pursued with a small molecule discovery strategy requires an understanding of the characteristics of the PPI interface and whether it has hotspots that can be leveraged by small molecules to achieve desired potency. Here, we assess the ability of mixed-solvent molecular dynamic (MSMD) simulations to detect hotspots at PPI interfaces. MSMD simulations using three cosolvents (acetonitrile, isopropanol, and pyrimidine) were performed on a large test set of 21 PPI targets that have been experimentally validated by small molecule inhibitors. We compare MSMD, which includes explicit solvent and full protein flexibility, to a simpler approach that does not include dynamics or explicit solvent (SiteMap) and find that MSMD simulations reveal additional information about the characteristics of these targets and the ability for small molecules to inhibit the PPI interface. In the few cases were MSMD simulations did not detect hotspots, we explore the shortcomings of this technique and propose future improvements. Finally, using Interleukin-2 as an example, we highlight the advantage of the MSMD approach for detecting transient cryptic druggable pockets that exists at PPI interfaces.

Discovery and development of 5-[(5S,9R)-9-(4-cyanophenyl)-3-(3,5-dichlorophenyl)-1-methyl-2,4-dioxo-1,3,7-triazaspiro[4.4]non-7-yl-methyl]-3-thiophenecarboxylic acid (BMS-587101)--a small molecule antagonist of leukocyte function associated antigen-1.

PubMed

Potin, Dominique; Launay, Michele; Monatlik, Francoise; Malabre, Patrice; Fabreguettes, Maud; Fouquet, Andre; Maillet, Magali; Nicolai, Eric; Dorgeret, Loïc; Chevallier, François; Besse, Dominique; Dufort, Monique; Caussade, François; Ahmad, Syed Z; Stetsko, Dawn K; Skala, Stacey; Davis, Patricia M; Balimane, Praveen; Patel, Karishma; Yang, Zheng; Marathe, Punit; Postelneck, Jennifer; Townsend, Robert M; Goldfarb, Valentina; Sheriff, Steven; Einspahr, Howard; Kish, Kevin; Malley, Mary F; DiMarco, John D; Gougoutas, Jack Z; Kadiyala, Pathanjali; Cheney, Daniel L; Tejwani, Ravindra W; Murphy, Denette K; Mcintyre, Kim W; Yang, Xiaoxia; Chao, Sam; Leith, Leslie; Xiao, Zili; Mathur, Arvind; Chen, Bang-Chi; Wu, Daugh-Rurng; Traeger, Sarah C; McKinnon, Murray; Barrish, Joel C; Robl, Jeffrey A; Iwanowicz, Edwin J; Suchard, Suzanne J; Dhar, T G Murali

2006-11-30

LFA-1 (leukocyte function-associated antigen-1), is a member of the beta2-integrin family and is expressed on all leukocytes. This letter describes the discovery and preliminary SAR of spirocyclic hydantoin based LFA-1 antagonists that culminated in the identification of analog 8 as a clinical candidate. We also report the first example of the efficacy of a small molecule LFA-1 antagonist in combination with CTLA-4Ig in an animal model of transplant rejection.

The Role of Histone Deacetylases in Neurodegenerative Diseases and Small-Molecule Inhibitors as a Potential Therapeutic Approach

NASA Astrophysics Data System (ADS)

Bürli, Roland W.; Thomas, Elizabeth; Beaumont, Vahri

Neurodegenerative disorders are devastating for patients and their social environment. Their etiology is poorly understood and complex. As a result, there is clearly an urgent need for therapeutic agents that slow down disease progress and alleviate symptoms. In this respect, interference with expression and function of multiple gene products at the epigenetic level has offered much promise, and histone deacetylases play a crucial role in these processes. This review presents an overview of the biological pathways in which these enzymes are involved and illustrates the complex network of proteins that governs their activity. An overview of small molecules that interfere with histone deacetylase function is provided.

Differential targeting of Gbetagamma-subunit signaling with small molecules.

PubMed

Bonacci, Tabetha M; Mathews, Jennifer L; Yuan, Chujun; Lehmann, David M; Malik, Sundeep; Wu, Dianqing; Font, Jose L; Bidlack, Jean M; Smrcka, Alan V

2006-04-21

G protein betagamma subunits have potential as a target for therapeutic treatment of a number of diseases. We performed virtual docking of a small-molecule library to a site on Gbetagamma subunits that mediates protein interactions. We hypothesized that differential targeting of this surface could allow for selective modulation of Gbetagamma subunit functions. Several compounds bound to Gbetagamma subunits with affinities from 0.1 to 60 muM and selectively modulated functional Gbetagamma-protein-protein interactions in vitro, chemotactic peptide signaling pathways in HL-60 leukocytes, and opioid receptor-dependent analgesia in vivo. These data demonstrate an approach for modulation of G protein-coupled receptor signaling that may represent an important therapeutic strategy.

Osteoclast-targeting small molecules for the treatment of neoplastic bone metastases.

PubMed

Kawatani, Makoto; Osada, Hiroyuki

2009-11-01

Osteoclasts are highly specialized cells that resorb bone, and their abnormal activity is implicated in a variety of human bone diseases. In neoplastic bone metastasis, the bone destruction caused by osteoclasts is not only associated with the formation and progression of metastatic lesions, but also could contribute to frequent complications such as severe pain and pathological fractures, which greatly diminish the quality of life of patients. Bisphosphonates, potent antiresorptive drugs, have been shown to have efficacy for treating bone metastases in many types of cancer, and the development of various molecularly targeted agents is currently proceeding. Thus, inhibition of osteoclast function is now established as an important treatment strategy for bony metastases. This review focuses on promising small molecules that disrupt osteoclast function and introduces our chemical/biological approach for identifying osteoclast-targeting small molecular inhibitors.

RNA interference for performance enhancement and detection in doping control.

PubMed

Kohler, Maxie; Schänzer, Wilhelm; Thevis, Mario

2011-10-01

RNA interference represents a comparably new route of regulating and manipulating specific gene expression. Promising results were obtained in experimental therapies aim at the treatment of different kinds of diseases including cancer, diabetes mellitus or Dychenne muscular dystrophy. While studies on down-regulation efficiency are often performed by analyzing the regulated protein, the direct detection of small, interfering RNA molecules and antisense oligonucleotides is of great interest for the investigation of the metabolism and degradation and also for the detection of a putative misuse of these molecules in sports. Myostatin down-regulation was shown to result in increased performance and muscle growth and the regulation of several other proteins could be relevant for performance enhancement. This mini-review summarizes current approaches for the mass spectrometric analysis of siRNA and antisense oligonucleotides from biological matrices and the available data on biodistribution, metabolism, and half-life of relevant substances are discussed. Copyright © 2011 John Wiley & Sons, Ltd.

Drug-loaded nanoparticles induce gene expression in human pluripotent stem cell derivatives

NASA Astrophysics Data System (ADS)

Gajbhiye, Virendra; Escalante, Leah; Chen, Guojun; Laperle, Alex; Zheng, Qifeng; Steyer, Benjamin; Gong, Shaoqin; Saha, Krishanu

2013-12-01

Tissue engineering and advanced manufacturing of human stem cells requires a suite of tools to control gene expression spatiotemporally in culture. Inducible gene expression systems offer cell-extrinsic control, typically through addition of small molecules, but small molecule inducers typically contain few functional groups for further chemical modification. Doxycycline (DXC), a potent small molecule inducer of tetracycline (Tet) transgene systems, was conjugated to a hyperbranched dendritic polymer (Boltorn H40) and subsequently reacted with polyethylene glycol (PEG). The resulting PEG-H40-DXC nanoparticle exhibited pH-sensitive drug release behavior and successfully controlled gene expression in stem-cell-derived fibroblasts with a Tet-On system. While free DXC inhibited fibroblast proliferation and matrix metalloproteinase (MMP) activity, PEG-H40-DXC nanoparticles maintained higher fibroblast proliferation levels and MMP activity. The results demonstrate that the PEG-H40-DXC nanoparticle system provides an effective tool to controlling gene expression in human stem cell derivatives.Tissue engineering and advanced manufacturing of human stem cells requires a suite of tools to control gene expression spatiotemporally in culture. Inducible gene expression systems offer cell-extrinsic control, typically through addition of small molecules, but small molecule inducers typically contain few functional groups for further chemical modification. Doxycycline (DXC), a potent small molecule inducer of tetracycline (Tet) transgene systems, was conjugated to a hyperbranched dendritic polymer (Boltorn H40) and subsequently reacted with polyethylene glycol (PEG). The resulting PEG-H40-DXC nanoparticle exhibited pH-sensitive drug release behavior and successfully controlled gene expression in stem-cell-derived fibroblasts with a Tet-On system. While free DXC inhibited fibroblast proliferation and matrix metalloproteinase (MMP) activity, PEG-H40-DXC nanoparticles maintained higher fibroblast proliferation levels and MMP activity. The results demonstrate that the PEG-H40-DXC nanoparticle system provides an effective tool to controlling gene expression in human stem cell derivatives. Electronic supplementary information (ESI) available: ESI containing 1H NMR spectra and additional fibroblast characterization data. See DOI: 10.1039/c3nr04794f

Molecular locks and keys: the role of small molecules in phytohormone research

PubMed Central

Fonseca, Sandra; Rosado, Abel; Vaughan-Hirsch, John; Bishopp, Anthony; Chini, Andrea

2014-01-01

Plant adaptation, growth and development rely on the integration of many environmental and endogenous signals that collectively determine the overall plant phenotypic plasticity. Plant signaling molecules, also known as phytohormones, are fundamental to this process. These molecules act at low concentrations and regulate multiple aspects of plant fitness and development via complex signaling networks. By its nature, phytohormone research lies at the interface between chemistry and biology. Classically, the scientific community has always used synthetic phytohormones and analogs to study hormone functions and responses. However, recent advances in synthetic and combinational chemistry, have allowed a new field, plant chemical biology, to emerge and this has provided a powerful tool with which to study phytohormone function. Plant chemical biology is helping to address some of the most enduring questions in phytohormone research such as: Are there still undiscovered plant hormones? How can we identify novel signaling molecules? How can plants activate specific hormone responses in a tissue-specific manner? How can we modulate hormone responses in one developmental context without inducing detrimental effects on other processes? The chemical genomics approaches rely on the identification of small molecules modulating different biological processes and have recently identified active forms of plant hormones and molecules regulating many aspects of hormone synthesis, transport and response. We envision that the field of chemical genomics will continue to provide novel molecules able to elucidate specific aspects of hormone-mediated mechanisms. In addition, compounds blocking specific responses could uncover how complex biological responses are regulated. As we gain information about such compounds we can design small alterations to the chemical structure to further alter specificity, enhance affinity or modulate the activity of these compounds. PMID:25566283

Septins - active GTPases or just GTP-binding proteins?

PubMed

Abbey, Megha; Gaestel, Matthias; Menon, Manoj B

2018-05-10

Septins are conserved cytoskeletal proteins with unique filament forming capabilities and roles in cytokinesis and cell morphogenesis. Septins undergo hetero-oligomerization and assemble into higher order structures including filaments, rings and cages. Hetero- and homotypic interactions of septin isoforms involve alternating GTPase (G)-domain interfaces and those mediated by N- and C-terminal extensions. While most septins bind GTP, display weak GTP-hydrolysis activity and incorporate guanine nucleotides in their interaction interfaces, studies using GTPase-inactivating mutations have failed to conclusively establish a crucial role for GTPase activity in mediating septin functions. In this mini-review, we will critically assess the role of GTP-binding and -hydrolysis on septin assembly and function. The relevance of G-domain activity will also be discussed in the context of human septin mutations as well as the development of specific small-molecules targeting septin polymerization. As structural determinants of septin oligomer interfaces, G-domains are attractive targets for ligand-based inhibition of septin assembly. Whether such an intervention can predictably alter septin function is a major question for future research. This article is protected by copyright. All rights reserved. © 2018 Wiley Periodicals, Inc.

High-throughput screening based on label-free detection of small molecule microarrays

NASA Astrophysics Data System (ADS)

Zhu, Chenggang; Fei, Yiyan; Zhu, Xiangdong

2017-02-01

Based on small-molecule microarrays (SMMs) and oblique-incidence reflectivity difference (OI-RD) scanner, we have developed a novel high-throughput drug preliminary screening platform based on label-free monitoring of direct interactions between target proteins and immobilized small molecules. The screening platform is especially attractive for screening compounds against targets of unknown function and/or structure that are not compatible with functional assay development. In this screening platform, OI-RD scanner serves as a label-free detection instrument which is able to monitor about 15,000 biomolecular interactions in a single experiment without the need to label any biomolecule. Besides, SMMs serves as a novel format for high-throughput screening by immobilization of tens of thousands of different compounds on a single phenyl-isocyanate functionalized glass slide. Based on the high-throughput screening platform, we sequentially screened five target proteins (purified target proteins or cell lysate containing target protein) in high-throughput and label-free mode. We found hits for respective target protein and the inhibition effects for some hits were confirmed by following functional assays. Compared to traditional high-throughput screening assay, the novel high-throughput screening platform has many advantages, including minimal sample consumption, minimal distortion of interactions through label-free detection, multi-target screening analysis, which has a great potential to be a complementary screening platform in the field of drug discovery.

Small molecule proteostasis regulators that reprogram the ER to reduce extracellular protein aggregation

PubMed Central

Plate, Lars; Cooley, Christina B; Chen, John J; Paxman, Ryan J; Gallagher, Ciara M; Madoux, Franck; Genereux, Joseph C; Dobbs, Wesley; Garza, Dan; Spicer, Timothy P; Scampavia, Louis; Brown, Steven J; Rosen, Hugh; Powers, Evan T; Walter, Peter; Hodder, Peter; Wiseman, R Luke; Kelly, Jeffery W

2016-01-01

Imbalances in endoplasmic reticulum (ER) proteostasis are associated with etiologically-diverse degenerative diseases linked to excessive extracellular protein misfolding and aggregation. Reprogramming of the ER proteostasis environment through genetic activation of the Unfolded Protein Response (UPR)-associated transcription factor ATF6 attenuates secretion and extracellular aggregation of amyloidogenic proteins. Here, we employed a screening approach that included complementary arm-specific UPR reporters and medium-throughput transcriptional profiling to identify non-toxic small molecules that phenocopy the ATF6-mediated reprogramming of the ER proteostasis environment. The ER reprogramming afforded by our molecules requires activation of endogenous ATF6 and occurs independent of global ER stress. Furthermore, our molecules phenocopy the ability of genetic ATF6 activation to selectively reduce secretion and extracellular aggregation of amyloidogenic proteins. These results show that small molecule-dependent ER reprogramming, achieved through preferential activation of the ATF6 transcriptional program, is a promising strategy to ameliorate imbalances in ER function associated with degenerative protein aggregation diseases. DOI: http://dx.doi.org/10.7554/eLife.15550.001 PMID:27435961

Drug transport across the blood–brain barrier

PubMed Central

Pardridge, William M

2012-01-01

The blood–brain barrier (BBB) prevents the brain uptake of most pharmaceuticals. This property arises from the epithelial-like tight junctions within the brain capillary endothelium. The BBB is anatomically and functionally distinct from the blood–cerebrospinal fluid barrier at the choroid plexus. Certain small molecule drugs may cross the BBB via lipid-mediated free diffusion, providing the drug has a molecular weight <400 Da and forms <8 hydrogen bonds. These chemical properties are lacking in the majority of small molecule drugs, and all large molecule drugs. Nevertheless, drugs can be reengineered for BBB transport, based on the knowledge of the endogenous transport systems within the BBB. Small molecule drugs can be synthesized that access carrier-mediated transport (CMT) systems within the BBB. Large molecule drugs can be reengineered with molecular Trojan horse delivery systems to access receptor-mediated transport (RMT) systems within the BBB. Peptide and antisense radiopharmaceuticals are made brain-penetrating with the combined use of RMT-based delivery systems and avidin–biotin technology. Knowledge on the endogenous CMT and RMT systems expressed at the BBB enable new solutions to the problem of BBB drug transport. PMID:22929442

Spectroscopy of PAHs with carbon side chains

NASA Astrophysics Data System (ADS)

Rouille, G.; Steglich, M.; Carpentier, Y.; Huisken, F.; Henning, T.

2011-05-01

The presence of polycyclic aromatic hydrocarbons (PAHs) in space has been inferred ever since sp ecific infrared emission bands were interpreted as their collective fingerprint. In parallel, it has been admitted that the famous diffuse interstellar bands (DIBs), which are absorption features observed in the visible wavelength range, are bands belonging to the electronic spectra of free-flying interstellar molecules yet to be identified. As neutral PAHs of medium and large sizes exhibit absorption bands in the range where the DIBs are found, these molecules, which also fulfill other criteria, have been proposed as potential carriers. Studies of small PAHs in solutions have shown that adding an ethynyl side chain (--CCH) to their structure causes their electronic transitions to shift toward longer wavelengths. This fact, added to the observations of interstellar polyynyl radicals, motivated our current research project on PAHs carrying polyynyl side chains. In a first stage, we are measuring the electronic spectra of small PAHs and of their ethynyl and butadiynyl (--CCCCH) derivatives at cryogenic temperatures in rare gas matrices. Then, measurements will be carried out in supersonic jets, providing us with spectra obtained under conditions relevant to the study of free-flying interstellar molecules. The results of IR absorption measurements will be included in our set of new data. As a complement to our laboratory study on the substituted PAHs, quantum chemical calculations are carried out to interprete and simulate their IR and vibronic spectra. We use the density functional theory approach and its time-dependent extension for calculating the electronic ground states and the electronically excited states, respectively. Through the analysis of the new data, it will be determined whether PAHs carrying polyynyl side chains can play a role in interstellar phenomena. The latest results of this on-going project will be presented.

Small molecule fluoride toxicity agonists.

PubMed

Nelson, James W; Plummer, Mark S; Blount, Kenneth F; Ames, Tyler D; Breaker, Ronald R

2015-04-23

Fluoride is a ubiquitous anion that inhibits a wide variety of metabolic processes. Here, we report the identification of a series of compounds that enhance fluoride toxicity in Escherichia coli and Streptococcus mutans. These molecules were isolated by using a high-throughput screen (HTS) for compounds that increase intracellular fluoride levels as determined via a fluoride riboswitch reporter fusion construct. A series of derivatives were synthesized to examine structure-activity relationships, leading to the identification of compounds with improved activity. Thus, we demonstrate that small molecule fluoride toxicity agonists can be identified by HTS from existing chemical libraries by exploiting a natural fluoride riboswitch. In addition, our findings suggest that some molecules might be further optimized to function as binary antibacterial agents when combined with fluoride. Copyright © 2015 Elsevier Ltd. All rights reserved.

Vibronic coupling simulations for linear and nonlinear optical processes: Simulation results

NASA Astrophysics Data System (ADS)

Silverstein, Daniel W.; Jensen, Lasse

2012-02-01

A vibronic coupling model based on time-dependent wavepacket approach is applied to simulate linear optical processes, such as one-photon absorbance and resonance Raman scattering, and nonlinear optical processes, such as two-photon absorbance and resonance hyper-Raman scattering, on a series of small molecules. Simulations employing both the long-range corrected approach in density functional theory and coupled cluster are compared and also examined based on available experimental data. Although many of the small molecules are prone to anharmonicity in their potential energy surfaces, the harmonic approach performs adequately. A detailed discussion of the non-Condon effects is illustrated by the molecules presented in this work. Linear and nonlinear Raman scattering simulations allow for the quantification of interference between the Franck-Condon and Herzberg-Teller terms for different molecules.

Intuitive Density Functional Theory-Based Energy Decomposition Analysis for Protein-Ligand Interactions.

PubMed

Phipps, M J S; Fox, T; Tautermann, C S; Skylaris, C-K

2017-04-11

First-principles quantum mechanical calculations with methods such as density functional theory (DFT) allow the accurate calculation of interaction energies between molecules. These interaction energies can be dissected into chemically relevant components such as electrostatics, polarization, and charge transfer using energy decomposition analysis (EDA) approaches. Typically EDA has been used to study interactions between small molecules; however, it has great potential to be applied to large biomolecular assemblies such as protein-protein and protein-ligand interactions. We present an application of EDA calculations to the study of ligands that bind to the thrombin protein, using the ONETEP program for linear-scaling DFT calculations. Our approach goes beyond simply providing the components of the interaction energy; we are also able to provide visual representations of the changes in density that happen as a result of polarization and charge transfer, thus pinpointing the functional groups between the ligand and protein that participate in each kind of interaction. We also demonstrate with this approach that we can focus on studying parts (fragments) of ligands. The method is relatively insensitive to the protocol that is used to prepare the structures, and the results obtained are therefore robust. This is an application to a real protein drug target of a whole new capability where accurate DFT calculations can produce both energetic and visual descriptors of interactions. These descriptors can be used to provide insights for tailoring interactions, as needed for example in drug design.

The fundamental unit of pain is the cell.

PubMed

Reichling, David B; Green, Paul G; Levine, Jon D

2013-12-01

The molecular/genetic era has seen the discovery of a staggering number of molecules implicated in pain mechanisms [18,35,61,69,96,133,150,202,224]. This has stimulated pharmaceutical and biotechnology companies to invest billions of dollars to develop drugs that enhance or inhibit the function of many these molecules. Unfortunately this effort has provided a remarkably small return on this investment. Inevitably, transformative progress in this field will require a better understanding of the functional links among the ever-growing ranks of "pain molecules," as well as their links with an even larger number of molecules with which they interact. Importantly, all of these molecules exist side-by-side, within a functional unit, the cell, and its adjacent matrix of extracellular molecules. To paraphrase a recent editorial in Science magazine [223], although we live in the Golden age of Genetics, the fundamental unit of biology is still arguably the cell, and the cell is the critical structural and functional setting in which the function of pain-related molecules must be understood. This review summarizes our current understanding of the nociceptor as a cell-biological unit that responds to a variety of extracellular inputs with a complex and highly organized interaction of signaling molecules. We also discuss the insights that this approach is providing into peripheral mechanisms of chronic pain and sex dependence in pain.

Small-molecule inhibitors of phosphatidylcholine transfer protein/StarD2 identified by high-throughput screening.

PubMed

Wagle, Neil; Xian, Jun; Shishova, Ekaterina Y; Wei, Jie; Glicksman, Marcie A; Cuny, Gregory D; Stein, Ross L; Cohen, David E

2008-12-01

Phosphatidylcholine transfer protein (PC-TP, also referred to as StarD2) is a highly specific intracellular lipid-binding protein that catalyzes the transfer of phosphatidylcholines between membranes in vitro. Recent studies have suggested that PC-TP in vivo functions to regulate fatty acid and glucose metabolism, possibly via interactions with selected other proteins. To begin to address the relationship between activity in vitro and biological function, we undertook a high-throughput screen to identify small-molecule inhibitors of the phosphatidylcholine transfer activity of PC-TP. After adapting a fluorescence quench assay to measure phosphatidylcholine transfer activity, we screened 114,752 compounds of a small-molecule library. The high-throughput screen identified 14 potential PC-TP inhibitors. Of these, 6 compounds exhibited characteristics consistent with specific inhibition of PC-TP activity, with IC(50) values that ranged from 4.1 to 95.0muM under conditions of the in vitro assay. These compounds should serve as valuable reagents to elucidate the biological function of PC-TP. Because mice with homozygous disruption of the PC-TP gene (Pctp) are sensitized to insulin action and relatively resistant to the development of atherosclerosis, these inhibitors may also prove to be of value in the management of diabetes and atherosclerotic cardiovascular diseases.

Development of protein A functionalized microcantilever immunosensors for the analyses of small molecules at parts per trillion levels.

PubMed

Tan, Weiming; Huang, Yuan; Nan, Tiegui; Xue, Changguo; Li, Zhaohu; Zhang, Qingchuan; Wang, Baomin

2010-01-15

Development of microcantilever biosensors for small molecules was exemplified with the beta-adrenergic agonist clenbuterol and the antibiotic chloramphenicol. In this paper, antibody sulfhydrylation and protein A were used to modify the microcantilever Au surface, and the antibody activities on the microcantilever were evaluated with direct competitive enzyme-linked immunosorbent assay (dcELISA). The activity of the antibodies immobilized on the microcantilever via protein A was 1.7-fold of that via the sulfhydrylation reagent 2-iminothiolane hydrochloride. A microcantilever immunosensor method with protein A as the functionalization reagent was established to detect the residues of clenbuterol and chloramphenicol at limits of detection (LOD) of approximately 0.1 and 0.2 ng/mL, respectively. Such LODs were better than that of the corresponding dcELISAs. The concentration of clenbuterol in a fortified feed sample detected with the microcantilever immunosensor after thorough extraction and purification agreed well with that detected with the dcELISA. Protein A showed to be simple and reproducible for functionalization of the antibodies on the Au surface and, thus, has common application values in microcantilever immunosensor development. The results suggest that microcantilever immunosensors be suitable for detection of small molecules, and the assay sensitivity is mainly related to the quality and activities of the antibodies.

Designing small molecule polyaromatic p- and n-type semiconductor materials for organic electronics

NASA Astrophysics Data System (ADS)

Collis, Gavin E.

2015-12-01

By combining computational aided design with synthetic chemistry, we are able to identify core 2D polyaromatic small molecule templates with the necessary optoelectronic properties for p- and n-type materials. By judicious selection of the functional groups, we can tune the physical properties of the material making them amenable to solution and vacuum deposition. In addition to solubility, we observe that the functional group can influence the thin film molecular packing. By developing structure-property relationships (SPRs) for these families of compounds we observe that some compounds are better suited for use in organic solar cells, while others, varying only slightly in structure, are favoured in organic field effect transistor devices. We also find that the processing conditions can have a dramatic impact on molecular packing (i.e. 1D vs 2D polymorphism) and charge mobility; this has implications for material and device long term stability. We have developed small molecule p- and n-type materials for organic solar cells with efficiencies exceeding 2%. Subtle variations in the functional groups of these materials produces p- and ntype materials with mobilities higher than 0.3 cm2/Vs. We are also interested in using our SPR approach to develop materials for sensor and bioelectronic applications.

Use of a small molecule as an initiator for interchain staudinger reaction: A new ATP sensing platform using product fluorescence.

PubMed

Yu, Huan; Zheng, Jing; Yang, Sheng; Asiri, Abdullah M; Alamry, Khalid A; Sun, Mingtai; Zhang, Kui; Wang, Suhua; Yang, Ronghua

2018-02-01

We demonstrated that a small molecule induced interchain Staudinger reaction can be employed for highly selective detection of adenosine triphosphate (ATP), an important energy-storage biomolecule. A designed ATP split aptamer (A1) was first functionalized with a weakly fluorescent coumarin derivative due to an azide group (azido-coumarin). The second DNA strand (A2) was covalently linked with triphenylphosphine, which could selectively and efficiently reduce azido to amino group through the Staudinger reaction. The A2 was then hybridized with a half of another designed longer DNA strand (T1). The second half of T1 was a split aptamer and selectively recognized ATP with A1 to form a sandwich structure. The specific interaction between ATP and the aptamers drew the two functionalized DNA strands (A1 and A2) together to initiate the interchain Staudinger reduction at fmol-nmol concentration level, hence produced fluorescent 7-aminocoumarin which could be used as an indicator for the presence of trace ATP. The reaction process had a concentration dependent manner with ATP in a large concentration range. Such a strategy of interchain Staudinger reaction can be extended to construct biosensors for other small functional molecules on the basis of judiciously designed aptamers. Copyright © 2017 Elsevier B.V. All rights reserved.

Inhibition of amyloid oligomerization into different supramolecular architectures by small molecules: mechanistic insights and design rules.

PubMed

Brahmachari, Sayanti; Paul, Ashim; Segal, Daniel; Gazit, Ehud

2017-05-01

Protein misfolding and aggregation have been associated with several human disorders, including Alzheimer's, Parkinson's and Huntington's diseases, as well as senile systemic amyloidosis and Type II diabetes. However, there is no current disease-modifying therapy available for the treatment of these disorders. In spite of extensive academic, pharmaceutical, medicinal and clinical research, a complete mechanistic model for this family of diseases is still lacking. In this review, we primarily discuss the different types of small molecular entities which have been used for the inhibition of the aggregation process of different amyloidogenic proteins under diseased conditions. These include small peptides, polyphenols, inositols, quinones and their derivatives, and metal chelator molecules. In recent years, these groups of molecules have been extensively studied using in vitro, in vivo and computational models to understand their mechanism of action and common structural features underlying the process of inhibition. A salient feature found to be instrumental in the process of inhibition is the balance between the aromatic unit that functions as the amyloid recognition unit and the hydrophilic amyloid breaker unit. The establishment of structure-function relationship for amyloid-modifying therapies by the various functional entities should serve as an important step toward the development of efficient therapeutics.

Multi-functional ultrathin Pd xCu 1-x and Pt~Pd xCu 1-x one-dimensional nanowire motifs for various small molecule oxidation reactions

DOE PAGES

Liu, Haiqing; Wong, Stanislaus S.; Adzic, Radoslav R.

2015-11-18

Developing novel electrocatalysts for small molecule oxidation processes, including formic acid oxidation (FAOR), methanol oxidation reaction (MOR), and ethanol oxidation reaction (EOR), denoting the key anodic reactions for their respective fuel cell configurations, is a significant and relevant theme of recent efforts in the field. Herein, in this report, we demonstrated a concerted effort to couple and combine the benefits of small size, anisotropic morphology, and tunable chemical composition in order to devise a novel “family” of functional architectures. In particular, we have fabricated not only ultrathin 1-D Pd 1–xCu x alloys but also Pt-coated Pd 1–xCu x (i.e., Pt~Pdmore » 1–xCu x; herein the ~ indicates an intimate association, but not necessarily actual bond formation, between the inner bimetallic core and the Pt outer shell) core–shell hierarchical nanostructures with readily tunable chemical compositions by utilizing a facile, surfactant-based, wet chemical synthesis coupled with a Cu underpotential deposition technique. Our main finding is that our series of as-prepared nanowires are functionally flexible. More precisely, we demonstrate that various examples within this “family” of structural motifs can be tailored for exceptional activity with all 3 of these important electrocatalytic reactions. In particular, we note that our series of Pd 1–xCu x nanowires all exhibit enhanced FAOR activities as compared with not only analogous Pd ultrathin nanowires but also commercial Pt and Pd standards, with Pd 9Cu representing the “optimal” composition. Moreover, our group of Pt~Pd 1–xCu x nanowires consistently outperformed not only commercial Pt NPs but also ultrathin Pt nanowires by several fold orders of magnitude for both the MOR and EOR reactions in alkaline media. As a result, the variation of the MOR and EOR performance with the chemical composition of our ultrathin Pt~Pd 1–xCu x nanowires was also discussed.« less

Inhibition of Oncogenic functionality of STAT3 Protein by Membrane Anchoring

NASA Astrophysics Data System (ADS)

Liu, Baoxu; Fletcher, Steven; Gunning, Patrick; Gradinaru, Claudiu

2009-03-01

Signal Transducer and Activator of Transcription 3 (STAT3) protein plays an important role in oncogenic processes. A novel molecular therapeutic approach to inhibit the oncogenic functionality of STAT3 is to design a prenylated small peptide sequence which could sequester STAT3 to the plasma membrane. We have also developed a novel fluorescein derivative label (F-NAc), which is much more photostable compared to the popular fluorescein label FITC. Remarkably, the new dye shows fluorescent properties that are invariant over a wide pH range, which is advantageous for our application. We have shown that F-NAc is suitable for single-molecule measurements and its properties are not affected by ligation to biomolecules. The membrane localization via high-affinity prenylated small-molecule binding agents is studied by encapsulating FNAc-labeled STAT3 and inhibitors within a liposome model cell system. The dynamics of the interaction between the protein and the prenylated ligands is investigated at single molecule level. The efficiency and stability of the STAT3 anchoring in lipid membranes are addressed via quantitative confocal imaging and single-molecule spectroscopy using a custom-built multiparameter fluorescence microscope.

Effects of Acids, Bases, and Heteroatoms on Proximal Radial Distribution Functions for Proteins.

PubMed

Nguyen, Bao Linh; Pettitt, B Montgomery

2015-04-14

The proximal distribution of water around proteins is a convenient method of quantifying solvation. We consider the effect of charged and sulfur-containing amino acid side-chain atoms on the proximal radial distribution function (pRDF) of water molecules around proteins using side-chain analogs. The pRDF represents the relative probability of finding any solvent molecule at a distance from the closest or surface perpendicular protein atom. We consider the near-neighbor distribution. Previously, pRDFs were shown to be universal descriptors of the water molecules around C, N, and O atom types across hundreds of globular proteins. Using averaged pRDFs, a solvent density around any globular protein can be reconstructed with controllable relative error. Solvent reconstruction using the additional information from charged amino acid side-chain atom types from both small models and protein averages reveals the effects of surface charge distribution on solvent density and improves the reconstruction errors relative to simulation. Solvent density reconstructions from the small-molecule models are as effective and less computationally demanding than reconstructions from full macromolecular models in reproducing preferred hydration sites and solvent density fluctuations.

Development and Implementation of a High-Throughput High-Content Screening Assay to Identify Inhibitors of Androgen Receptor Nuclear Localization in Castration-Resistant Prostate Cancer Cells

PubMed Central

Nguyen, Minh M.; Dar, Javid A.; Ai, Junkui; Wang, Yujuan; Masoodi, Khalid Z.; Shun, Tongying; Shinde, Sunita; Camarco, Daniel P.; Hua, Yun; Huryn, Donna M.; Wilson, Gabriela Mustata; Lazo, John S.; Nelson, Joel B.; Wipf, Peter

2016-01-01

Abstract Patients with castration-resistant prostate cancer (CRPC) can be treated with abiraterone, a potent inhibitor of androgen synthesis, or enzalutamide, a second-generation androgen receptor (AR) antagonist, both targeting AR signaling. However, most patients relapse after several months of therapy and a majority of patients with relapsed CRPC tumors express the AR target gene prostate-specific antigen (PSA), suggesting that AR signaling is reactivated and can be targeted again to inhibit the relapsed tumors. Novel small molecules capable of inhibiting AR function may lead to urgently needed therapies for patients resistant to abiraterone, enzalutamide, and/or other previously approved antiandrogen therapies. Here, we describe a high-throughput high-content screening (HCS) campaign to identify small-molecule inhibitors of AR nuclear localization in the C4-2 CRPC cell line stably transfected with GFP-AR-GFP (2GFP-AR). The implementation of this HCS assay to screen a National Institutes of Health library of 219,055 compounds led to the discovery of 3 small molecules capable of inhibiting AR nuclear localization and function in C4-2 cells, demonstrating the feasibility of using this cell-based phenotypic assay to identify small molecules targeting the subcellular localization of AR. Furthermore, the three hit compounds provide opportunities to develop novel AR drugs with potential for therapeutic intervention in CRPC patients who have relapsed after treatment with antiandrogens, such as abiraterone and/or enzalutamide. PMID:27187604

PDF cycling in the dorsal protocerebrum of the Drosophila brain is not necessary for circadian clock function.

PubMed

Kula, Elzbieta; Levitan, Edwin S; Pyza, Elzbieta; Rosbash, Michael

2006-04-01

In Drosophila, the neuropeptide pigment-dispersing factor (PDF) is a likely circadian molecule, secreted by central pacemaker neurons (LNvs). PDF is expressed in both small and large LNvs (sLNvs and lLNvs), and there are striking circadian oscillations of PDF staining intensity in the small cell termini, which require a functional molecular clock. This cycling may be relevant to the proposed role of PDF as a synchronizer of the clock system or as an output signal connecting pacemaker cells to locomotor activity centers. In this study, the authors use a generic neuropeptide fusion protein (atrial natriuretic factor-green fluorescent protein [ANF-GFP]) and show that it can be expressed in the same neurons as PDF itself. Yet, ANF-GFP as well as PDF itself does not manifest any cyclical accumulation in sLNv termini in adult transgenic flies. Surprisingly, the absence of detectable PDF cycling is not accompanied by any detectable behavioral pheno-type, since these transgenic flies have normal morning and evening anticipation in a light-dark cycle (LD) and are fully rhythmic in constant darkness (DD). The molecular clock is also not compromised. The results suggest that robust PDF cycling in sLNv termini plays no more than a minor role in the Drosophila circadian system and is apparently not even necessary for clock output function.

Single-molecule photon emission statistics for systems with explicit time dependence: Generating function approach

NASA Astrophysics Data System (ADS)

Peng, Yonggang; Xie, Shijie; Zheng, Yujun; Brown, Frank L. H.

2009-12-01

Generating function calculations are extended to allow for laser pulse envelopes of arbitrary shape in numerical applications. We investigate photon emission statistics for two-level and V- and Λ-type three-level systems under time-dependent excitation. Applications relevant to electromagnetically induced transparency and photon emission from single quantum dots are presented.

Improved Dye Stability in Single-Molecule Fluorescence Experiments

NASA Astrophysics Data System (ADS)

EcheverrÍa Aitken, Colin; Marshall, R. Andrew; Pugi, Joseph D.

Complex biological systems challenge existing single-molecule methods. In particular, dye stability limits observation time in singlemolecule fluorescence applications. Current approaches to improving dye performance involve the addition of enzymatic oxygen scavenging systems and small molecule additives. We present an enzymatic oxygen scavenging system that improves dye stability in single-molecule experiments. Compared to the currently-employed glucose-oxidase/catalase system, the protocatechuate-3,4-dioxygenase system achieves lower dissolved oxygen concentration and stabilizes single Cy3, Cy5, and Alexa488 fluorophores. Moreover, this system possesses none of the limitations associated with the glucose oxidase/catalase system. We also tested the effects of small molecule additives in this system. Biological reducing agents significantly destabilize the Cy5 fluorophore as a function of reducing potential. In contrast, anti-oxidants stabilize the Cy3 and Alexa488 fluorophores. We recommend use of the protocatechuate-3,4,-dioxygenase system with antioxidant additives, and in the absence of biological reducing agents. This system should have wide application to single-molecule fluorescence experiments.

Clinical trial considerations on male contraception and collection of pregnancy information from female partner: update.

PubMed

Banholzer, Maria Longauer; Wandel, Christoph; Barrow, Paul; Mannino, Marie; Schmitt, Georg; Guérard, Melanie; Müller, Lutz; Greig, Gerard; Amemiya, Kenjie; Peck, Richard; Singer, Thomas; Doessegger, Lucette

2016-12-01

This is an update to our 2012 publication on clinical trial considerations on male contraception and collection of pregnancy information from female partner, after critical review of recent (draft) guidances released by the International Council for Harmonisation [ICH] the Clinical Trial Facilitation Group [CTFG] and the US Food & Drug Administration [FDA]. Relevant aspects of the new guidance documents are discussed in the context of male contraception and pregnancy reporting from female partner in clinical trials and the approach is updated accordingly. Genotoxicity The concept of a threshold is introduced using acceptable daily intake/permissible daily exposure to define genotoxicity requirements, hence highly effective contraception in order to avoid conception. The duration for highly effective contraception has been extended from 74 to 90 days from the end of relevant systemic exposure. Teratogenicity Pharmacokinetic considerations to estimate safety margins have been contextualized with regard to over- and underestimation of the risk of teratogenicity transmitted by a vaginal dose. The duration of male contraception after the last dose takes into account the end of relevant systemic exposure if measured, or a default period of five half-lives after last dose for small molecules and two half-lives for immunoglobulins (mAbs). Measures to prevent exposure of the conceptus via a vaginal dose apply to reproductively competent or vasectomized men, unless measurements fail to detect the compound in seminal fluid. Critical review of new guidance documents provides a comparison across approaches and resulted in an update of our previous publication. Separate algorithms for small molecules and monoclonal antibodies are proposed to guide the recommendations for contraception for male trial participants and pregnancy reporting from female partners. No male contraception is required if the dose is below a defined threshold for genotoxic concern applicable to small molecules. For men treated with teratogenic mAbs, condom use to prevent exposure of a potentially pregnant partner is unlikely to be recommended because of the minimal female exposure anticipated following a vaginal dose. The proposed safety margins for teratogenicity may evolve with further knowledge.

A systematic benchmark of the ab initio Bethe-Salpeter equation approach for low-lying optical excitations of small organic molecules

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

Bruneval, Fabien; Molecular Foundry, Lawrence Berkeley National Laboratory, Berkeley, California 94720; Department of Physics, University of California, Berkeley, California 94720

2015-06-28

The predictive power of the ab initio Bethe-Salpeter equation (BSE) approach, rigorously based on many-body Green’s function theory but incorporating information from density functional theory, has already been demonstrated for the optical gaps and spectra of solid-state systems. Interest in photoactive hybrid organic/inorganic systems has recently increased and so has the use of the BSE for computing neutral excitations of organic molecules. However, no systematic benchmarks of the BSE for neutral electronic excitations of organic molecules exist. Here, we study the performance of the BSE for the 28 small molecules in Thiel’s widely used time-dependent density functional theory benchmark setmore » [Schreiber et al., J. Chem. Phys. 128, 134110 (2008)]. We observe that the BSE produces results that depend critically on the mean-field starting point employed in the perturbative approach. We find that this starting point dependence is mainly introduced through the quasiparticle energies obtained at the intermediate GW step and that with a judicious choice of starting mean-field, singlet excitation energies obtained from BSE are in excellent quantitative agreement with higher-level wavefunction methods. The quality of the triplet excitations is slightly less satisfactory.« less

Extracting Social Information from Chemosensory Cues: Consideration of Several Scenarios and Their Functional Implications

PubMed Central

Ben-Shaul, Yoram

2015-01-01

Across all sensory modalities, stimuli can vary along multiple dimensions. Efficient extraction of information requires sensitivity to those stimulus dimensions that provide behaviorally relevant information. To derive social information from chemosensory cues, sensory systems must embed information about the relationships between behaviorally relevant traits of individuals and the distributions of the chemical cues that are informative about these traits. In simple cases, the mere presence of one particular compound is sufficient to guide appropriate behavior. However, more generally, chemosensory information is conveyed via relative levels of multiple chemical cues, in non-trivial ways. The computations and networks needed to derive information from multi-molecule stimuli are distinct from those required by single molecule cues. Our current knowledge about how socially relevant information is encoded by chemical blends, and how it is extracted by chemosensory systems is very limited. This manuscript explores several scenarios and the neuronal computations required to identify them. PMID:26635515

Regulation of antiapoptotic MCL-1 function by gossypol: mechanistic insights from in vitro reconstituted systems.

PubMed

Etxebarria, Aitor; Landeta, Olatz; Antonsson, Bruno; Basañez, Gorka

2008-12-01

Small-molecule drugs that induce apoptosis in tumor cells by activation of the BCL-2-regulated mitochondrial outer membrane permeabilization (MOMP) pathway hold promise for rational anticancer therapies. Accumulating evidence indicates that the natural product gossypol and its derivatives can kill tumor cells by targeting antiapoptotic BCL-2 family members in such a manner as to trigger MOMP. However, due to the inherent complexity of the cellular apoptotic network, the precise mechanisms by which interactions between gossypol and individual BCL-2 family members lead to MOMP remain poorly understood. Here, we used simplified systems bearing physiological relevance to examine the impact of gossypol on the function of MCL-1, a key determinant for survival of various human malignancies that has become a highly attractive target for anticancer drug design. First, using a reconstituted liposomal system that recapitulates basic aspects of the BCL-2-regulated MOMP pathway, we demonstrate that MCL-1 inhibits BAX permeabilizing function via a "dual-interaction" mechanism, while submicromolar concentrations of gossypol reverse MCL-1-mediated inhibition of functional BAX activation. Solution-based studies showed that gossypol competes with BAX/BID BH3 ligands for binding to MCL-1 hydrophobic groove, thereby providing with a mechanistic explanation for how gossypol restores BAX permeabilizing function in the presence of MCL-1. By contrast, no evidence was found indicating that gossypol transforms MCL-1 into a BAX-like pore-forming molecule. Altogether, our findings validate MCL-1 as a direct target of gossypol, and highlight that making this antiapoptotic protein unable to inhibit BAX-driven MOMP may represent one important mechanism by which gossypol exerts its cytotoxic effect in selected cancer cells.

Organic Nanoflowers from a Wide Variety of Molecules Templated by a Hierarchical Supramolecular Scaffold.

PubMed

Negrón, Luis M; Díaz, Tanya L; Ortiz-Quiles, Edwin O; Dieppa-Matos, Diómedes; Madera-Soto, Bismark; Rivera, José M

2016-03-15

Nanoflowers (NFs) are flowered-shaped particles with overall sizes or features in the nanoscale. Beyond their pleasing aesthetics, NFs have found a number of applications ranging from catalysis, to sensing, to drug delivery. Compared to inorganic based NFs, their organic and hybrid counterparts are relatively underdeveloped mostly because of the lack of a reliable and versatile method for their construction. We report here a method for constructing NFs from a wide variety of biologically relevant molecules (guests), ranging from small molecules, like doxorubicin, to biomacromolecules, like various proteins and plasmid DNA. The method relies on the encapsulation of the guests within a hierarchically structured particle made from supramolecular G-quadruplexes. The size and overall flexibility of the guests dictate the broad morphological features of the resulting NFs, specifically, small and rigid guests favor the formation of NFs with spiky petals, while large and/or flexible guests promote NFs with wide petals. The results from experiments using confocal fluorescence microscopy, and scanning electron microscopy provides the basis for the proposed mechanism for the NF formation.

K+ Efflux-Independent NLRP3 Inflammasome Activation by Small Molecules Targeting Mitochondria.

PubMed

Groß, Christina J; Mishra, Ritu; Schneider, Katharina S; Médard, Guillaume; Wettmarshausen, Jennifer; Dittlein, Daniela C; Shi, Hexin; Gorka, Oliver; Koenig, Paul-Albert; Fromm, Stephan; Magnani, Giovanni; Ćiković, Tamara; Hartjes, Lara; Smollich, Joachim; Robertson, Avril A B; Cooper, Matthew A; Schmidt-Supprian, Marc; Schuster, Michael; Schroder, Kate; Broz, Petr; Traidl-Hoffmann, Claudia; Beutler, Bruce; Kuster, Bernhard; Ruland, Jürgen; Schneider, Sabine; Perocchi, Fabiana; Groß, Olaf

2016-10-18

Imiquimod is a small-molecule ligand of Toll-like receptor-7 (TLR7) that is licensed for the treatment of viral infections and cancers of the skin. Imiquimod has TLR7-independent activities that are mechanistically unexplained, including NLRP3 inflammasome activation in myeloid cells and apoptosis induction in cancer cells. We investigated the mechanism of inflammasome activation by imiquimod and the related molecule CL097 and determined that K + efflux was dispensable for NLRP3 activation by these compounds. Imiquimod and CL097 inhibited the quinone oxidoreductases NQO2 and mitochondrial Complex I. This induced a burst of reactive oxygen species (ROS) and thiol oxidation, and led to NLRP3 activation via NEK7, a recently identified component of this inflammasome. Metabolic consequences of Complex I inhibition and endolysosomal effects of imiquimod might also contribute to NLRP3 activation. Our results reveal a K + efflux-independent mechanism for NLRP3 activation and identify targets of imiquimod that might be clinically relevant. Copyright © 2016 Elsevier Inc. All rights reserved.

Molecular self-assembly for biological investigations and nanoscale lithography

NASA Astrophysics Data System (ADS)

Cheunkar, Sarawut

Small, diffusible molecules when recognized by their binding partners, such as proteins and antibodies, trigger enzymatic activity, cell communication, and immune response. Progress in analytical methods enabling detection, characterization, and visualization of biological dynamics at the molecular level will advance our exploration of complex biological systems. In this dissertation, analytical platforms were fabricated to capture membrane-associated receptors, which are essential proteins in cell signaling pathways. The neurotransmitter serotonin and its biological precursor were immobilized on gold substrates coated with self-assembled monolayers (SAMs) of oligo(ethylene glycol)alkanethiols and their reactive derivatives. The SAM-coated substrates present the biologically selective affinity of immobilized molecules to target native membrane-associated receptors. These substrates were also tested for biospecificity using antibodies. In addition, small-molecule-functionalized platforms, expressing neurotransmitter pharmacophores, were employed to examine kinetic interactions between G-protein-coupled receptors and their associated neurotransmitters. The binding interactions were monitored using a quartz crystal microbalance equipped with liquid-flow injection. The interaction kinetics of G-protein-coupled serotonin 1A receptor and 5-hydroxytyptophan-functionalized surfaces were studied in a real-time, label-free environment. Key binding parameters, such as equilibrium dissociation constants, binding rate constants, and dissociative half-life, were extracted. These parameters are critical for understanding and comparing biomolecular interactions in modern biomedical research. By integrating self-assembly, surface functionalization, and nanofabrication, small-molecule microarrays were created for high-throughput screening. A hybrid soft-lithography, called microcontact insertion printing, was used to pattern small molecules at the dilute scales necessary for highly selective biorecognition. By carefully tuning the polar surface energy of polymeric stamps, problems associated with patterning hydrophilic tether molecules inserted into hydrophilic preformed SAMs are surmounted. The patterned substrates presenting neurotransmitter precursors selectively capture membrane-associated receptors. These advances provide new avenues for fabricating small-molecule arrays. Furthermore, a novel strategy based on a conventional microcontact printing, called chemical lift-off lithography, was invented to overcome the micrometer-scale resolution limits of molecular ink diffusion in soft lithography. Self-assembled monolayers of hydroxyl-terminated alkanethiols, preformed on gold substrates, were selectively removed by oxygen-plasma-treated polymeric stamps in a subtractive stamping process with high pattern fidelity. The covalent interactions formed at the stamp-substrate interface are believed to be responsible for removing not only alkanethiol molecules but also a monolayer of gold atoms from the substrates. A variety of high-resolution patterned features were fabricated, and stamps were cleaned and reused many times without feature deterioration. The remaining SAMs acted as resists for etching exposed gold features. Monolayer backfilling into lifted-off areas enabled patterned protein capture, and 40-nanometer chemical patterns were achieved.

Autocatalytic, bistable, oscillatory networks of biologically relevant organic reactions.

PubMed

Semenov, Sergey N; Kraft, Lewis J; Ainla, Alar; Zhao, Mengxia; Baghbanzadeh, Mostafa; Campbell, Victoria E; Kang, Kyungtae; Fox, Jerome M; Whitesides, George M

2016-09-29

Networks of organic chemical reactions are important in life and probably played a central part in its origin. Network dynamics regulate cell division, circadian rhythms, nerve impulses and chemotaxis, and guide the development of organisms. Although out-of-equilibrium networks of chemical reactions have the potential to display emergent network dynamics such as spontaneous pattern formation, bistability and periodic oscillations, the principles that enable networks of organic reactions to develop complex behaviours are incompletely understood. Here we describe a network of biologically relevant organic reactions (amide formation, thiolate-thioester exchange, thiolate-disulfide interchange and conjugate addition) that displays bistability and oscillations in the concentrations of organic thiols and amides. Oscillations arise from the interaction between three subcomponents of the network: an autocatalytic cycle that generates thiols and amides from thioesters and dialkyl disulfides; a trigger that controls autocatalytic growth; and inhibitory processes that remove activating thiol species that are produced during the autocatalytic cycle. In contrast to previous studies that have demonstrated oscillations and bistability using highly evolved biomolecules (enzymes and DNA) or inorganic molecules of questionable biochemical relevance (for example, those used in Belousov-Zhabotinskii-type reactions), the organic molecules we use are relevant to metabolism and similar to those that might have existed on the early Earth. By using small organic molecules to build a network of organic reactions with autocatalytic, bistable and oscillatory behaviour, we identify principles that explain the ways in which dynamic networks relevant to life could have developed. Modifications of this network will clarify the influence of molecular structure on the dynamics of reaction networks, and may enable the design of biomimetic networks and of synthetic self-regulating and evolving chemical systems.

Autocatalytic, bistable, oscillatory networks of biologically relevant organic reactions

NASA Astrophysics Data System (ADS)

Semenov, Sergey N.; Kraft, Lewis J.; Ainla, Alar; Zhao, Mengxia; Baghbanzadeh, Mostafa; Campbell, Victoria E.; Kang, Kyungtae; Fox, Jerome M.; Whitesides, George M.

2016-09-01

Networks of organic chemical reactions are important in life and probably played a central part in its origin. Network dynamics regulate cell division, circadian rhythms, nerve impulses and chemotaxis, and guide the development of organisms. Although out-of-equilibrium networks of chemical reactions have the potential to display emergent network dynamics such as spontaneous pattern formation, bistability and periodic oscillations, the principles that enable networks of organic reactions to develop complex behaviours are incompletely understood. Here we describe a network of biologically relevant organic reactions (amide formation, thiolate-thioester exchange, thiolate-disulfide interchange and conjugate addition) that displays bistability and oscillations in the concentrations of organic thiols and amides. Oscillations arise from the interaction between three subcomponents of the network: an autocatalytic cycle that generates thiols and amides from thioesters and dialkyl disulfides; a trigger that controls autocatalytic growth; and inhibitory processes that remove activating thiol species that are produced during the autocatalytic cycle. In contrast to previous studies that have demonstrated oscillations and bistability using highly evolved biomolecules (enzymes and DNA) or inorganic molecules of questionable biochemical relevance (for example, those used in Belousov-Zhabotinskii-type reactions), the organic molecules we use are relevant to metabolism and similar to those that might have existed on the early Earth. By using small organic molecules to build a network of organic reactions with autocatalytic, bistable and oscillatory behaviour, we identify principles that explain the ways in which dynamic networks relevant to life could have developed. Modifications of this network will clarify the influence of molecular structure on the dynamics of reaction networks, and may enable the design of biomimetic networks and of synthetic self-regulating and evolving chemical systems.

Scanning Tunneling Microscopy Study of the Structure and Interaction Between Carbon Monoxide and Hydrogen on the Ru(0001) Surface

DOE PAGES

Lechner, Barbara A. J.; Feng, Xiaofeng; Feibelman, Peter J.; ...

2017-07-28

Here, we use scanning tunneling microscopy (STM) to investigate the spatial arrangement of carbon monoxide (CO) and hydrogen (H) co-adsorbed on a model catalyst surface, Ru(0001). We find that at cryogenic temperatures CO forms small triangular islands of up to 21 molecules with hydrogen segregated outside of the islands. Furthermore, whereas for small island sizes (3-6 CO molecules) the molecules adsorb at hcp sites, a registry shift towards top sites occurs for larger islands (10-21 CO molecules). To characterize the CO structures better and to help interpret the data, we carried out density functional theory (DFT) calculations of the structuremore » and simulations of the STM images, which reveal a delicate interplay between the repulsions of the different species.« less

Raman imaging of molecular dynamics during cellular events

NASA Astrophysics Data System (ADS)

Fujita, Katsumasa

2017-07-01

To overcome the speed limitation in Raman imaging, we have developed a microscope system that detects Raman spectra from hundreds of points in a sample simultaneously. The sample was illuminated by a line-shaped focus, and Raman scattering from the illuminated positions was measured simultaneously by an imaging spectrophotometer. We applied the line-illumination technique to observe the dynamics of intracellular molecules during cellular events. We found that intracellular cytochrome c can be clearly imaged by resonant Raman scattering. We demonstrated label-free imaging of redistribution of cytochrome c during apoptosis and osteoblastic mineralization. We also proposed alkyne-tagged Raman imaging to observe small molecules in living cells. Due to its small size and the unique Raman band, alkyne can tag molecules without strong perturbation to molecular functions and with the capability to be detected separately from endogenous molecules.

Scanning Tunneling Microscopy Study of the Structure and Interaction Between Carbon Monoxide and Hydrogen on the Ru(0001) Surface

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

Lechner, Barbara A. J.; Feng, Xiaofeng; Feibelman, Peter J.

Here, we use scanning tunneling microscopy (STM) to investigate the spatial arrangement of carbon monoxide (CO) and hydrogen (H) co-adsorbed on a model catalyst surface, Ru(0001). We find that at cryogenic temperatures CO forms small triangular islands of up to 21 molecules with hydrogen segregated outside of the islands. Furthermore, whereas for small island sizes (3-6 CO molecules) the molecules adsorb at hcp sites, a registry shift towards top sites occurs for larger islands (10-21 CO molecules). To characterize the CO structures better and to help interpret the data, we carried out density functional theory (DFT) calculations of the structuremore » and simulations of the STM images, which reveal a delicate interplay between the repulsions of the different species.« less

Confined water: a Mercedes-Benz model study.

PubMed

Urbic, T; Vlachy, V; Dill, K A

2006-03-16

We study water that is confined within small geometric spaces. We use the Mercedes-Benz (MB) model of water, in NVT and muVT Monte Carlo computer simulations. For MB water molecules between two planes separated by a distance d, we explore the structures, hydrogen bond networks, and thermodynamics as a function of d, temperature T, and water chemical potential mu. We find that squeezing the planes close enough together leads to a vaporization of waters out of the cavity. This vaporization transition has a corresponding peak in the heat capacity of the water. We also find that, in small pores, hydrogen bonding is not isotropic but, rather, it preferentially forms chains along the axis of the cavity. This may be relevant for fast proton transport in pores. Our simulations show oscillations in the forces between the inert plates, due to water structure, even for plate separations of 5-10 water diameters, consistent with experiments by Israelachvili et al. [Nature 1983, 306, 249]. Finally, we find that confinement affects water's heat capacity, consistent with recent experiments of Tombari et al. on Vycor nanopores [J. Chem. Phys. 2005, 122, 104712].

Two Novel Determinants of Etoposide Resistance in Small Cell Lung Cancer

PubMed Central

Lawson, Malcolm H; Cummings, Natalie M; Rassl, Doris M; Russell, Roslin; Brenton, James D; Rintoul, Robert C; Murphy, Gillian

2011-01-01

Patient survival in small cell lung cancer (SCLC) is limited by acquired chemoresistance. Here we report the use of a biologically relevant model to identify novel candidate genes mediating in vivo acquired resistance to etoposide. Candidate genes derived from a cDNA microarray analysis were cloned and transiently overexpressed to evaluate their potential functional roles. We identified two promising genes in the DNA repair enzyme DNA Polymerase β and in the neuroendocrine transcription factor NKX2.2. Specific inhibition of DNA Polymerase β reduced the numbers of cells surviving treatment with etoposide and increased the amount of DNA damage in cells. Conversely, stable overexpression of NKX2.2 increased cell survival in response to etoposide in SCLC cell lines. Consistent with these findings, we found that an absence of nuclear staining for NKX2.2 in SCLC primary tumors was an independent predictor of improved outcomes in chemotherapy-treated patients. Taken together, our findings justify future prospective studies to confirm the roles of these molecules in mediating chemotherapy resistance in SCLC. PMID:21642373

Differential Targeting of Gβγ-Subunit Signaling with Small Molecules

NASA Astrophysics Data System (ADS)

Bonacci, Tabetha M.; Mathews, Jennifer L.; Yuan, Chujun; Lehmann, David M.; Malik, Sundeep; Wu, Dianqing; Font, Jose L.; Bidlack, Jean M.; Smrcka, Alan V.

2006-04-01

G protein βγ subunits have potential as a target for therapeutic treatment of a number of diseases. We performed virtual docking of a small-molecule library to a site on Gβγ subunits that mediates protein interactions. We hypothesized that differential targeting of this surface could allow for selective modulation of Gβγ subunit functions. Several compounds bound to Gβγ subunits with affinities from 0.1 to 60 μM and selectively modulated functional Gβγ-protein-protein interactions in vitro, chemotactic peptide signaling pathways in HL-60 leukocytes, and opioid receptor-dependent analgesia in vivo. These data demonstrate an approach for modulation of G protein-coupled receptor signaling that may represent an important therapeutic strategy.

Branched terthiophenes in organic electronics: from small molecules to polymers.

PubMed

Scheuble, Martin; Goll, Miriam; Ludwigs, Sabine

2015-01-01

A zoo of chemical structures is accessible when the branched unit 2,2':3',2″-terthiophene (3T) is included both in structurally well-defined small molecules and polymer-like architectures. The first part of this review article highlights literature on all-thiophene based branched oligomers including dendrimers as well as combinations of 3T-units with functional moieties for light-harvesting systems. Motivated by the perfectly branched macromolecular dendrimers both electropolymerization as well as chemical approaches are presented as methods for the preparation of branched polythiophenes with different branching densities. Structure-function relationships between the molecular architecture and optical and electronic properties are discussed throughout the article. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Printing Proteins as Microarrays for High-Throughput Function Determination

NASA Astrophysics Data System (ADS)

MacBeath, Gavin; Schreiber, Stuart L.

2000-09-01

Systematic efforts are currently under way to construct defined sets of cloned genes for high-throughput expression and purification of recombinant proteins. To facilitate subsequent studies of protein function, we have developed miniaturized assays that accommodate extremely low sample volumes and enable the rapid, simultaneous processing of thousands of proteins. A high-precision robot designed to manufacture complementary DNA microarrays was used to spot proteins onto chemically derivatized glass slides at extremely high spatial densities. The proteins attached covalently to the slide surface yet retained their ability to interact specifically with other proteins, or with small molecules, in solution. Three applications for protein microarrays were demonstrated: screening for protein-protein interactions, identifying the substrates of protein kinases, and identifying the protein targets of small molecules.

Monolayer coated gold nanoparticles for delivery applications

PubMed Central

Rana, Subinoy; Bajaj, Avinash; Mout, Rubul; Rotello, Vincent M.

2011-01-01

Gold nanoparticles (AuNPs) provide attractive vehicles for delivery of drugs, genetic materials, proteins, and small molecules. AuNPs feature low core toxicity coupled with the ability to parametrically control particle size and surface properties. In this review, we focus on engineering of the AuNP surface monolayer, highlighting recent advances in tuning monolayer structures for efficient delivery of drugs and biomolecules. This review covers two broad categories of particle functionalization, organic monolayers and biomolecule coatings, and discusses their applications in drug, DNA/RNA, protein and small molecule delivery. PMID:21925556

Sequence-Specific Affinity Chromatography of Bacterial Small Regulatory RNA-Binding Proteins from Bacterial Cells.

PubMed

Gans, Jonathan; Osborne, Jonathan; Cheng, Juliet; Djapgne, Louise; Oglesby-Sherrouse, Amanda G

2018-01-01

Bacterial small RNA molecules (sRNAs) are increasingly recognized as central regulators of bacterial stress responses and pathogenesis. In many cases, RNA-binding proteins are critical for the stability and function of sRNAs. Previous studies have adopted strategies to genetically tag an sRNA of interest, allowing isolation of RNA-protein complexes from cells. Here we present a sequence-specific affinity purification protocol that requires no prior genetic manipulation of bacterial cells, allowing isolation of RNA-binding proteins bound to native RNA molecules.

The small molecule probe PT-Yellow labels the renal proximal tubules in zebrafish.

PubMed

Sander, Veronika; Patke, Shantanu; Sahu, Srikanta; Teoh, Chai Lean; Peng, Zhenzhen; Chang, Young-Tae; Davidson, Alan J

2015-01-01

We report the development of a small fluorescent molecule, BDNCA3-D2, herein referred to as PT-Yellow. Soaking zebrafish embryos in PT-Yellow or intraperitoneal injection into adults results in non-toxic in vivo fluorescent labeling of the renal proximal tubules, the major site of blood filtrate reabsorption and a common target of injury in acute kidney injury. We demonstrate the applicability of this new compound as a rapid and simple readout for zebrafish kidney filtration and proximal tubule reabsorption function.

Hydrogen sulphide in cardiovascular system: A cascade from interaction between sulphur atoms and signalling molecules.

PubMed

Wang, Ming-Jie; Cai, Wen-Jie; Zhu, Yi-Chun

2016-05-15

As a gasotransmitter, hydrogen sulphide exerts its extensive physiological and pathophysiological effects in mammals. The interaction between sulphur atoms and signalling molecules forms a cascade that modulates cellular functions and homeostasis. In this review, we focus on the signalling mechanism underlying the effect of hydrogen sulphide in the cardiovascular system and metabolism as well as the biological relevance to human diseases. Copyright © 2016 Elsevier Inc. All rights reserved.

Surface-supported Ag islands stabilized by a quantum size effect: Their interaction with small molecules relevant to ethylene epoxidation

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

Shao, Dahai

2013-05-15

This dissertation focuses on how QSE-stabilized, surface-supported Ag nanoclusters will interact with ethylene or oxygen. Experiments are performed to determine whether the QSE-mediated Ag islands react differently toward adsorption of ethylene or oxygen, or whether the adsorption of these small molecules will affect the QSE-mediated stability of Ag islands. Studies of the interaction of oxygen with Ag/Si(111)-7×7 were previously reported, but these studies were performed at a low Ag coverage where 3D Ag islands were not formed. So the study of such a system at a higher Ag coverage will be a subject of this work. The interaction of ethylenemore » with Ag/Si(111)-7×7, as well as the interaction of oxygen with Ag/NiAl(110) are also important parts of this study.« less

Signaling gateway molecule pages—a data model perspective

PubMed Central

Dinasarapu, Ashok Reddy; Saunders, Brian; Ozerlat, Iley; Azam, Kenan; Subramaniam, Shankar

2011-01-01

Summary: The Signaling Gateway Molecule Pages (SGMP) database provides highly structured data on proteins which exist in different functional states participating in signal transduction pathways. A molecule page starts with a state of a native protein, without any modification and/or interactions. New states are formed with every post-translational modification or interaction with one or more proteins, small molecules or class molecules and with each change in cellular location. State transitions are caused by a combination of one or more modifications, interactions and translocations which then might be associated with one or more biological processes. In a characterized biological state, a molecule can function as one of several entities or their combinations, including channel, receptor, enzyme, transcription factor and transporter. We have also exported SGMP data to the Biological Pathway Exchange (BioPAX) and Systems Biology Markup Language (SBML) as well as in our custom XML. Availability: SGMP is available at www.signaling-gateway.org/molecule. Contact: shankar@ucsd.edu Supplementary information: Supplementary data are available at Bioinformatics online. PMID:21505029

Hypoxia-inducible factor stabilizers and other small-molecule erythropoiesis-stimulating agents in current and preventive doping analysis.

PubMed

Beuck, Simon; Schänzer, Wilhelm; Thevis, Mario

2012-11-01

Increasing the blood's capacity for oxygen transport by erythropoiesis-stimulating agents (ESAs) constitutes a prohibited procedure of performance enhancement according to the World Anti-Doping Agency (WADA). The advent of orally bio-available small-molecule ESAs such as hypoxia-inducible factor (HIF) stabilizers in the development of novel anti-anaemia therapies expands the list of potential ESA doping techniques. Here, the erythropoiesis-stimulating properties and doping relevance of experimental HIF-stabilizers, such as cobaltous chloride, 3,4-dihydroxybenzoic acid or GSK360A, amongst others, are discussed. The stage of clinical trials is reviewed for the anti-anaemia drug candidates FG-2216, FG-4592, GSK1278863, AKB-6548, and BAY85-3934. Currently available methods and strategies for the determination of selected HIF stabilizers in sports drug testing are based on liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS). For the support of further analytical assay development, patents claiming distinct compounds for the use in HIF-mediated therapies are evaluated and exemplary molecular structures of HIF stabilizers presented. Moreover, data concerning the erythropoiesis-enhancing effects of the GATA inhibitors K7174 and K11706 as well as the lipidic small-molecule ESA PBI-1402 are elucidated the context of doping analysis. Copyright © 2012 John Wiley & Sons, Ltd.

Text Mining Improves Prediction of Protein Functional Sites

PubMed Central

Cohn, Judith D.; Ravikumar, Komandur E.

2012-01-01

We present an approach that integrates protein structure analysis and text mining for protein functional site prediction, called LEAP-FS (Literature Enhanced Automated Prediction of Functional Sites). The structure analysis was carried out using Dynamics Perturbation Analysis (DPA), which predicts functional sites at control points where interactions greatly perturb protein vibrations. The text mining extracts mentions of residues in the literature, and predicts that residues mentioned are functionally important. We assessed the significance of each of these methods by analyzing their performance in finding known functional sites (specifically, small-molecule binding sites and catalytic sites) in about 100,000 publicly available protein structures. The DPA predictions recapitulated many of the functional site annotations and preferentially recovered binding sites annotated as biologically relevant vs. those annotated as potentially spurious. The text-based predictions were also substantially supported by the functional site annotations: compared to other residues, residues mentioned in text were roughly six times more likely to be found in a functional site. The overlap of predictions with annotations improved when the text-based and structure-based methods agreed. Our analysis also yielded new high-quality predictions of many functional site residues that were not catalogued in the curated data sources we inspected. We conclude that both DPA and text mining independently provide valuable high-throughput protein functional site predictions, and that integrating the two methods using LEAP-FS further improves the quality of these predictions. PMID:22393388

Pharmacogenomic Biomarkers: an FDA Perspective on Utilization in Biological Product Labeling.

PubMed

Schuck, Robert N; Grillo, Joseph A

2016-05-01

Precision medicine promises to improve both the efficacy and safety of therapeutic products by better informing why some patients respond well to a drug, and some experience adverse reactions, while others do not. Pharmacogenomics is a key component of precision medicine and can be utilized to select optimal doses for patients, more precisely identify individuals who will respond to a treatment and avoid serious drug-related toxicities. Since pharmacogenomic biomarker information can help inform drug dosing, efficacy, and safety, pharmacogenomic data are critically reviewed by FDA staff to ensure effective use of pharmacogenomic strategies in drug development and appropriate incorporation into product labels. Pharmacogenomic information may be provided in drug or biological product labeling to inform health care providers about the impact of genotype on response to a drug through description of relevant genomic markers, functional effects of genomic variants, dosing recommendations based on genotype, and other applicable genomic information. The format and content of labeling for biologic drugs will generally follow that of small molecule drugs; however, there are notable differences in pharmacogenomic information that might be considered useful for biologic drugs in comparison to small molecule drugs. Furthermore, the rapid entry of biologic drugs for treatment of rare genetic diseases and molecularly defined subsets of common diseases will likely lead to increased use of pharmacogenomic information in biologic drug labels in the near future. In this review, we outline the general principles of therapeutic product labeling and discuss the utilization of pharmacogenomic information in biologic drug labels.

The fundamental unit of pain is the cell.

PubMed

Reichling, David B; Green, Paul G; Levine, Jon D

2013-12-01

The molecular/genetic era has seen the discovery of a staggering number of molecules implicated in pain mechanisms [18,35,61,69,96,133,150,202,224]. This has stimulated pharmaceutical and biotechnology companies to invest billions of dollars to develop drugs that enhance or inhibit the function of many these molecules. Unfortunately this effort has provided a remarkably small return on this investment. Inevitably, transformative progress in this field will require a better understanding of the functional links among the ever-growing ranks of "pain molecules," as well as their links with an even larger number of molecules with which they interact. Importantly, all of these molecules exist side-by-side, within a functional unit, the cell, and its adjacent matrix of extracellular molecules. To paraphrase a recent editorial in Science magazine [223], although we live in the Golden age of Genetics, the fundamental unit of biology is still arguably the cell, and the cell is the critical structural and functional setting in which the function of pain-related molecules must be understood. This review summarizes our current understanding of the nociceptor as a cell-biological unit that responds to a variety of extracellular inputs with a complex and highly organized interaction of signaling molecules. We also discuss the insights that this approach is providing into peripheral mechanisms of chronic pain and sex dependence in pain. Copyright © 2013 International Association for the Study of Pain. Published by Elsevier B.V. All rights reserved.

In vitro selection of shape-changing DNA nanostructures capable of binding-induced cargo release.

PubMed

Oh, Seung Soo; Plakos, Kory; Xiao, Yi; Eisenstein, Michael; Soh, H Tom

2013-11-26

Many biological systems employ allosteric regulatory mechanisms, which offer a powerful means of directly linking a specific binding event to a wide spectrum of molecular functionalities. There is considerable interest in generating synthetic allosteric regulators that can perform useful molecular functions for applications in diagnostics, imaging and targeted therapies, but generating such molecules through either rational design or directed evolution has proven exceptionally challenging. To address this need, we present an in vitro selection strategy for generating conformation-switching DNA nanostructures that selectively release a small-molecule payload in response to binding of a specific trigger molecule. As an exemplar, we have generated a DNA nanostructure that hybridizes with a separate 'cargo strand' containing an abasic site. This abasic site stably sequesters a fluorescent cargo molecule in an inactive state until the DNA nanostructure encounters an ATP trigger molecule. This ATP trigger causes the nanostructure to release the cargo strand, thereby liberating the fluorescent payload and generating a detectable fluorescent readout. Our DNA nanostructure is highly sensitive, with an EC50 of 30 μM, and highly specific, releasing its payload in response to ATP but not to other chemically similar nucleotide triphosphates. We believe that this selection approach could be generalized to generate synthetic nanostructures capable of selective and controlled release of other small-molecule cargos in response to a variety of triggers, for both research and clinical applications.

Prion-like nanofibrils of small molecules (PriSM): A new frontier at the intersection of supramolecular chemistry and cell biology.

PubMed

Zhou, Jie; Du, Xuewen; Xu, Bing

2015-01-01

Formed by non-covalent interactions and not defined at genetic level, the assemblies of small molecules in biology are complicated and less explored. A common morphology of the supramolecular assemblies of small molecules is nanofibrils, which coincidentally resembles the nanofibrils formed by proteins such as prions. So these supramolecular assemblies are termed as prion-like nanofibrils of small molecules (PriSM). Emerging evidence from several unrelated fields over the past decade implies the significance of PriSM in biology and medicine. This perspective aims to highlight some recent advances of the research on PriSM. This paper starts with description of the intriguing similarities between PriSM and prions, discusses the paradoxical features of PriSM, introduces the methods for elucidating the biological functions of PriSM, illustrates several examples of beneficial aspects of PriSM, and finishes with the promises and current challenges in the research of PriSM. We anticipate that the research of PriSM will contribute to the fundamental understanding at the intersection of supramolecular chemistry and cell biology and ultimately lead to a new paradigm of molecular (or supramolecular) therapeutics for biomedicine.

Prion-like nanofibrils of small molecules (PriSM): A new frontier at the intersection of supramolecular chemistry and cell biology

PubMed Central

Zhou, Jie; Du, Xuewen; Xu, Bing

2015-01-01

Abstract Formed by non-covalent interactions and not defined at genetic level, the assemblies of small molecules in biology are complicated and less explored. A common morphology of the supramolecular assemblies of small molecules is nanofibrils, which coincidentally resembles the nanofibrils formed by proteins such as prions. So these supramolecular assemblies are termed as prion-like nanofibrils of small molecules (PriSM). Emerging evidence from several unrelated fields over the past decade implies the significance of PriSM in biology and medicine. This perspective aims to highlight some recent advances of the research on PriSM. This paper starts with description of the intriguing similarities between PriSM and prions, discusses the paradoxical features of PriSM, introduces the methods for elucidating the biological functions of PriSM, illustrates several examples of beneficial aspects of PriSM, and finishes with the promises and current challenges in the research of PriSM. We anticipate that the research of PriSM will contribute to the fundamental understanding at the intersection of supramolecular chemistry and cell biology and ultimately lead to a new paradigm of molecular (or supramolecular) therapeutics for biomedicine. PMID:25738892

Novel dual small-molecule HIV inhibitors: scaffolds and discovery strategies.

PubMed

Song, Anran; Yu, Haiqing; Wang, Changyuan; Zhu, Xingqi; Liu, Kexin; Ma, Xiaodong

2015-01-01

Searching for safe and effective treatments for HIV infection is still a great challenge worldwide in spite of the 27 marketed anti-HIV drugs and the powerful highly active antiretroviral therapy (HAART). As a promising prospect for generation of new HIV therapy drugs, multiple ligands (MDLs) were greatly focused on recently due to their lower toxicity, simplified dosing and patient adherence than single-target drugs. Till now, by disrupting two active sites or steps of HIV replications, a number of HIV dual inhibitors, such as CD4-gssucap120 inhibitors, CXCR4-gp20 inhibitors, RT-CXCR4 inhibitors, RT-protease inhibitors, RT-integrase inhibitors, and RTassociated functions inhibitors have been identified. Generally, these dual inhibitors were discovered mainly through screening approaches and design strategies. Of these compounds, the molecules bearing small skeletons exhibited strong anti-HIV activity and aroused great attention recently. Reviewing the progress of the dual small-molecule HIV inhibitors from the point of view of their scaffolds and discovery strategies will provide valuable information for producing more effective anti-HIV drugs. In this regard, novel dual small-molecule HIV inhibitors were illustrated, and their discovery paradigms as the major contents were also summarized in this manuscript.

Pre-clinical evaluation of small molecule LOXL2 inhibitors in breast cancer

PubMed Central

Chang, Joan; Lucas, Morghan C.; Leonte, Lidia E.; Garcia-Montolio, Marc; Singh, Lukram Babloo; Findlay, Alison D.; Deodhar, Mandar; Foot, Jonathan S.; Jarolimek, Wolfgang; Timpson, Paul; Erler, Janine T.; Cox, Thomas R.

2017-01-01

Lysyl Oxidase-like 2 (LOXL2), a member of the lysyl oxidase family of amine oxidases is known to be important in normal tissue development and homeostasis, as well as the onset and progression of solid tumors. Here we tested the anti-tumor properties of two generations of novel small molecule LOXL2 inhibitor in the MDA-MB-231 human model of breast cancer. We confirmed a functional role for LOXL2 activity in the progression of primary breast cancer. Inhibition of LOXL2 activity inhibited the growth of primary tumors and reduced primary tumor angiogenesis. Dual inhibition of LOXL2 and LOX showed a greater effect and also led to a lower overall metastatic burden in the lung and liver. Our data provides the first evidence to support a role for LOXL2 specific small molecule inhibitors as a potential therapy in breast cancer. PMID:28199967

Remote control of therapeutic T cells through a small molecule-gated chimeric receptor

PubMed Central

Wu, Chia-Yung; Roybal, Kole T.; Puchner, Elias M.; Onuffer, James; Lim, Wendell A.

2016-01-01

There is growing promise in using engineered cells as therapeutic agents. For example, synthetic Chimeric Antigen Receptors (CARs) can redirect T cells to recognize and eliminate tumor cells expressing specific antigens. Despite promising clinical results, excessive activity and poor control over such engineered T cells can cause severe toxicities. We present the design of “ON-switch” CARs that enable small molecule-control over T cell therapeutic functions, while still retaining antigen specificity. In these split receptors, antigen binding and intracellular signaling components only assemble in the presence of a heterodimerizing small molecule. This titratable pharmacologic regulation could allow physicians to precisely control the timing, location, and dosage of T cell activity, thereby mitigating toxicity. This work illustrates the potential of combining cellular engineering with orthogonal chemical tools to yield safer therapeutic cells that tightly integrate both cell autonomous recognition and user control. PMID:26405231

Remote control of therapeutic T cells through a small molecule-gated chimeric receptor.

PubMed

Wu, Chia-Yung; Roybal, Kole T; Puchner, Elias M; Onuffer, James; Lim, Wendell A

2015-10-16

There is growing interest in using engineered cells as therapeutic agents. For example, synthetic chimeric antigen receptors (CARs) can redirect T cells to recognize and eliminate tumor cells expressing specific antigens. Despite promising clinical results, these engineered T cells can exhibit excessive activity that is difficult to control and can cause severe toxicity. We designed "ON-switch" CARs that enable small-molecule control over T cell therapeutic functions while still retaining antigen specificity. In these split receptors, antigen-binding and intracellular signaling components assemble only in the presence of a heterodimerizing small molecule. This titratable pharmacologic regulation could allow physicians to precisely control the timing, location, and dosage of T cell activity, thereby mitigating toxicity. This work illustrates the potential of combining cellular engineering with orthogonal chemical tools to yield safer therapeutic cells that tightly integrate cell-autonomous recognition and user control. Copyright © 2015, American Association for the Advancement of Science.

Diamond nanowires for highly sensitive matrix-free mass spectrometry analysis of small molecules.

PubMed

Coffinier, Yannick; Szunerits, Sabine; Drobecq, Hervé; Melnyk, Oleg; Boukherroub, Rabah

2012-01-07

This paper reports on the use of boron-doped diamond nanowires (BDD NWs) as an inorganic substrate for matrix-free laser desorption/ionization mass spectrometry (LDI-MS) analysis of small molecules. The diamond nanowires are prepared by reactive ion etching (RIE) with oxygen plasma of highly boron-doped (the boron level is 10(19) B cm(-3)) or undoped nanocrystalline diamond substrates. The resulting diamond nanowires are coated with a thin silicon oxide layer that confers a superhydrophilic character to the surface. To minimize droplet spreading, the nanowires were chemically functionalized with octadecyltrichlorosilane (OTS) and then UV/ozone treated to reach a final water contact angle of 120°. The sub-bandgap absorption under UV laser irradiation and the heat confinement inside the nanowires allowed desorption/ionization, most likely via a thermal mechanism, and mass spectrometry analysis of small molecules. A detection limit of 200 zeptomole for verapamil was demonstrated.

A small-molecule switch for Golgi sulfotransferases.

PubMed

de Graffenried, Christopher L; Laughlin, Scott T; Kohler, Jennifer J; Bertozzi, Carolyn R

2004-11-30

The study of glycan function is a major frontier in biology that could benefit from small molecules capable of perturbing carbohydrate structures on cells. The widespread role of sulfotransferases in modulating glycan function makes them prime targets for small-molecule modulators. Here, we report a system for conditional activation of Golgi-resident sulfotransferases using a chemical inducer of dimerization. Our approach capitalizes on two features shared by these enzymes: their requirement of Golgi localization for activity on cellular substrates and the modularity of their catalytic and localization domains. Fusion of these domains to the proteins FRB and FKBP enabled their induced assembly by the natural product rapamycin. We applied this strategy to the GlcNAc-6-sulfotransferases GlcNAc6ST-1 and GlcNAc6ST-2, which collaborate in the sulfation of L-selectin ligands. Both the activity and specificity of the inducible enzymes were indistinguishable from their WT counterparts. We further generated rapamycin-inducible chimeric enzymes comprising the localization domain of a sulfotransferase and the catalytic domain of a glycosyltransferase, demonstrating the generality of the system among other Golgi enzymes. The approach provides a means for studying sulfate-dependent processes in cellular systems and, potentially, in vivo.

BET Acetyl-Lysine Binding Proteins Control Pathological Cardiac Hypertrophy

PubMed Central

Spiltoir, Jessica I.; Stratton, Matthew S.; Cavasin, Maria A.; Demos-Davies, Kim; Reid, Brian G.; Qi, Jun; Bradner, James E.; McKinsey, Timothy A.

2014-01-01

Cardiac hypertrophy is an independent predictor of adverse outcomes in patients with heart failure, and thus represents an attractive target for novel therapeutic intervention. JQ1, a small molecule inhibitor of bromodomain and extraterminal (BET) acetyl-lysine reader proteins, was identified in a high throughput screen designed to discover novel small molecule regulators of cardiomyocyte hypertrophy. JQ1 dose-dependently blocked agonist-dependent hypertrophy of cultured neonatal rat ventricular myocytes (NRVMs) and reversed the prototypical gene program associated with pathological cardiac hypertrophy. JQ1 also blocked left ventricular hypertrophy (LVH) and improved cardiac function in adult mice subjected to transverse aortic constriction (TAC). The BET family consists of BRD2, BRD3, BRD4 and BRDT. BRD4 protein expression was increased during cardiac hypertrophy, and hypertrophic stimuli promoted recruitment of BRD4 to the transcriptional start site (TSS) of the gene encoding atrial natriuretic factor (ANF). Binding of BRD4 to the ANF TSS was associated with increased phosphorylation of local RNA polymerase II. These findings define a novel function for BET proteins as signal-responsive regulators of cardiac hypertrophy, and suggest that small molecule inhibitors of these epigenetic reader proteins have potential as therapeutics for heart failure. PMID:23939492

Rescue of p53 function by small-molecule RITA in cervical carcinoma by blocking E6-mediated degradation.

PubMed

Zhao, Carolyn Ying; Szekely, Laszlo; Bao, Wenjie; Selivanova, Galina

2010-04-15

Proteasomal degradation of p53 by human papilloma virus (HPV) E6 oncoprotein plays a pivotal role in the survival of cervical carcinoma cells. Abrogation of HPV-E6-dependent p53 destruction can therefore be a good strategy to combat cervical carcinomas. Here, we show that a small-molecule reactivation of p53 and induction of tumor cell apoptosis (RITA) is able to induce the accumulation of p53 and rescue its tumor suppressor function in cells containing high-risk HPV16 and HPV18 by inhibiting HPV-E6-mediated proteasomal degradation. RITA blocks p53 ubiquitination by preventing p53 interaction with E6-associated protein, required for HPV-E6-mediated degradation. RITA activates the transcription of proapoptotic p53 targets Noxa, PUMA, and BAX, and repressed the expression of pro-proliferative factors CyclinB1, CDC2, and CDC25C, resulting in p53-dependent apoptosis and cell cycle arrest. Importantly, RITA showed substantial suppression of cervical carcinoma xenografts in vivo. These results provide a proof of principle for the treatment of cervical cancer in a p53-dependent manner by using small molecules that target p53. (c)2010 AACR.

Efficacy of a Mer and Flt3 tyrosine kinase small molecule inhibitor, UNC1666, in acute myeloid leukemia

PubMed Central

Lee-Sherick, Alisa B.; Zhang, Weihe; Menachof, Kelly K.; Hill, Amanda A.; Rinella, Sean; Kirkpatrick, Gregory; Page, Lauren S.; Stashko, Michael A.; Jordan, Craig T.; Wei, Qi; Liu, Jing; Zhang, Dehui; DeRyckere, Deborah; Wang, Xiaodong; Frye, Stephen; Earp, H. Shelton; Graham, Douglas K.

2015-01-01

Mer and Flt3 receptor tyrosine kinases have been implicated as therapeutic targets in acute myeloid leukemia (AML). In this manuscript we describe UNC1666, a novel ATP-competitive small molecule tyrosine kinase inhibitor, which potently diminishes Mer and Flt3 phosphorylation in AML. Treatment with UNC1666 mediated biochemical and functional effects in AML cell lines expressing Mer or Flt3 internal tandem duplication (ITD), including decreased phosphorylation of Mer, Flt3 and downstream effectors Stat, Akt and Erk, induction of apoptosis in up to 98% of cells, and reduction of colony formation by greater than 90%, compared to treatment with vehicle. These effects were dose-dependent, with inhibition of downstream signaling and functional effects correlating with the degree of Mer or Flt3 kinase inhibition. Treatment of primary AML patient samples expressing Mer and/or Flt3-ITD with UNC1666 also inhibited Mer and Flt3 intracellular signaling, induced apoptosis, and inhibited colony formation. In summary, UNC1666 is a novel potent small molecule tyrosine kinase inhibitor that decreases oncogenic signaling and myeloblast survival, thereby validating dual Mer/Flt3 inhibition as an attractive treatment strategy for AML. PMID:25762638

Application of reference-modified density functional theory: Temperature and pressure dependences of solvation free energy.

PubMed

Sumi, Tomonari; Maruyama, Yutaka; Mitsutake, Ayori; Mochizuki, Kenji; Koga, Kenichiro

2018-02-05

Recently, we proposed a reference-modified density functional theory (RMDFT) to calculate solvation free energy (SFE), in which a hard-sphere fluid was introduced as the reference system instead of an ideal molecular gas. Through the RMDFT, using an optimal diameter for the hard-sphere reference system, the values of the SFE calculated at room temperature and normal pressure were in good agreement with those for more than 500 small organic molecules in water as determined by experiments. In this study, we present an application of the RMDFT for calculating the temperature and pressure dependences of the SFE for solute molecules in water. We demonstrate that the RMDFT has high predictive ability for the temperature and pressure dependences of the SFE for small solute molecules in water when the optimal reference hard-sphere diameter determined for each thermodynamic condition is used. We also apply the RMDFT to investigate the temperature and pressure dependences of the thermodynamic stability of an artificial small protein, chignolin, and discuss the mechanism of high-temperature and high-pressure unfolding of the protein. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Meta-analysis of global metabolomics and proteomics data to link alterations with phenotype

DOE PAGES

Patti, Gary J.; Tautenhahn, Ralf; Fonslow, Bryan R.; ...

2011-01-01

Global metabolomics has emerged as a powerful tool to interrogate cellular biochemistry at the systems level by tracking alterations in the levels of small molecules. One approach to define cellular dynamics with respect to this dysregulation of small molecules has been to consider metabolic flux as a function of time. While flux measurements have proven effective for model organisms, acquiring multiple time points at appropriate temporal intervals for many sample types (e.g., clinical specimens) is challenging. As an alternative, meta-analysis provides another strategy for delineating metabolic cause and effect perturbations. That is, the combination of untargeted metabolomic data from multiplemore » pairwise comparisons enables the association of specific changes in small molecules with unique phenotypic alterations. We recently developed metabolomic software called metaXCMS to automate these types of higher order comparisons. Here we discuss the potential of metaXCMS for analyzing proteomic datasets and highlight the biological value of combining meta-results from both metabolomic and proteomic analyses. The combined meta-analysis has the potential to facilitate efforts in functional genomics and the identification of metabolic disruptions related to disease pathogenesis.« less

Small molecule inhibition of fibroblast growth factor receptors in cancer.

PubMed

Liang, Guang; Chen, Gaozhi; Wei, Xiaoyan; Zhao, Yunjie; Li, Xiaokun

2013-10-01

Fibroblast growth factors (FGFs) signal through FGF receptors (FGFRs), which are a sub-family of the superfamily of receptor tyrosine kinases, to regulate human development and metabolism. Uncontrolled FGF signaling is responsible for diverse array of developmental disorders, most notably skeletal syndromes due to FGFR gain-of-function mutations. Studies in the last few years have provided significant evidence for the importance of FGF signaling in the pathogenesis of diverse cancers, including endometrial and bladder cancers. FGFs are both potent mitogenic and angiogenic factors and can contribute to carcinogenesis by stimulating cell proliferation and tumor angiogenesis. Gene knockout and pharmacological inhibition of FGFRs in in vivo and in vitro models validate FGFRs as a target for cancer treatment. Considerable efforts are being expended to develop specific, small-molecule inhibitors for treating FGFR-driven cancers. Recent reviews on the FGF/FGFR system have focused primarily on signaling, pathophysiology, and functions in cancer. In this article, we review the key roles of FGFR in cancer, provide an update on the status of clinical trials with small-molecule FGFR inhibitors, and discuss how the current structural data on FGFR kinases guide the design and characterization of new FGFR inhibitors. Copyright © 2013 Elsevier Ltd. All rights reserved.

Array Formatting of the Heat-Transfer Method (HTM) for the Detection of Small Organic Molecules by Molecularly Imprinted Polymers

PubMed Central

Wackers, Gideon; Vandenryt, Thijs; Cornelis, Peter; Kellens, Evelien; Thoelen, Ronald; De Ceuninck, Ward; Losada-Pérez, Patricia; van Grinsven, Bart; Peeters, Marloes; Wagner, Patrick

2014-01-01

In this work we present the first steps towards a molecularly imprinted polymer (MIP)-based biomimetic sensor array for the detection of small organic molecules via the heat-transfer method (HTM). HTM relies on the change in thermal resistance upon binding of the target molecule to the MIP-type receptor. A flow-through sensor cell was developed, which is segmented into four quadrants with a volume of 2.5 μL each, allowing four measurements to be done simultaneously on a single substrate. Verification measurements were conducted, in which all quadrants received a uniform treatment and all four channels exhibited a similar response. Subsequently, measurements were performed in quadrants, which were functionalized with different MIP particles. Each of these quadrants was exposed to the same buffer solution, spiked with different molecules, according to the MIP under analysis. With the flow cell design we could discriminate between similar small organic molecules and observed no significant cross-selectivity. Therefore, the MIP array sensor platform with HTM as a readout technique, has the potential to become a low-cost analysis tool for bioanalytical applications. PMID:24955945

Adsorption of H2O, H2, O2, CO, NO, and CO2 on graphene/g-C3N4 nanocomposite investigated by density functional theory

NASA Astrophysics Data System (ADS)

Wu, Hong-Zhang; Bandaru, Sateesh; Liu, Jin; Li, Li-Li; Wang, Zhenling

2018-02-01

Motivated by the photocatalytic reactions of small molecules on g-C3N4 by these insights, we sought to explore the adsorption of H2O and CO2 molecules on the graphene side and H2O, H2, O2, CO, NO, and CO2 molecules on the g-C3N4 side of hybrid g-C3N4/graphene nanocomposite using first-principles calculations. The atomic structure and electronic properties of hybrid g-C3N4/graphene nanocomposite is explored. The adsorption of small molecules on graphene/g-C3N4 nanocomposite is thoroughly investigated. The computational studies revels that all small molecules on graphene/g-C3N4 nanocomposite are the physisorption. The adsorption characteristics of H2O and CO2 molecules on the graphene side are similar to that on graphene. The adsorption of H2O, H2, O2, CO, NO, and CO2 molecules on the g-C3N4 side always leads to a buckle structure of graphene/g-C3N4 nanocomposite. Graphene as a substrate can significantly relax the buckle degree of g-C3N4 in g-C3N4/graphene nanocomposite.

A toxic RNA catalyzes the in cellulo synthesis of its own inhibitor.

PubMed

Rzuczek, Suzanne G; Park, HaJeung; Disney, Matthew D

2014-10-06

Potent modulators of RNA function can be assembled in cellulo by using the cell as a reaction vessel and a disease-causing RNA as a catalyst. When designing small molecule effectors of function, a balance between permeability and potency must be struck. Low molecular weight compounds are more permeable whereas higher molecular weight compounds are more potent. The advantages of both types of compounds could be synergized if low molecular weight molecules could be transformed into potent, multivalent ligands by a reaction that is catalyzed by binding to a target in cells expressing a genetic defect. It was shown that this approach is indeed viable in cellulo. Small molecule modules with precisely positioned alkyne and azide moieties bind adjacent internal loops in r(CCUG)(exp), the causative agent of myotonic dystrophy type 2 (DM2), and are transformed into oligomeric, potent inhibitors of DM2 RNA dysfunction by a Huisgen 1,3-dipolar cycloaddition reaction, a variant of click chemistry. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Adsorption of CO and O2 molecules on Li metal adsorbed graphene: Search for graphene based gas sensors

NASA Astrophysics Data System (ADS)

Kaur, Gagandeep; Gupta, Shuchi; Sachdeva, Ritika; Dharamvir, Keya

2018-05-01

Adsorption of small gas molecules (such as CO and O2) on pristine graphene (PG) and Li-adsorbed graphene (PG-Li) have been investigated using first principles methods within density functional theory (DFT). We also notice that PG-Li has a higher chemical reactivity towards the gas molecules as compared to PG and these molecules have higher adsorption energy on this surface. Moreover, the strong interactions between PG-Li and the adsorbed molecules (as compared to PG and gas molecules) induce dramatic changes to the electronic properties of PG adsorbed with Li and make PG-Li a promising candidate as sensing material for CO and O2 gases.

Studying a Drug-like, RNA-Focused Small Molecule Library Identifies Compounds That Inhibit RNA Toxicity in Myotonic Dystrophy.

PubMed

Rzuczek, Suzanne G; Southern, Mark R; Disney, Matthew D

2015-12-18

There are many RNA targets in the transcriptome to which small molecule chemical probes and lead therapeutics are desired. However, identifying compounds that bind and modulate RNA function in cellulo is difficult. Although rational design approaches have been developed, they are still in their infancies and leave many RNAs "undruggable". In an effort to develop a small molecule library that is biased for binding RNA, we computationally identified "drug-like" compounds from screening collections that have favorable properties for binding RNA and for suitability as lead drugs. As proof-of-concept, this collection was screened for binding to and modulating the cellular dysfunction of the expanded repeating RNA (r(CUG)(exp)) that causes myotonic dystrophy type 1. Hit compounds bind the target in cellulo, as determined by the target identification approach Competitive Chemical Cross-Linking and Isolation by Pull-down (C-ChemCLIP), and selectively improve several disease-associated defects. The best compounds identified from our 320-member library are more potent in cellulo than compounds identified by high-throughput screening (HTS) campaigns against this RNA. Furthermore, the compound collection has a higher hit rate (9% compared to 0.01-3%), and the bioactive compounds identified are not charged; thus, RNA can be "drugged" with compounds that have favorable pharmacological properties. Finally, this RNA-focused small molecule library may serve as a useful starting point to identify lead "drug-like" chemical probes that affect the biological (dys)function of other RNA targets by direct target engagement.

Label-free electrochemical biosensing of small-molecule inhibition on O-GlcNAc glycosylation.

PubMed

Yang, Yu; Gu, Yuxin; Wan, Bin; Ren, Xiaomin; Guo, Liang-Hong

2017-09-15

O-linked N-acetylglucosamine (O-GlcNAc) transferase (OGT) plays a critical role in modulating protein function in many cellular processes and human diseases such as Alzheimer's disease and type II diabetes, and has emerged as a promising new target. Specific inhibitors of OGT could be valuable tools to probe the biological functions of O-GlcNAcylation, but a lack of robust nonradiometric assay strategies to detect glycosylation, has impeded efforts to identify such compounds. Here we have developed a novel label-free electrochemical biosensor for the detection of peptide O-GlcNAcylation using protease-protection strategy and electrocatalytic oxidation of tyrosine mediated by osmium bipyridine as a signal reporter. There is a large difference in the abilities of proteolysis of the glycosylated and the unglycosylated peptides by protease, thus providing a sensing mechanism for OGT activity. When the O-GlcNAcylation is achieved, the glycosylated peptides cannot be cleaved by proteinase K and result in a high current response on indium tin oxide (ITO) electrode. However, when the O-GlcNAcylation is successfully inhibited using a small molecule, the unglycosylated peptides can be cleaved easily and lead to low current signal. Peptide O-GlcNAcylation reaction was performed in the presence of a well-defined small-molecule OGT inhibitor. The results indicated that the biosensor could be used to screen the OGT inhibitors effectively. Our label-free electrochemical method is a promising candidate for protein glycosylation pathway research in screening small-molecule inhibitors of OGT. Copyright © 2017 Elsevier B.V. All rights reserved.

Carbohydrate Recognition by Boronolectins, Small Molecules, and Lectins

PubMed Central

Jin, Shan; Cheng, Yunfeng; Reid, Suazette; Li, Minyong; Wang, Binghe

2009-01-01

Carbohydrates are known to mediate a large number of biological and pathological events. Small and macromolecules capable of carbohydrate recognition have great potentials as research tools, diagnostics, vectors for targeted delivery of therapeutic and imaging agents, and therapeutic agents. However, this potential is far from being realized. One key issue is the difficulty in the development of “binders” capable of specific recognition of carbohydrates of biological relevance. This review discusses systematically the general approaches that are available in developing carbohydrate sensors and “binders/receptors,” and their applications. The focus is on discoveries during the last five years. PMID:19291708

Rational Design of Small Molecules Targeting Oncogenic Noncoding RNAs from Sequence.

PubMed

Disney, Matthew D; Angelbello, Alicia J

2016-12-20

The discovery of RNA catalysis in the 1980s and the dissemination of the human genome sequence at the start of this century inspired investigations of the regulatory roles of noncoding RNAs in biology. In fact, the Encyclopedia of DNA Elements (ENCODE) project has shown that only 1-2% of the human genome encodes protein, yet 75% is transcribed into RNA. Functional studies both preceding and following the ENCODE project have shown that these noncoding RNAs have important roles in regulating gene expression, developmental timing, and other critical functions. RNA's diverse roles are often a consequence of the various folds that it adopts. The single-stranded nature of the biopolymer enables it to adopt intramolecular folds with noncanonical pairings to lower its free energy. These folds can be scaffolds to bind proteins or to form frameworks to interact with other RNAs. Not surprisingly, dysregulation of certain noncoding RNAs has been shown to be causative of disease. Given this as the background, it is easy to see why it would be useful to develop methods that target RNA and manipulate its biology in rational and predictable ways. The antisense approach has afforded strategies to target RNAs via Watson-Crick base pairing and has typically focused on targeting partially unstructured regions of RNA. Small molecule strategies to target RNA would be desirable not only because compounds could be lead optimized via medicinal chemistry but also because structured regions within an RNA of interest could be targeted to directly interfere with RNA folds that contribute to disease. Additionally, small molecules have historically been the most successful drug candidates. Until recently, the ability to design small molecules that target non-ribosomal RNAs has been elusive, creating the perception that they are "undruggable". In this Account, approaches to demystify targeting RNA with small molecules are described. Rather than bulk screening for compounds that bind to singular targets, which is the purview of the pharmaceutical industry and academic institutions with high throughput screening facilities, we focus on methods that allow for the rational design of small molecules toward biological RNAs. One enabling and foundational technology that has been developed is two-dimensional combinatorial screening (2DCS), a library-versus-library selection approach that allows the identification of the RNA motif binding preferences of small molecules from millions of combinations. A landscape map of the 2DCS-defined and annotated RNA motif-small molecule interactions is then placed into Inforna, a computational tool that allows one to mine these interactions against an RNA of interest or an entire transcriptome. Indeed, this approach has been enabled by tools to annotate RNA structure from sequence, an invaluable asset to the RNA community and this work, and has allowed for the rational identification of "druggable" RNAs in a target agnostic fashion.

Recognition Tunneling

PubMed Central

Lindsay, Stuart; He, Jin; Sankey, Otto; Hapala, Prokop; Jelinek, Pavel; Zhang, Peiming; Chang, Shuai; Huang, Shuo

2010-01-01

Single molecules in a tunnel junction can now be interrogated reliably using chemically-functionalized electrodes. Monitoring stochastic bonding fluctuations between a ligand bound to one electrode and its target bound to a second electrode (“tethered molecule-pair” configuration) gives insight into the nature of the intermolecular bonding at a single molecule-pair level, and defines the requirements for reproducible tunneling data. Simulations show that there is an instability in the tunnel gap at large currents, and this results in a multiplicity of contacts with a corresponding spread in the measured currents. At small currents (i.e. large gaps) the gap is stable, and functionalizing a pair of electrodes with recognition reagents (the “free analyte” configuration) can generate a distinct tunneling signal when an analyte molecule is trapped in the gap. This opens up a new interface between chemistry and electronics with immediate implications for rapid sequencing of single DNA molecules. PMID:20522930

Identification of DNA primase inhibitors via a combined fragment-based and virtual screening

NASA Astrophysics Data System (ADS)

Ilic, Stefan; Akabayov, Sabine R.; Arthanari, Haribabu; Wagner, Gerhard; Richardson, Charles C.; Akabayov, Barak

2016-11-01

The structural differences between bacterial and human primases render the former an excellent target for drug design. Here we describe a technique for selecting small molecule inhibitors of the activity of T7 DNA primase, an ideal model for bacterial primases due to their common structural and functional features. Using NMR screening, fragment molecules that bind T7 primase were identified and then exploited in virtual filtration to select larger molecules from the ZINC database. The molecules were docked to the primase active site using the available primase crystal structure and ranked based on their predicted binding energies to identify the best candidates for functional and structural investigations. Biochemical assays revealed that some of the molecules inhibit T7 primase-dependent DNA replication. The binding mechanism was delineated via NMR spectroscopy. Our approach, which combines fragment based and virtual screening, is rapid and cost effective and can be applied to other targets.

Autophagic compound database: A resource connecting autophagy-modulating compounds, their potential targets and relevant diseases.

PubMed

Deng, Yiqi; Zhu, Lingjuan; Cai, Haoyang; Wang, Guan; Liu, Bo

2018-06-01

Autophagy, a highly conserved lysosomal degradation process in eukaryotic cells, can digest long-lived proteins and damaged organelles through vesicular trafficking pathways. Nowadays, mechanisms of autophagy have been gradually elucidated and thus the discovery of small-molecule drugs targeting autophagy has always been drawing much attention. So far, some autophagy-related web servers have been available online to facilitate scientists to obtain the information relevant to autophagy conveniently, such as HADb, CTLPScanner, iLIR server and ncRDeathDB. However, to the best of our knowledge, there is not any web server available about the autophagy-modulating compounds. According to published articles, all the compounds and their relations with autophagy were anatomized. Subsequently, an online Autophagic Compound Database (ACDB) (http://www.acdbliulab.com/) was constructed, which contained information of 357 compounds with 164 corresponding signalling pathways and potential targets in different diseases. We achieved a great deal of information of autophagy-modulating compounds, including compounds, targets/pathways and diseases. ACDB is a valuable resource for users to access to more than 300 curated small-molecule compounds correlated with autophagy. Autophagic compound database will facilitate to the discovery of more novel therapeutic drugs in the near future. © 2017 John Wiley & Sons Ltd.

Controlling release from 3D printed medical devices using CLIP and drug-loaded liquid resins.

PubMed

Bloomquist, Cameron J; Mecham, Michael B; Paradzinsky, Mark D; Janusziewicz, Rima; Warner, Samuel B; Luft, J Christopher; Mecham, Sue J; Wang, Andrew Z; DeSimone, Joseph M

2018-05-28

Mass customization along with the ability to generate designs using medical imaging data makes 3D printing an attractive method for the fabrication of patient-tailored drug and medical devices. Herein we describe the application of Continuous Liquid Interface Production (CLIP) as a method to fabricate biocompatible and drug-loaded devices with controlled release properties, using liquid resins containing active pharmaceutical ingredients (API). In this work, we characterize how the release kinetics of a model small molecule, rhodamine B-base (RhB), are affected by device geometry, network crosslink density, and the polymer composition of polycaprolactone- and poly (ethylene glycol)-based networks. To demonstrate the applicability of using API-loaded liquid resins with CLIP, the UV stability was evaluated for a panel of clinically-relevant small molecule drugs. Finally, select formulations were tested for biocompatibility, degradation and encapsulation of docetaxel (DTXL) and dexamethasone-acetate (DexAc). Formulations were shown to be biocompatible over the course of 175 days of in vitro degradation and the clinically-relevant drugs could be encapsulated and released in a controlled fashion. This study reveals the potential of the CLIP manufacturing platform to serve as a method for the fabrication of patient-specific medical and drug-delivery devices for personalized medicine. Copyright © 2018. Published by Elsevier B.V.

SInCRe—structural interactome computational resource for Mycobacterium tuberculosis

PubMed Central

Metri, Rahul; Hariharaputran, Sridhar; Ramakrishnan, Gayatri; Anand, Praveen; Raghavender, Upadhyayula S.; Ochoa-Montaño, Bernardo; Higueruelo, Alicia P.; Sowdhamini, Ramanathan; Chandra, Nagasuma R.; Blundell, Tom L.; Srinivasan, Narayanaswamy

2015-01-01

We have developed an integrated database for Mycobacterium tuberculosis H37Rv (Mtb) that collates information on protein sequences, domain assignments, functional annotation and 3D structural information along with protein–protein and protein–small molecule interactions. SInCRe (Structural Interactome Computational Resource) is developed out of CamBan (Cambridge and Bangalore) collaboration. The motivation for development of this database is to provide an integrated platform to allow easily access and interpretation of data and results obtained by all the groups in CamBan in the field of Mtb informatics. In-house algorithms and databases developed independently by various academic groups in CamBan are used to generate Mtb-specific datasets and are integrated in this database to provide a structural dimension to studies on tuberculosis. The SInCRe database readily provides information on identification of functional domains, genome-scale modelling of structures of Mtb proteins and characterization of the small-molecule binding sites within Mtb. The resource also provides structure-based function annotation, information on small-molecule binders including FDA (Food and Drug Administration)-approved drugs, protein–protein interactions (PPIs) and natural compounds that bind to pathogen proteins potentially and result in weakening or elimination of host–pathogen protein–protein interactions. Together they provide prerequisites for identification of off-target binding. Database URL: http://proline.biochem.iisc.ernet.in/sincre PMID:26130660

Butter feeding enhances TNF-alpha production from macrophages and lymphocyte adherence in murine small intestinal microvessels.

PubMed

Fujiyama, Yoichi; Hokari, Ryota; Miura, Soichiro; Watanabe, Chikako; Komoto, Shunsuke; Oyama, Tokushige; Kurihara, Chie; Nagata, Hiroshi; Hibi, Toshifumi

2007-11-01

Dietary fat is known to modulate immune functions. Intake of an animal fat-rich diet has been linked to increased risk of inflammation; however, little is known about how animal fat ingestion directly affects intestinal immune function. The objective of this study was to assess the effect of butter feeding on lymphocyte migration in intestinal mucosa and the changes in adhesion molecules and cytokines involved in this effect. T-lymphocytes isolated from the spleen were fluorescence-labeled and injected into recipient mice. Butter was administered into the duodenum, and villus microvessels of the small intestinal mucosa were observed under an intravital microscope. mRNA expression of adhesion molecules and cytokines in the intestinal mucosa were determined by quantitative PCR. The effect of butter feeding on tumor necrosis factor (TNF)-alpha mRNA expression of intestinal macrophages was also determined. Intraluminal butter administration significantly increased lymphocyte adherence to intestinal microvessels accompanied by increases in expression levels of adhesion molecules ICAM-1, MAdCAM-1 and VCAM-1. This accumulation was significantly attenuated by anti-MAdCAM-1 and anti-ICAM-1 antibodies. Butter administration significantly increased TNF-alpha in the lamina proprial macrophages but not interleukin-6. Anti-TNF-alpha treatment attenuated the enhanced expression of adhesion molecules induced by butter administration. T-lymphocyte adherence to microvessels of the small intestinal mucosa was significantly enhanced after butter ingestion. This enhancement is due to increase in expression levels of adhesion molecules of the intestinal mucosa, which is mediated by TNF-alpha from macrophages in the intestinal lamina propria.

Engineering surfaces for bioconjugation: developing strategies and quantifying the extent of the reactions.

PubMed

Gauvreau, Virginie; Chevallier, Pascale; Vallières, Karine; Petitclerc, Eric; Gaudreault, René C; Laroche, Gaétan

2004-01-01

This study presents two-step and multistep reactions for modifying the surface of plasma-functionalized poly(tetrafluoroethylene) (PTFE) surfaces for subsequent conjugation of biologically relevant molecules. First, PTFE films were treated by a radiofrequency glow discharge (RFGD) ammonia plasma to introduce amino groups on the fluoropolymer surface. This plasma treatment is well optimized and allows the incorporation of a relative surface concentration of approximately 2-3.5% of amino groups, as assessed by chemical derivatization followed by X-ray photoelectron spectroscopy (XPS). In a second step, these amino groups were further reacted with various chemical reagents to provide the surface with chemical functionalities such as maleimides, carboxylic acids, acetals, aldehydes, and thiols, that could be used later on to conjugate a wide variety of biologically relevant molecules such as proteins, DNA, drugs, etc. In the present study, glutaric and cis-aconitic anhydrides were evaluated for their capability to provide carboxylic functions to the PTFE plasma-treated surface. Bromoacetaldehyde diethylacetal was reacted with the aminated PTFE surface, providing a diethylacetal function, which is a latent form of aldehyde functionality. Reactions with cross-linkers such as sulfo-succinimidyl derivatives (sulfo-SMCC, sulfo-SMPB) were evaluated to provide a highly reactive maleimide function suitable for further chemical reactions with thiolated molecules. Traut reagent (2-iminothiolane) was also conjugated to introduce a thiol group onto the fluoropolymer surface. PTFE-modified surfaces were analyzed by XPS with a particular attention to quantify the extent of the reactions that occurred on the polymer. Finally, surface immobilization of fibronectin performed using either glutaric anhydride or sulfo-SMPB activators demonstrated the importance of selecting the appropriate conjugation strategy to retain the protein biological activity.

Small-Molecule Photostabilizing Agents are Modifiers of Lipid Bilayer Properties

PubMed Central

Alejo, Jose L.; Blanchard, Scott C.; Andersen, Olaf S.

2013-01-01

Small-molecule photostabilizing or protective agents (PAs) provide essential support for the stability demands on fluorescent dyes in single-molecule spectroscopy and fluorescence microscopy. These agents are employed also in studies of cell membranes and model systems mimicking lipid bilayer environments, but there is little information about their possible effects on membrane structure and physical properties. Given the impact of amphipathic small molecules on bilayer properties such as elasticity and intrinsic curvature, we investigated the effects of six commonly used PAs—cyclooctatetraene (COT), para-nitrobenzyl alcohol (NBA), Trolox (TX), 1,4-diazabicyclo[2.2.2]octane (DABCO), para-nitrobenzoic acid (pNBA), and n-propyl gallate (nPG)—on bilayer properties using a gramicidin A (gA)-based fluorescence quench assay to probe for PA-induced changes in the gramicidin monomer↔dimer equilibrium. The experiments were done using fluorophore-loaded large unilamellar vesicles that had been doped with gA, and changes in the gA monomer↔dimer equilibrium were assayed using a gA channel-permeable fluorescence quencher (Tl+). Changes in bilayer properties caused by, e.g., PA adsorption at the bilayer/solution interface that alter the equilibrium constant for gA channel formation, and thus the number of conducting gA channels in the large unilamellar vesicle membrane, will be detectable as changes in the rate of Tl+ influx—the fluorescence quench rate. Over the experimentally relevant millimolar concentration range, TX, NBA, and pNBA, caused comparable increases in gA channel activity. COT, also in the millimolar range, caused a slight decrease in gA channel activity. nPG increased channel activity at submillimolar concentrations. DABCO did not alter gA activity. Five of the six tested PAs thus alter lipid bilayer properties at experimentally relevant concentrations, which becomes important for the design and analysis of fluorescence studies in cells and model membrane systems. We therefore tested combinations of COT, NBA, and TX; the combinations altered the fluorescence quench rate less than would be predicted assuming their effects on bilayer properties were additive. The combination of equimolar concentrations of COT and NBA caused minimal changes in the fluorescence quench rate. PMID:23746513

Gas-phase study on uridine: Conformation and X-ray photofragmentation

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

Itälä, Eero, E-mail: ersita@utu.fi; Kooser, Kuno; Levola, Helena

2015-05-21

Fragmentation of RNA nucleoside uridine, induced by carbon 1s core ionization, has been studied. The measurements by combined electron and ion spectroscopy have been performed in gas phase utilizing synchrotron radiation. As uridine is a combination of d-ribose and uracil, which have been studied earlier with the same method, this study also considers the effect of chemical environment and the relevant functional groups. Furthermore, since in core ionization the initial core hole is always highly localized, charge migration prior to fragmentation has been studied here. This study also demonstrates the destructive nature of core ionization as in most cases themore » C 1s ionization of uridine leads to concerted explosions producing only small fragments with masses ≤43 amu. In addition to fragmentation patterns, we found out that upon evaporation the sugar part of the uridine molecule attains hexagonal form.« less

Niche-based screening identifies small-molecule inhibitors of leukemia stem cells.

PubMed

Hartwell, Kimberly A; Miller, Peter G; Mukherjee, Siddhartha; Kahn, Alissa R; Stewart, Alison L; Logan, David J; Negri, Joseph M; Duvet, Mildred; Järås, Marcus; Puram, Rishi; Dancik, Vlado; Al-Shahrour, Fatima; Kindler, Thomas; Tothova, Zuzana; Chattopadhyay, Shrikanta; Hasaka, Thomas; Narayan, Rajiv; Dai, Mingji; Huang, Christina; Shterental, Sebastian; Chu, Lisa P; Haydu, J Erika; Shieh, Jae Hung; Steensma, David P; Munoz, Benito; Bittker, Joshua A; Shamji, Alykhan F; Clemons, Paul A; Tolliday, Nicola J; Carpenter, Anne E; Gilliland, D Gary; Stern, Andrew M; Moore, Malcolm A S; Scadden, David T; Schreiber, Stuart L; Ebert, Benjamin L; Golub, Todd R

2013-12-01

Efforts to develop more effective therapies for acute leukemia may benefit from high-throughput screening systems that reflect the complex physiology of the disease, including leukemia stem cells (LSCs) and supportive interactions with the bone marrow microenvironment. The therapeutic targeting of LSCs is challenging because LSCs are highly similar to normal hematopoietic stem and progenitor cells (HSPCs) and are protected by stromal cells in vivo. We screened 14,718 compounds in a leukemia-stroma co-culture system for inhibition of cobblestone formation, a cellular behavior associated with stem-cell function. Among those compounds that inhibited malignant cells but spared HSPCs was the cholesterol-lowering drug lovastatin. Lovastatin showed anti-LSC activity in vitro and in an in vivo bone marrow transplantation model. Mechanistic studies demonstrated that the effect was on target, via inhibition of HMG-CoA reductase. These results illustrate the power of merging physiologically relevant models with high-throughput screening.

Niche-based screening identifies small-molecule inhibitors of leukemia stem cells

PubMed Central

Mukherjee, Siddhartha; Kahn, Alissa R; Stewart, Alison L; Logan, David J; Negri, Joseph M; Duvet, Mildred; Järås, Marcus; Puram, Rishi; Dancik, Vlado; Al-Shahrour, Fatima; Kindler, Thomas; Tothova, Zuzana; Chattopadhyay, Shrikanta; Hasaka, Thomas; Narayan, Rajiv; Dai, Mingji; Huang, Christina; Shterental, Sebastian; Chu, Lisa P; Haydu, J Erika; Shieh, Jae Hung; Steensma, David P; Munoz, Benito; Bittker, Joshua A; Shamji, Alykhan F; Clemons, Paul A; Tolliday, Nicola J; Carpenter, Anne E; Gilliland, D Gary; Stern, Andrew M; Moore, Malcolm A S; Scadden, David T; Schreiber, Stuart L; Ebert, Benjamin L; Golub, Todd R

2014-01-01

Efforts to develop more effective therapies for acute leukemia may benefit from high-throughput screening systems that reflect the complex physiology of the disease, including leukemia stem cells (LSCs) and supportive interactions with the bone-marrow microenvironment. The therapeutic targeting of LSCs is challenging because LSCs are highly similar to normal hematopoietic stem and progenitor cells (HSPCs) and are protected by stromal cells in vivo. We screened 14,718 compounds in a leukemia-stroma co-culture system for inhibition of cobblestone formation, a cellular behavior associated with stem-cell function. Among those that inhibited malignant cells but spared HSPCs was the cholesterol-lowering drug lovastatin. Lovastatin showed anti-LSC activity in vitro and in an in vivo bone marrow transplantation model. Mechanistic studies demonstrated that the effect was on-target, via inhibition of HMGCoA reductase. These results illustrate the power of merging physiologically-relevant models with high-throughput screening. PMID:24161946

Machine Learning Estimates of Natural Product Conformational Energies

PubMed Central

Rupp, Matthias; Bauer, Matthias R.; Wilcken, Rainer; Lange, Andreas; Reutlinger, Michael; Boeckler, Frank M.; Schneider, Gisbert

2014-01-01

Machine learning has been used for estimation of potential energy surfaces to speed up molecular dynamics simulations of small systems. We demonstrate that this approach is feasible for significantly larger, structurally complex molecules, taking the natural product Archazolid A, a potent inhibitor of vacuolar-type ATPase, from the myxobacterium Archangium gephyra as an example. Our model estimates energies of new conformations by exploiting information from previous calculations via Gaussian process regression. Predictive variance is used to assess whether a conformation is in the interpolation region, allowing a controlled trade-off between prediction accuracy and computational speed-up. For energies of relaxed conformations at the density functional level of theory (implicit solvent, DFT/BLYP-disp3/def2-TZVP), mean absolute errors of less than 1 kcal/mol were achieved. The study demonstrates that predictive machine learning models can be developed for structurally complex, pharmaceutically relevant compounds, potentially enabling considerable speed-ups in simulations of larger molecular structures. PMID:24453952

Chemical Foundations of Hydrogen Sulfide Biology

PubMed Central

Li, Qian; Lancaster, Jack R.

2013-01-01

Following nitric oxide (nitrogen monoxide) and carbon monoxide, hydrogen sulfide (or its newer systematic name sulfane, H2S) became the third small molecule that can be both toxic and beneficial depending on the concentration. In spite of its impressive therapeutic potential, the underlying mechanisms for its beneficial effects remain unclear. Any novel mechanism has to obey fundamental chemical principles. H2S chemistry was studied long before its biological relevance was discovered, however, with a few exceptions, these past works have received relatively little attention in the path of exploring the mechanistic conundrum of H2S biological functions. This review calls attention to the basic physical and chemical properties of H2S, focuses on the chemistry between H2S and its three potential biological targets: oxidants, metals and thiol derivatives, discusses the applications of these basics into H2S biology and methodology, and introduces the standard terminology to this youthful field. PMID:23850631

A whole organism screen identifies novel regulators of fat storage

PubMed Central

Lemieux, George A.; Liu, Jason; Mayer, Nasima; Bainton, Roland J.; Ashrafi, Kaveh; Werb, Zena

2011-01-01

The regulation of energy homeostasis integrates diverse biological processes ranging from behavior to metabolism and is linked fundamentally to numerous disease states. To identify new molecules that can bypass homeostatic compensatory mechanisms of energy balance in intact animals, we screened for small molecule modulators of C. elegans fat content. We report on several molecules that modulate fat storage without obvious deleterious effects on feeding, growth, and reproduction. A subset of these compounds also altered fat storage in mammalian and insect cell culture. We found that one of the newly identified compounds exerts its effects in C. elegans through a pathway that requires novel functions of an AMP-activated kinase catalytic subunit and a transcription factor previously unassociated with fat regulation. Thus, our strategy identifies small molecules that are effective within the context of intact animals and reveals relationships between new pathways that operate across phyla to influence energy homeostasis. PMID:21390037

Recent progress in human telomere RNA structure and function.

PubMed

Xu, Yan

2018-06-14

Human telomeric DNA is transcribed into telomeric RNA in cells. Telomeric RNA performs the fundamental biological functions such as regulation and protection of chromosome ends. This digest highlights the human telomere RNA G-quadruplex structures, telomere RNA functions, G-quadruplex-binding small molecules, and future prospects. Copyright © 2018 Elsevier Ltd. All rights reserved.

Long-Term Priming by Three Small Molecules Is a Promising Strategy for Enhancing Late Endothelial Progenitor Cell Bioactivities.

PubMed

Kim, Yeon-Ju; Ji, Seung Taek; Kim, Da Yeon; Jung, Seok Yun; Kang, Songhwa; Park, Ji Hye; Jang, Woong Bi; Yun, Jisoo; Ha, Jongseong; Lee, Dong Hyung; Kwon, Sang-Mo

2018-06-12

Endothelial progenitor cells (EPCs) and outgrowth endothelial cells (OECs) play a pivotal role in vascular regeneration in ischemic tissues; however, their therapeutic application in clinical settings is limited due to the low quality and quantity of patient-derived circulating EPCs. To solve this problem, we evaluated whether three priming small molecules (tauroursodeoxycholic acid, fucoidan, oleuropein) could enhance the angiogenic potential of EPCs. Such enhancement would promote the cellular bioactivities and help to develop functionally improved EPC therapeutics for ischemic diseases by accelerating the priming effect of the defined physiological molecules. We found that preconditioning of each of the three small molecules significantly induced the differentiation potential of CD34+ stem cells into EPC lineage cells. Notably, long-term priming of OECs with the three chemical cocktail (OEC-3C) increased the proliferation potential of EPCs via ERK activation. The migration, invasion, and tube-forming capacities were also significantly enhanced in OEC-3Cs compared with unprimed OECs. Further, the cell survival ratio was dramatically increased in OEC-3Cs against H2O2-induced oxidative stress via the augmented expression of Bcl-2, a prosurvival protein. In conclusion, we identified three small molecules for enhancing the bioactivities of ex vivo-expanded OECs for vascular repair. Long-term 3C priming might be a promising methodology for EPC-based therapy against ischemic diseases.

Effects of Acids, Bases, and Heteroatoms on Proximal Radial Distribution Functions for Proteins

PubMed Central

Nguyen, Bao Linh; Pettitt, B. Montgomery

2015-01-01

The proximal distribution of water around proteins is a convenient method of quantifying solvation. We consider the effect of charged and sulfur-containing amino acid side-chain atoms on the proximal radial distribution function (pRDF) of water molecules around proteins using side-chain analogs. The pRDF represents the relative probability of finding any solvent molecule at a distance from the closest or surface perpendicular protein atom. We consider the near-neighbor distribution. Previously, pRDFs were shown to be universal descriptors of the water molecules around C, N, and O atom types across hundreds of globular proteins. Using averaged pRDFs, a solvent density around any globular protein can be reconstructed with controllable relative error. Solvent reconstruction using the additional information from charged amino acid side-chain atom types from both small models and protein averages reveals the effects of surface charge distribution on solvent density and improves the reconstruction errors relative to simulation. Solvent density reconstructions from the small-molecule models are as effective and less computationally demanding than reconstructions from full macromolecular models in reproducing preferred hydration sites and solvent density fluctuations. PMID:26388706

Single molecule optical measurements of orientation and rotations of biological macromolecules.

PubMed

Shroder, Deborah Y; Lippert, Lisa G; Goldman, Yale E

2016-11-22

Subdomains of macromolecules often undergo large orientation changes during their catalytic cycles that are essential for their activity. Tracking these rearrangements in real time opens a powerful window into the link between protein structure and functional output. Site-specific labeling of individual molecules with polarized optical probes and measurement of their spatial orientation can give insight into the crucial conformational changes, dynamics, and fluctuations of macromolecules. Here we describe the range of single molecule optical technologies that can extract orientation information from these probes, review the relevant types of probes and labeling techniques, and highlight the advantages and disadvantages of these technologies for addressing specific inquiries.

The personal receiving document management and the realization of email function in OAS

NASA Astrophysics Data System (ADS)

Li, Biqing; Li, Zhao

2017-05-01

This software is an independent software system, suitable for small and medium enterprises, contains personal office, scientific research project management and system management functions, independently run in relevant environment, and to solve practical needs. This software is an independent software system, using the current popular B/S (browser/server) structure and ASP.NET technology development, using the Windows 7 operating system, Microsoft SQL Server2005 Visual2008 and database as a development platform, suitable for small and medium enterprises, contains personal office, scientific research project management and system management functions, independently run in relevant environment, and to solve practical needs.

Constraint Network Analysis (CNA): a Python software package for efficiently linking biomacromolecular structure, flexibility, (thermo-)stability, and function.

PubMed

Pfleger, Christopher; Rathi, Prakash Chandra; Klein, Doris L; Radestock, Sebastian; Gohlke, Holger

2013-04-22

For deriving maximal advantage from information on biomacromolecular flexibility and rigidity, results from rigidity analyses must be linked to biologically relevant characteristics of a structure. Here, we describe the Python-based software package Constraint Network Analysis (CNA) developed for this task. CNA functions as a front- and backend to the graph-based rigidity analysis software FIRST. CNA goes beyond the mere identification of flexible and rigid regions in a biomacromolecule in that it (I) provides a refined modeling of thermal unfolding simulations that also considers the temperature-dependence of hydrophobic tethers, (II) allows performing rigidity analyses on ensembles of network topologies, either generated from structural ensembles or by using the concept of fuzzy noncovalent constraints, and (III) computes a set of global and local indices for quantifying biomacromolecular stability. This leads to more robust results from rigidity analyses and extends the application domain of rigidity analyses in that phase transition points ("melting points") and unfolding nuclei ("structural weak spots") are determined automatically. Furthermore, CNA robustly handles small-molecule ligands in general. Such advancements are important for applying rigidity analysis to data-driven protein engineering and for estimating the influence of ligand molecules on biomacromolecular stability. CNA maintains the efficiency of FIRST such that the analysis of a single protein structure takes a few seconds for systems of several hundred residues on a single core. These features make CNA an interesting tool for linking biomacromolecular structure, flexibility, (thermo-)stability, and function. CNA is available from http://cpclab.uni-duesseldorf.de/software for nonprofit organizations.

Nanopore Device for Reversible Ion and Molecule Sensing or Migration

NASA Technical Reports Server (NTRS)

Seger, R. Adam (Inventor); Pourmand, Nader (Inventor); Actis, Paolo (Inventor); Singaram, Bakthan (Inventor); Vilozny, Boaz (Inventor)

2015-01-01

Disclosed are methods and devices for detection of ion migration and binding, utilizing a nanopipette adapted for use in an electrochemical sensing circuit. The nanopipette may be functionalized on its interior bore with metal chelators for binding and sensing metal ions or other specific binding molecules such as boronic acid for binding and sensing glucose. Such a functionalized nanopipette is comprised in an electrical sensor that detects when the nanopipette selectively and reversibly binds ions or small molecules. Also disclosed is a nanoreactor, comprising a nanopipette, for controlling precipitation in aqueous solutions by voltage-directed ion migration, wherein ions may be directed out of the interior bore by a repulsing charge in the bore.

Increasing rigor in NMR-based metabolomics through validated and open source tools

PubMed Central

Eghbalnia, Hamid R; Romero, Pedro R; Westler, William M; Baskaran, Kumaran; Ulrich, Eldon L; Markley, John L

2016-01-01

The metabolome, the collection of small molecules associated with an organism, is a growing subject of inquiry, with the data utilized for data-intensive systems biology, disease diagnostics, biomarker discovery, and the broader characterization of small molecules in mixtures. Owing to their close proximity to the functional endpoints that govern an organism’s phenotype, metabolites are highly informative about functional states. The field of metabolomics identifies and quantifies endogenous and exogenous metabolites in biological samples. Information acquired from nuclear magnetic spectroscopy (NMR), mass spectrometry (MS), and the published literature, as processed by statistical approaches, are driving increasingly wider applications of metabolomics. This review focuses on the role of databases and software tools in advancing the rigor, robustness, reproducibility, and validation of metabolomics studies. PMID:27643760

Synthetic, structural mimetics of the β-hairpin flap of HIV-1 protease inhibit enzyme function.

PubMed

Chauhan, Jay; Chen, Shen-En; Fenstermacher, Katherine J; Naser-Tavakolian, Aurash; Reingewertz, Tali; Salmo, Rosene; Lee, Christian; Williams, Emori; Raje, Mithun; Sundberg, Eric; DeStefano, Jeffrey J; Freire, Ernesto; Fletcher, Steven

2015-11-01

Small-molecule mimetics of the β-hairpin flap of HIV-1 protease (HIV-1 PR) were designed based on a 1,4-benzodiazepine scaffold as a strategy to interfere with the flap-flap protein-protein interaction, which functions as a gated mechanism to control access to the active site. Michaelis-Menten kinetics suggested our small-molecules are competitive inhibitors, which indicates the mode of inhibition is through binding the active site or sterically blocking access to the active site and preventing flap closure, as designed. More generally, a new bioactive scaffold for HIV-1PR inhibition has been discovered, with the most potent compound inhibiting the protease with a modest K(i) of 11 μM. Copyright © 2015 Elsevier Ltd. All rights reserved.

Small Molecules Targeting the miRNA-Binding Domain of Argonaute 2: From Computer-Aided Molecular Design to RNA Immunoprecipitation.

PubMed

Bellissimo, Teresa; Masciarelli, Silvia; Poser, Elena; Genovese, Ilaria; Del Rio, Alberto; Colotti, Gianni; Fazi, Francesco

2017-01-01

The development of small-molecule-based target therapy design for human disease and cancer is object of growing attention. Recently, specific microRNA (miRNA) mimicking compounds able to bind the miRNA-binding domain of Argonaute 2 protein (AGO2) to inhibit miRNA loading and its functional activity were described. Computer-aided molecular design techniques and RNA immunoprecipitation represent suitable approaches to identify and experimentally determine if a compound is able to impair the loading of miRNAs on AGO2 protein. Here, we describe these two methodologies that we recently used to select a specific compound able to interfere with the AGO2 functional activity and able to improve the retinoic acid-dependent myeloid differentiation of leukemic cells.

Increasing rigor in NMR-based metabolomics through validated and open source tools.

PubMed

Eghbalnia, Hamid R; Romero, Pedro R; Westler, William M; Baskaran, Kumaran; Ulrich, Eldon L; Markley, John L

2017-02-01

The metabolome, the collection of small molecules associated with an organism, is a growing subject of inquiry, with the data utilized for data-intensive systems biology, disease diagnostics, biomarker discovery, and the broader characterization of small molecules in mixtures. Owing to their close proximity to the functional endpoints that govern an organism's phenotype, metabolites are highly informative about functional states. The field of metabolomics identifies and quantifies endogenous and exogenous metabolites in biological samples. Information acquired from nuclear magnetic spectroscopy (NMR), mass spectrometry (MS), and the published literature, as processed by statistical approaches, are driving increasingly wider applications of metabolomics. This review focuses on the role of databases and software tools in advancing the rigor, robustness, reproducibility, and validation of metabolomics studies. Copyright © 2016. Published by Elsevier Ltd.

Fluorescein isothiocyanate-labeled human plasma fibronectin in extracellular matrix remodeling.

PubMed

Hoffmann, Celine; Leroy-Dudal, Johanne; Patel, Salima; Gallet, Olivier; Pauthe, Emmanuel

2008-01-01

Fluorescein isothiocyanate (FITC) is a well-known probe for labeling biologically relevant proteins. However, the impact of the labeling procedure on protein structure and biological activities remains unclear. In this work, FITC-labeled human plasma fibronectin (Fn) was developed to gain insight into the dynamic relationship between cells and Fn. The similarities and differences concerning the structure and function between Fn-FITC and standard Fn were evaluated using biochemical as well as cellular approaches. By varying the FITC/Fn ratio, we demonstrated that overlabeling (>10 FITC molecules/Fn molecule) induces probe fluorescence quenching, protein aggregation, and cell growth modifications. A correct balance between reliable fluorescence for detection and no significant modifications to structure and biological function compared with standard Fn was obtained with a final ratio of 3 FITC molecules per Fn molecule (Fn-FITC3). Fn-FITC3, similar to standard Fn, is correctly recruited into the cell matrix network. Also, Fn-FITC3 is proposed to be a powerful molecular tool to investigate Fn organization and cellular behavior concomitantly.

Quantum rotor model for a Bose-Einstein condensate of dipolar molecules.

PubMed

Armaitis, J; Duine, R A; Stoof, H T C

2013-11-22

We show that a Bose-Einstein condensate of heteronuclear molecules in the regime of small and static electric fields is described by a quantum rotor model for the macroscopic electric dipole moment of the molecular gas cloud. We solve this model exactly and find the symmetric, i.e., rotationally invariant, and dipolar phases expected from the single-molecule problem, but also an axial and planar nematic phase due to many-body effects. Investigation of the wave function of the macroscopic dipole moment also reveals squeezing of the probability distribution for the angular momentum of the molecules.

MPAI (mass probes aided ionization) method for total analysis of biomolecules by mass spectrometry.

PubMed

Honda, Aki; Hayashi, Shinichiro; Hifumi, Hiroki; Honma, Yuya; Tanji, Noriyuki; Iwasawa, Naoko; Suzuki, Yoshio; Suzuki, Koji

2007-01-01

We have designed and synthesized various mass probes, which enable us to effectively ionize various molecules to be detected with mass spectrometry. We call the ionization method using mass probes the "MPAI (mass probes aided ionization)" method. We aim at the sensitive detection of various biological molecules, and also the detection of bio-molecules by a single mass spectrometry serially without changing the mechanical settings. Here, we review mass probes for small molecules with various functional groups and mass probes for proteins. Further, we introduce newly developed mass probes for proteins for highly sensitive detection.

Chemogenomics: a discipline at the crossroad of high throughput technologies, biomarker research, combinatorial chemistry, genomics, cheminformatics, bioinformatics and artificial intelligence.

PubMed

Maréchal, Eric

2008-09-01

Chemogenomics is the study of the interaction of functional biological systems with exogenous small molecules, or in broader sense the study of the intersection of biological and chemical spaces. Chemogenomics requires expertises in biology, chemistry and computational sciences (bioinformatics, cheminformatics, large scale statistics and machine learning methods) but it is more than the simple apposition of each of these disciplines. Biological entities interacting with small molecules can be isolated proteins or more elaborate systems, from single cells to complete organisms. The biological space is therefore analyzed at various postgenomic levels (genomic, transcriptomic, proteomic or any phenotypic level). The space of small molecules is partially real, corresponding to commercial and academic collections of compounds, and partially virtual, corresponding to the chemical space possibly synthesizable. Synthetic chemistry has developed novel strategies allowing a physical exploration of this universe of possibilities. A major challenge of cheminformatics is to charter the virtual space of small molecules using realistic biological constraints (bioavailability, druggability, structural biological information). Chemogenomics is a descendent of conventional pharmaceutical approaches, since it involves the screening of chemolibraries for their effect on biological targets, and benefits from the advances in the corresponding enabling technologies and the introduction of new biological markers. Screening was originally motivated by the rigorous discovery of new drugs, neglecting and throwing away any molecule that would fail to meet the standards required for a therapeutic treatment. It is now the basis for the discovery of small molecules that might or might not be directly used as drugs, but which have an immense potential for basic research, as probes to explore an increasing number of biological phenomena. Concerns about the environmental impact of chemical industry open new fields of research for chemogenomics.

Small Molecule Interactome Mapping by Photoaffinity Labeling Reveals Binding Site Hotspots for the NSAIDs.

PubMed

Gao, Jinxu; Mfuh, Adelphe; Amako, Yuka; Woo, Christina M

2018-03-28

Many therapeutics elicit cell-type specific polypharmacology that is executed by a network of molecular recognition events between a small molecule and the whole proteome. However, measurement of the structures that underpin the molecular associations between the proteome and even common therapeutics, such as the nonsteroidal anti-inflammatory drugs (NSAIDs), is limited by the inability to map the small molecule interactome. To address this gap, we developed a platform termed small molecule interactome mapping by photoaffinity labeling (SIM-PAL) and applied it to the in cellulo direct characterization of specific NSAID binding sites. SIM-PAL uses (1) photochemical conjugation of NSAID derivatives in the whole proteome and (2) enrichment and isotope-recoding of the conjugated peptides for (3) targeted mass spectrometry-based assignment. Using SIM-PAL, we identified the NSAID interactome consisting of over 1000 significantly enriched proteins and directly characterized nearly 200 conjugated peptides representing direct binding sites of the photo-NSAIDs with proteins from Jurkat and K562 cells. The enriched proteins were often identified as parts of complexes, including known targets of NSAID activity (e.g., NF-κB) and novel interactions (e.g., AP-2, proteasome). The conjugated peptides revealed direct NSAID binding sites from the cell surface to the nucleus and a specific binding site hotspot for the three photo-NSAIDs on histones H2A and H2B. NSAID binding stabilized COX-2 and histone H2A by cellular thermal shift assay. Since small molecule stabilization of protein complexes is a gain of function regulatory mechanism, it is conceivable that NSAIDs affect biological processes through these broader proteomic interactions. SIM-PAL enabled characterization of NSAID binding site hotspots and is amenable to map global binding sites for virtually any molecule of interest.

Detoxification function of the Arabidopsis sulphotransferase AtSOT12 by sulphonation of xenobiotics.

PubMed

Chen, Jinhua; Gao, Liqiong; Baek, Dongwon; Liu, Chunlin; Ruan, Ying; Shi, Huazhong

2015-08-01

Cytosolic sulphotransferases have been implicated in inactivation of endogenous steroid hormones and detoxification of xenobiotics in human and animals. Yet, the function of plant sulphotransferases in xenobiotic sulphonation and detoxification has not been reported. In this study, we show that the Arabidopsis sulphotransferase AtSOT12 could sulphonate the bacterial-produced toxin cycloheximide. Loss-of-function mutant sot12 exhibited hypersensitive phenotype to cycloheximide, and expression of AtSOT12 protein in yeast cells conferred resistance to this toxic compound. AtSOT12 exhibited broad specificity and could sulphonate a variety of xenobiotics including phenolic and polycyclic compounds. Enzyme kinetics analysis indicated that AtSOT12 has different selectivity for simple phenolics with different side chains, and the position of the side chain in the simple phenolic compounds affects substrate binding affinity and catalytic efficiency. We proposed that the broad specificity and induced production of AtSOT12 may have rendered this enzyme to not only modify endogenous molecules such as salicylic acid as we previously reported, but also sulphonate pathogen-produced toxic small molecules to protect them from infection. Sulphonation of small molecules in plants may constitute a rapid way to inactivate or change the physiochemical properties of biologically active molecules that could have profound effects on plant growth, development and defence. © 2015 John Wiley & Sons Ltd.

Pharmacologic Induction of CD8+ T Cell Memory: Better Living Through Chemistry

PubMed Central

Gattinoni, Luca; Klebanoff, Christopher A.; Restifo, Nicholas P.

2011-01-01

The generation of a robust population of memory T cells is critical for effective vaccine and cell-based therapies to prevent and treat infectious diseases and cancer. A series of recent papers have established a new, cell-intrinsic approach in which small molecules target key metabolic and developmental pathways to enhance the formation and maintenance of highly functional CD8+ memory T cells. These findings raise the exciting new possibility of using small molecules, many of which are already approved for human use, for the pharmacologic induction of immunologic memory. PMID:20371454

Rational tuning of high-energy visible light absorption for panchromatic small molecules by a two-dimensional conjugation approach

DOE PAGES

He, B.; Zherebetskyy, D.; Wang, H.; ...

2016-02-29

We have demonstrated a rational two-dimensional (2D) conjugation approach towards achieving panchromatic absorption of small molecules. Furthermore, by extending the conjugation on two orthogonal axes of an electron acceptor, namely, bay-annulated indigo (BAI), the optical absorptions could be tuned independently in both high- and low-energy regions. The unconventional modulation of the high-energy absorption is rationalized by density functional theory (DFT) calculations. Finally, we determine that a 2D tuning strategy provides novel guidelines for the design of molecular materials with tailored optoelectronic properties.

Spatial Molecular Architecture of the Microbial Community of a Peltigera Lichen.

PubMed

Garg, Neha; Zeng, Yi; Edlund, Anna; Melnik, Alexey V; Sanchez, Laura M; Mohimani, Hosein; Gurevich, Alexey; Miao, Vivian; Schiffler, Stefan; Lim, Yan Wei; Luzzatto-Knaan, Tal; Cai, Shengxin; Rohwer, Forest; Pevzner, Pavel A; Cichewicz, Robert H; Alexandrov, Theodore; Dorrestein, Pieter C

2016-01-01

Microbes are commonly studied as individual species, but they exist as mixed assemblages in nature. At present, we know very little about the spatial organization of the molecules, including natural products that are produced within these microbial networks. Lichens represent a particularly specialized type of symbiotic microbial assemblage in which the component microorganisms exist together. These composite microbial assemblages are typically comprised of several types of microorganisms representing phylogenetically diverse life forms, including fungi, photosymbionts, bacteria, and other microbes. Here, we employed matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) imaging mass spectrometry to characterize the distributions of small molecules within a Peltigera lichen. In order to probe how small molecules are organized and localized within the microbial consortium, analytes were annotated and assigned to their respective producer microorganisms using mass spectrometry-based molecular networking and metagenome sequencing. The spatial analysis of the molecules not only reveals an ordered layering of molecules within the lichen but also supports the compartmentalization of unique functions attributed to various layers. These functions include chemical defense (e.g., antibiotics), light-harvesting functions associated with the cyanobacterial outer layer (e.g., chlorophyll), energy transfer (e.g., sugars) surrounding the sun-exposed cyanobacterial layer, and carbohydrates that may serve a structural or storage function and are observed with higher intensities in the non-sun-exposed areas (e.g., complex carbohydrates). IMPORTANCE Microbial communities have evolved over centuries to live symbiotically. The direct visualization of such communities at the chemical and functional level presents a challenge. Overcoming this challenge may allow one to visualize the spatial distributions of specific molecules involved in symbiosis and to define their functional roles in shaping the community structure. In this study, we examined the diversity of microbial genes and taxa and the presence of biosynthetic gene clusters by metagenomic sequencing and the compartmentalization of organic chemical components within a lichen using mass spectrometry. This approach allowed the identification of chemically distinct sections within this composite organism. Using our multipronged approach, various fungal natural products, not previously reported from lichens, were identified and two different fungal layers were visualized at the chemical level.

Spatial Molecular Architecture of the Microbial Community of a Peltigera Lichen

PubMed Central

Garg, Neha; Zeng, Yi; Edlund, Anna; Melnik, Alexey V.; Mohimani, Hosein; Gurevich, Alexey; Miao, Vivian; Schiffler, Stefan; Lim, Yan Wei; Luzzatto-Knaan, Tal; Cai, Shengxin; Rohwer, Forest; Pevzner, Pavel A.; Cichewicz, Robert H.; Alexandrov, Theodore

2016-01-01

ABSTRACT Microbes are commonly studied as individual species, but they exist as mixed assemblages in nature. At present, we know very little about the spatial organization of the molecules, including natural products that are produced within these microbial networks. Lichens represent a particularly specialized type of symbiotic microbial assemblage in which the component microorganisms exist together. These composite microbial assemblages are typically comprised of several types of microorganisms representing phylogenetically diverse life forms, including fungi, photosymbionts, bacteria, and other microbes. Here, we employed matrix-assisted laser desorption ionization–time of flight (MALDI-TOF) imaging mass spectrometry to characterize the distributions of small molecules within a Peltigera lichen. In order to probe how small molecules are organized and localized within the microbial consortium, analytes were annotated and assigned to their respective producer microorganisms using mass spectrometry-based molecular networking and metagenome sequencing. The spatial analysis of the molecules not only reveals an ordered layering of molecules within the lichen but also supports the compartmentalization of unique functions attributed to various layers. These functions include chemical defense (e.g., antibiotics), light-harvesting functions associated with the cyanobacterial outer layer (e.g., chlorophyll), energy transfer (e.g., sugars) surrounding the sun-exposed cyanobacterial layer, and carbohydrates that may serve a structural or storage function and are observed with higher intensities in the non-sun-exposed areas (e.g., complex carbohydrates). IMPORTANCE Microbial communities have evolved over centuries to live symbiotically. The direct visualization of such communities at the chemical and functional level presents a challenge. Overcoming this challenge may allow one to visualize the spatial distributions of specific molecules involved in symbiosis and to define their functional roles in shaping the community structure. In this study, we examined the diversity of microbial genes and taxa and the presence of biosynthetic gene clusters by metagenomic sequencing and the compartmentalization of organic chemical components within a lichen using mass spectrometry. This approach allowed the identification of chemically distinct sections within this composite organism. Using our multipronged approach, various fungal natural products, not previously reported from lichens, were identified and two different fungal layers were visualized at the chemical level. PMID:28028548

The Formation of Racemic Amino Acids by UV Photolysis of Interstellar Ice Analogs

NASA Technical Reports Server (NTRS)

Bernstein, Max P.; Dworkin, Jason P.; Sandford, Scott A.; Cooper, George; Allamandola, Louis J.; DeVincenzi, Donald (Technical Monitor)

2001-01-01

Small biologically relevant organic molecules including the amino acids glycine, alanine, and marine were formed in the laboratory by the UV (Ultraviolet) photolysis of realistic interstellar ice analogs, composed primarily of H2O, and including CH3OH, NH3, and HCN, under interstellar conditions. N-formyl glycine, cycloserine (4-amino-3-isoxazolidinone), and glycerol were detected before hydrolysis, and glycine, racemic alanine, racemic marine, glycerol, ethanolamine, and glyceric acid were found after hydrolysis. This suggests that some meteoritic amino acids (and other molecules) may be the direct result of interstellar ice photochemistry, expanding the current paradigm that they formed by reactions in liquid water on meteorite parent bodies.

Identification of small non-coding RNA classes expressed in swine whole blood during HP-PRRSV infection

USDA-ARS?s Scientific Manuscript database

It has been established that reduced susceptibility to porcine reproductive and respiratory syndrome virus (PRRSV) has a genetic component. This genetic component may take the form of small non-coding RNAs (sncRNA), which are molecules that function as regulators of gene expression. Various sncRNAs ...

An interaction map of small-molecule kinase inhibitors with anaplastic lymphoma kinase (ALK) mutants in ALK-positive non-small cell lung cancer.

PubMed

Ai, Xinghao; Shen, Shengping; Shen, Lan; Lu, Shun

2015-05-01

Human anaplastic lymphoma kinase (ALK) has become a well-established target for the treatment of ALK-positive non-small cell lung cancer (NSCLC). Here, we have profiled seven small-molecule inhibitors, including 2 that are approved drugs, against a panel of clinically relevant mutations in ALK tyrosine kinase (TK) domain, aiming at a comprehensive understanding of molecular mechanism and biological implication underlying inhibitor response to ALK TK mutation. We find that (i) the gatekeeper mutation L1196M causes crizotinib resistance by simultaneously increasing and decreasing the binding affinities of, respectively, ATP and inhibitor to ALK, whereas the secondary mutation C1156Y, which is located far away from the ATP-binding site of ALK TK domain, causes the resistance by inducing marked allosteric effect on the site, (ii) the 2nd and 3rd generation kinase inhibitors exhibit relatively high sensitivity towards ALK mutants as compared to 1st generation inhibitors, (iii) the pan-kinase inhibitor staurosporine is insensitive for most mutations due to its high structural compatibility, and (iv) ATP affinity to ALK is generally reduced upon most clinically relevant mutations. Furthermore, we also identify six novel mutation-inhibitor pairs that are potentially associated with drug resistance. In addition, the G1202R and C1156Y mutations are expected to generally cause resistance for many existing inhibitors, since they can address significant effect on the geometric shape and physicochemical property of ALK active pocket. Copyright © 2015 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

Highly sensitive detection of target molecules using a new fluorescence-based bead assay

NASA Astrophysics Data System (ADS)

Scheffler, Silvia; Strauß, Denis; Sauer, Markus

2007-07-01

Development of immunoassays with improved sensitivity, specificity and reliability are of major interest in modern bioanalytical research. We describe the development of a new immunomagnetic fluorescence detection (IM-FD) assay based on specific antigen/antibody interactions and on accumulation of the fluorescence signal on superparamagnetic PE beads in combination with the use of extrinsic fluorescent labels. IM-FD can be easily modified by varying the order of coatings and assay conditions. Depending on the target molecule, antibodies (ABs), entire proteins, or small protein epitopes can be used as capture molecules. The presence of target molecules is detected by fluorescence microscopy using fluorescently labeled secondary or detection antibodies. Here, we demonstrate the potential of the new assay detecting the two tumor markers IGF-I and p53 antibodies in the clinically relevant concentration range. Our data show that the fluorescence-based bead assay exhibits a large dynamic range and a high sensitivity down to the subpicomolar level.

Förster-Induced Energy Transfer in Functionalized Graphene

PubMed Central

2014-01-01

Carbon nanostructures are ideal substrates for functionalization with molecules since they consist of a single atomic layer giving rise to an extraordinary sensitivity to changes in their surrounding. The functionalization opens a new research field of hybrid nanostructures with tailored properties. Here, we present a microscopic view on the substrate–molecule interaction in the exemplary hybrid material consisting of graphene functionalized with perylene molecules. First experiments on similar systems have been recently realized illustrating an extremely efficient transfer of excitation energy from adsorbed molecules to the carbon substrate, a process with a large application potential for high-efficiency photovoltaic devices and biomedical imaging and sensing. So far, there has been no microscopically founded explanation for the observed energy transfer. Based on first-principle calculations, we have explicitly investigated the different transfer mechanisms revealing the crucial importance of Förster coupling. Due to the efficient Coulomb interaction in graphene, we obtain strong Förster rates in the range of 1/fs. We investigate its dependence on the substrate–molecule distance R and describe the impact of the momentum transfer q for an efficient energy transfer. Furthermore, we find that the Dexter transfer mechanism is negligibly small due to the vanishing overlap between the involved strongly localized orbital functions. The gained insights are applicable to a variety of carbon-based hybrid nanostructures. PMID:24808936

Challenges for single molecule electronic devices with nanographene and organic molecules. Do single molecules offer potential as elements of electronic devices in the next generation?

NASA Astrophysics Data System (ADS)

Enoki, Toshiaki; Kiguchi, Manabu

2018-03-01

Interest in utilizing organic molecules to fabricate electronic materials has existed ever since organic (molecular) semiconductors were first discovered in the 1950s. Since then, scientists have devoted serious effort to the creation of various molecule-based electronic systems, such as molecular metals and molecular superconductors. Single-molecule electronics and the associated basic science have emerged over the past two decades and provided hope for the development of highly integrated molecule-based electronic devices in the future (after the Si-based technology era has ended). Here, nanographenes (nano-sized graphene) with atomically precise structures are among the most promising molecules that can be utilized for electronic/spintronic devices. To manipulate single small molecules for an electronic device, a single molecular junction has been developed. It is a powerful tool that allows even small molecules to be utilized. External electric, magnetic, chemical, and mechanical perturbations can change the physical and chemical properties of molecules in a way that is different from bulk materials. Therefore, the various functionalities of molecules, along with changes induced by external perturbations, allows us to create electronic devices that we cannot create using current top-down Si-based technology. Future challenges that involve the incorporation of condensed matter physics, quantum chemistry calculations, organic synthetic chemistry, and electronic device engineering are expected to open a new era in single-molecule device electronic technology.

Increasing the Endoplasmic Reticulum Pool of the F508del Allele of the Cystic Fibrosis Transmembrane Conductance Regulator Leads to Greater Folding Correction by Small Molecule Therapeutics.

PubMed

Chung, W Joon; Goeckeler-Fried, Jennifer L; Havasi, Viktoria; Chiang, Annette; Rowe, Steven M; Plyler, Zackery E; Hong, Jeong S; Mazur, Marina; Piazza, Gary A; Keeton, Adam B; White, E Lucile; Rasmussen, Lynn; Weissman, Allan M; Denny, R Aldrin; Brodsky, Jeffrey L; Sorscher, Eric J

2016-01-01

Small molecules that correct the folding defects and enhance surface localization of the F508del mutation in the Cystic Fibrosis Transmembrane conductance Regulator (CFTR) comprise an important therapeutic strategy for cystic fibrosis lung disease. However, compounds that rescue the F508del mutant protein to wild type (WT) levels have not been identified. In this report, we consider obstacles to obtaining robust and therapeutically relevant levels of F508del CFTR. For example, markedly diminished steady state amounts of F508del CFTR compared to WT CFTR are present in recombinant bronchial epithelial cell lines, even when much higher levels of mutant transcript are present. In human primary airway cells, the paucity of Band B F508del is even more pronounced, although F508del and WT mRNA concentrations are comparable. Therefore, to augment levels of "repairable" F508del CFTR and identify small molecules that then correct this pool, we developed compound library screening protocols based on automated protein detection. First, cell-based imaging measurements were used to semi-quantitatively estimate distribution of F508del CFTR by high content analysis of two-dimensional images. We evaluated ~2,000 known bioactive compounds from the NIH Roadmap Molecular Libraries Small Molecule Repository in a pilot screen and identified agents that increase the F508del protein pool. Second, we analyzed ~10,000 compounds representing diverse chemical scaffolds for effects on total CFTR expression using a multi-plate fluorescence protocol and describe compounds that promote F508del maturation. Together, our findings demonstrate proof of principle that agents identified in this fashion can augment the level of endoplasmic reticulum (ER) resident "Band B" F508del CFTR suitable for pharmacologic correction. As further evidence in support of this strategy, PYR-41-a compound that inhibits the E1 ubiquitin activating enzyme-was shown to synergistically enhance F508del rescue by C18, a small molecule corrector. Our combined results indicate that increasing the levels of ER-localized CFTR available for repair provides a novel route to correct F508del CFTR.

Evaluation of genotoxicity testing of FDA approved large molecule therapeutics.

PubMed

Sawant, Satin G; Fielden, Mark R; Black, Kurt A

2014-10-01

Large molecule therapeutics (MW>1000daltons) are not expected to enter the cell and thus have reduced potential to interact directly with DNA or related physiological processes. Genotoxicity studies are therefore not relevant and typically not required for large molecule therapeutic candidates. Regulatory guidance supports this approach; however there are examples of marketed large molecule therapeutics where sponsors have conducted genotoxicity studies. A retrospective analysis was performed on genotoxicity studies of United States FDA approved large molecule therapeutics since 1998 identified through the Drugs@FDA website. This information was used to provide a data-driven rationale for genotoxicity evaluations of large molecule therapeutics. Fifty-three of the 99 therapeutics identified were tested for genotoxic potential. None of the therapeutics tested showed a positive outcome in any study except the peptide glucagon (GlucaGen®) showing equivocal in vitro results, as stated in the product labeling. Scientific rationale and data from this review indicate that testing of a majority of large molecule modalities do not add value to risk assessment and support current regulatory guidance. Similarly, the data do not support testing of peptides containing only natural amino acids. Peptides containing non-natural amino acids and small molecules in conjugated products may need to be tested. Copyright © 2014 Elsevier Inc. All rights reserved.

Differential expression and emerging functions of non-coding RNAs in cold adaptation.

PubMed

Frigault, Jacques J; Morin, Mathieu D; Morin, Pier Jr

2017-01-01

Several species undergo substantial physiological and biochemical changes to confront the harsh conditions associated with winter. Small mammalian hibernators and cold-hardy insects are examples of natural models of cold adaptation that have been amply explored. While the molecular picture associated with cold adaptation has started to become clearer in recent years, notably through the use of high-throughput experimental approaches, the underlying cold-associated functions attributed to several non-coding RNAs, including microRNAs (miRNAs) and long non-coding RNAs (lncRNAs), remain to be better characterized. Nevertheless, key pioneering work has provided clues on the likely relevance of these molecules in cold adaptation. With an emphasis on mammalian hibernation and insect cold hardiness, this work first reviews various molecular changes documented so far in these processes. The cascades leading to miRNA and lncRNA production as well as the mechanisms of action of these non-coding RNAs are subsequently described. Finally, we present examples of differentially expressed non-coding RNAs in models of cold adaptation and elaborate on the potential significance of this modulation with respect to low-temperature adaptation.

Manipulating and Visualizing Molecular Interactions in Customized Nanoscale Spaces

NASA Astrophysics Data System (ADS)

Stabile, Francis; Henkin, Gil; Berard, Daniel; Shayegan, Marjan; Leith, Jason; Leslie, Sabrina

We present a dynamically adjustable nanofluidic platform for formatting the conformations of and visualizing the interaction kinetics between biomolecules in solution, offering new time resolution and control of the reaction processes. This platform extends convex lens-induced confinement (CLiC), a technique for imaging molecules under confinement, by introducing a system for in situ modification of the chemical environment; this system uses a deep microchannel to diffusively exchange reagents within the nanoscale imaging region, whose height is fixed by a nanopost array. To illustrate, we visualize and manipulate salt-induced, surfactant-induced, and enzyme-induced reactions between small-molecule reagents and DNA molecules, where the conformations of the DNA molecules are formatted by the imposed nanoscale confinement. By using nanofabricated, nonabsorbing, low-background glass walls to confine biomolecules, our nanofluidic platform facilitates quantitative exploration of physiologically and biotechnologically relevant processes at the nanoscale. This device provides new kinetic information about dynamic chemical processes at the single-molecule level, using advancements in the CLiC design including a microchannel-based diffuser and postarray-based dialysis slit.

A small-molecule fragment that emulates binding of receptor and broadly neutralizing antibodies to influenza A hemagglutinin.

PubMed

Kadam, Rameshwar U; Wilson, Ian A

2018-04-17

The influenza virus hemagglutinin (HA) glycoprotein mediates receptor binding and membrane fusion during viral entry in host cells. Blocking these key steps in viral infection has applications for development of novel antiinfluenza therapeutics as well as vaccines. However, the lack of structural information on how small molecules can gain a foothold in the small, shallow receptor-binding site (RBS) has hindered drug design against this important target on the viral pathogen. Here, we report on the serendipitous crystallization-based discovery of a small-molecule N -cyclohexyltaurine, commonly known as the buffering agent CHES, that is able to bind to both group-1 and group-2 HAs of influenza A viruses. X-ray structural characterization of group-1 H5N1 A/Vietnam/1203/2004 (H5/Viet) and group-2 H3N2 A/Hong Kong/1/1968 (H3/HK68) HAs at 2.0-Å and 2.57-Å resolution, respectively, revealed that N -cyclohexyltaurine binds to the heart of the conserved HA RBS. N -cyclohexyltaurine mimics the binding mode of the natural receptor sialic acid and RBS-targeting bnAbs through formation of similar hydrogen bonds and CH-π interactions with the HA. In H3/HK68, N -cyclohexyltaurine also binds to a conserved pocket in the stem region, thereby exhibiting a dual-binding mode in group-2 HAs. These long-awaited structural insights into RBS recognition by a noncarbohydrate-based small molecule enhance our knowledge of how to target this important functional site and can serve as a template to guide the development of novel broad-spectrum small-molecule therapeutics against influenza virus.

Androgen Receptor Functional Analyses by High Throughput Imaging: Determination of Ligand, Cell Cycle, and Mutation-Specific Effects

PubMed Central

Szafran, Adam T.; Szwarc, Maria; Marcelli, Marco; Mancini, Michael A.

2008-01-01

Background Understanding how androgen receptor (AR) function is modulated by exposure to steroids, growth factors or small molecules can have important mechanistic implications for AR-related disease therapies (e.g., prostate cancer, androgen insensitivity syndrome, AIS), and in the analysis of environmental endocrine disruptors. Methodology/Principal Findings We report the development of a high throughput (HT) image-based assay that quantifies AR subcellular and subnuclear distribution, and transcriptional reporter gene activity on a cell-by-cell basis. Furthermore, simultaneous analysis of DNA content allowed determination of cell cycle position and permitted the analysis of cell cycle dependent changes in AR function in unsynchronized cell populations. Assay quality for EC50 coefficients of variation were 5–24%, with Z' values reaching 0.91. This was achieved by the selective analysis of cells expressing physiological levels of AR, important because minor over-expression resulted in elevated nuclear speckling and decreased transcriptional reporter gene activity. A small screen of AR-binding ligands, including known agonists, antagonists, and endocrine disruptors, demonstrated that nuclear translocation and nuclear “speckling” were linked with transcriptional output, and specific ligands were noted to differentially affect measurements for wild type versus mutant AR, suggesting differing mechanisms of action. HT imaging of patient-derived AIS mutations demonstrated a proof-of-principle personalized medicine approach to rapidly identify ligands capable of restoring multiple AR functions. Conclusions/Significance HT imaging-based multiplex screening will provide a rapid, systems-level analysis of compounds/RNAi that may differentially affect wild type AR or clinically relevant AR mutations. PMID:18978937

Midbond basis functions for weakly bound complexes

NASA Astrophysics Data System (ADS)

Shaw, Robert A.; Hill, J. Grant

2018-06-01

Weakly bound systems present a difficult problem for conventional atom-centred basis sets due to large separations, necessitating the use of large, computationally expensive bases. This can be remedied by placing a small number of functions in the region between molecules in the complex. We present compact sets of optimised midbond functions for a range of complexes involving noble gases, alkali metals and small molecules for use in high accuracy coupled -cluster calculations, along with a more robust procedure for their optimisation. It is shown that excellent results are possible with double-zeta quality orbital basis sets when a few midbond functions are added, improving both the interaction energy and the equilibrium bond lengths of a series of noble gas dimers by 47% and 8%, respectively. When used in conjunction with explicitly correlated methods, near complete basis set limit accuracy is readily achievable at a fraction of the cost that using a large basis would entail. General purpose auxiliary sets are developed to allow explicitly correlated midbond function studies to be carried out, making it feasible to perform very high accuracy calculations on weakly bound complexes.

A density functional theory for association of fluid molecules with a functionalized surface: fluid-wall single and double bonding.

PubMed

Haghmoradi, Amin; Wang, Le; Chapman, Walter G

2017-02-01

In this manuscript we extend Wertheim's two-density formalism beyond its first order to model a system of fluid molecules with a single association site close to a planar hard wall with association sites on its surface in a density functional theory framework. The association sites of the fluid molecules are small enough that they can form only one bond, while the wall association sites are large enough to bond with more than one fluid molecule. The effects of temperature and of bulk fluid and wall site densities on the fluid density profile, extent of association, and competition between single and double bonding of fluid segments at the wall sites versus distance from the wall are presented. The theory predictions are compared with new Monte Carlo simulation results and they are in good agreement. The theory captures the surface coverage over wide ranges of temperature and bulk density by introducing the effect of steric hindrance in fluid association at a wall site.

Urea transporter proteins as targets for small-molecule diuretics.

PubMed

Esteva-Font, Cristina; Anderson, Marc O; Verkman, Alan S

2015-02-01

Conventional diuretics such as furosemide and thiazides target salt transporters in kidney tubules, but urea transporters (UTs) have emerged as alternative targets. UTs are a family of transmembrane channels expressed in a variety of mammalian tissues, in particular the kidney. UT knockout mice and humans with UT mutations exhibit reduced maximal urinary osmolality, demonstrating that UTs are necessary for the concentration of urine. Small-molecule screening has identified potent and selective inhibitors of UT-A, the UT protein expressed in renal tubule epithelial cells, and UT-B, the UT protein expressed in vasa recta endothelial cells. Data from UT knockout mice and from rodents administered UT inhibitors support the diuretic action of UT inhibition. The kidney-specific expression of UT-A1, together with high selectivity of the small-molecule inhibitors, means that off-target effects of such small-molecule drugs should be minimal. This Review summarizes the structure, expression and function of UTs, and looks at the evidence supporting the validity of UTs as targets for the development of salt-sparing diuretics with a unique mechanism of action. UT-targeted inhibitors may be useful alone or in combination with conventional diuretics for therapy of various oedemas and hyponatraemias, potentially including those refractory to treatment with current diuretics.

AM-37 and ST-36 Are Small Molecule Bombesin Receptor Antagonists

PubMed Central

Moody, Terry W.; Tashakkori, Nicole; Mantey, Samuel A.; Moreno, Paola; Ramos-Alvarez, Irene; Leopoldo, Marcello; Jensen, Robert T.

2017-01-01

While peptide antagonists for the gastrin-releasing peptide receptor (BB2R), neuromedin B receptor (BB1R), and bombesin (BB) receptor subtype-3 (BRS-3) exist, there is a need to develop non-peptide small molecule inhibitors for all three BBR. The BB agonist (BA)1 binds with high affinity to the BB1R, BB2R, and BRS-3. In this communication, small molecule BBR antagonists were evaluated using human lung cancer cells. AM-37 and ST-36 inhibited binding to human BB1R, BB2R, and BRS-3 with similar affinity (Ki = 1.4–10.8 µM). AM-13 and AM-14 were approximately an order of magnitude less potent than AM-37 and ST-36. The ability of BA1 to elevate cytosolic Ca2+ in human lung cancer cells transfected with BB1R, BB2R, and BRS-3 was antagonized by AM-37 and ST-36. BA1 increased tyrosine phosphorylation of the EGFR and ERK in lung cancer cells, which was blocked by AM-37 and ST-36. AM-37 and ST-36 reduced the growth of lung cancer cells that have BBR. The results indicate that AM-37 and ST-36 function as small molecule BB receptor antagonists. PMID:28785244

AM-37 and ST-36 Are Small Molecule Bombesin Receptor Antagonists.

PubMed

Moody, Terry W; Tashakkori, Nicole; Mantey, Samuel A; Moreno, Paola; Ramos-Alvarez, Irene; Leopoldo, Marcello; Jensen, Robert T

2017-01-01

While peptide antagonists for the gastrin-releasing peptide receptor (BB 2 R), neuromedin B receptor (BB 1 R), and bombesin (BB) receptor subtype-3 (BRS-3) exist, there is a need to develop non-peptide small molecule inhibitors for all three BBR. The BB agonist (BA)1 binds with high affinity to the BB 1 R, BB 2 R, and BRS-3. In this communication, small molecule BBR antagonists were evaluated using human lung cancer cells. AM-37 and ST-36 inhibited binding to human BB 1 R, BB 2 R, and BRS-3 with similar affinity ( K i = 1.4-10.8 µM). AM-13 and AM-14 were approximately an order of magnitude less potent than AM-37 and ST-36. The ability of BA1 to elevate cytosolic Ca 2+ in human lung cancer cells transfected with BB 1 R, BB 2 R, and BRS-3 was antagonized by AM-37 and ST-36. BA1 increased tyrosine phosphorylation of the EGFR and ERK in lung cancer cells, which was blocked by AM-37 and ST-36. AM-37 and ST-36 reduced the growth of lung cancer cells that have BBR. The results indicate that AM-37 and ST-36 function as small molecule BB receptor antagonists.

Near-membrane electric field calcium ion dehydration.

PubMed

Barger, James P; Dillon, Patrick F

2016-12-01

The dehydration of ion-water complexes prior to ion channel transit has focused on channel protein-mediated dissociation of water. Ion dehydration by the membrane electric field has not previously been considered. Near membrane electric fields have previously been shown to cause the disassociation of non-covalently bound small molecule-small molecule, small molecule-protein, and protein-protein complexes. It is well known that cosmotropic, structure making ions such as calcium and sodium significantly bind multiple water ions in solution. It is also known that these ions are often not hydrated as they pass through membrane ion channels. Using capillary electrophoresis, the range of electric fields needed to strip water molecules from calcium ions has been measured. Ion migration velocity is a linear function of the electric field. At low electric fields, the migration rate of calcium ion was shown to be linearly related to the applied electric field. Using a form of the Stoke's equation applicable to ion migration, the hydrated calcium radius was found to be 0.334nm, corresponding to a water hydration shell of 5.09 water molecules. At higher electric fields, the slope of the calcium migration velocity as a function of the electric field increased, which was modeled as a decrease in the radius of the migrating ion as the water was removed. Using a tanh function to model the transition of the ion from a hydrated to a stripped state, the transition had a midpoint at 446V/cm, and was 88% complete at 587V/cm with a correlation coefficient of 0.9996. The migration velocity of the stripped calcium ion was found to be a function of both the decrease in radius and an increase in the effective, electronic viscosity of the dipole medium through which the dehydrated ion moved. The size of the electric field needed to dehydrate calcium occurs 6-7nm from the cell membrane. Calcium ions within this distance from the membrane will be devoid of water molecules when they reach the calcium selective channel pore entrances, all known to be approximately 1-2nm from the membrane. No matter what the calcium pore structure, calcium ions reaching the channel entrance will be devoid of a water shell. Copyright © 2016 Elsevier Ltd. All rights reserved.

Activation of the stress proteome as a mechanism for small molecule therapeutics.

PubMed

Brose, Rebecca Deering; Shin, Gloria; McGuinness, Martina C; Schneidereith, Tonya; Purvis, Shirley; Dong, Gao X; Keefer, Jeffrey; Spencer, Forrest; Smith, Kirby D

2012-10-01

Various small molecule pharmacologic agents with different known functions produce similar outcomes in diverse Mendelian and complex disorders, suggesting that they may induce common cellular effects. These molecules include histone deacetylase inhibitors, 4-phenylbutyrate (4PBA) and trichostatin A, and two small molecules without direct histone deacetylase inhibitor activity, hydroxyurea (HU) and sulforaphane. In some cases, the therapeutic effects of histone deacetylase inhibitors have been attributed to an increase in expression of genes related to the disease-causing gene. However, here we show that the pharmacological induction of mitochondrial biogenesis was necessary for the potentially therapeutic effects of 4PBA or HU in two distinct disease models, X-linked adrenoleukodystrophy and sickle cell disease. We hypothesized that a common cellular response to these four molecules is induction of mitochondrial biogenesis and peroxisome proliferation and activation of the stress proteome, or adaptive cell survival response. Treatment of human fibroblasts with these four agents induced mitochondrial and peroxisomal biogenesis as monitored by flow cytometry, immunofluorescence and/or western analyses. In treated normal human fibroblasts, all four agents induced the adaptive cell survival response: heat shock, unfolded protein, autophagic and antioxidant responses and the c-jun N-terminal kinase pathway, at the transcriptional and translational levels. Thus, activation of the evolutionarily conserved stress proteome and mitochondrial biogenesis may be a common cellular response to such small molecule therapy and a common basis of therapeutic action in various diseases. Modulation of this novel therapeutic target could broaden the range of treatable diseases without directly targeting the causative genetic abnormalities.

Activation of the stress proteome as a mechanism for small molecule therapeutics

PubMed Central

Brose, Rebecca Deering; Shin, Gloria; McGuinness, Martina C.; Schneidereith, Tonya; Purvis, Shirley; Dong, Gao X.; Keefer, Jeffrey; Spencer, Forrest; Smith, Kirby D.

2012-01-01

Various small molecule pharmacologic agents with different known functions produce similar outcomes in diverse Mendelian and complex disorders, suggesting that they may induce common cellular effects. These molecules include histone deacetylase inhibitors, 4-phenylbutyrate (4PBA) and trichostatin A, and two small molecules without direct histone deacetylase inhibitor activity, hydroxyurea (HU) and sulforaphane. In some cases, the therapeutic effects of histone deacetylase inhibitors have been attributed to an increase in expression of genes related to the disease-causing gene. However, here we show that the pharmacological induction of mitochondrial biogenesis was necessary for the potentially therapeutic effects of 4PBA or HU in two distinct disease models, X-linked adrenoleukodystrophy and sickle cell disease. We hypothesized that a common cellular response to these four molecules is induction of mitochondrial biogenesis and peroxisome proliferation and activation of the stress proteome, or adaptive cell survival response. Treatment of human fibroblasts with these four agents induced mitochondrial and peroxisomal biogenesis as monitored by flow cytometry, immunofluorescence and/or western analyses. In treated normal human fibroblasts, all four agents induced the adaptive cell survival response: heat shock, unfolded protein, autophagic and antioxidant responses and the c-jun N-terminal kinase pathway, at the transcriptional and translational levels. Thus, activation of the evolutionarily conserved stress proteome and mitochondrial biogenesis may be a common cellular response to such small molecule therapy and a common basis of therapeutic action in various diseases. Modulation of this novel therapeutic target could broaden the range of treatable diseases without directly targeting the causative genetic abnormalities. PMID:22752410

High-throughput screening of small molecules in miniaturized mammalian cell-based assays involving post-translational modifications.

PubMed

Stockwell, B R; Haggarty, S J; Schreiber, S L

1999-02-01

Fully adapting a forward genetic approach to mammalian systems requires efficient methods to alter systematically gene products without prior knowledge of gene sequences, while allowing for the subsequent characterization of these alterations. Ideally, these methods would also allow function to be altered in a temporally controlled manner. We report the development of a miniaturized cell-based assay format that enables a genetic-like approach to understanding cellular pathways in mammalian systems using small molecules, rather than mutations, as the source of gene-product alterations. This whole-cell immunodetection assay can sensitively detect changes in specific cellular macromolecules in high-density arrays of mammalian cells. Furthermore, it is compatible with screening large numbers of small molecules in nanoliter to microliter culture volumes. We refer to this assay format as a 'cytoblot', and demonstrate the use of cytoblotting to monitor biosynthetic processes such as DNA synthesis, and post-translational processes such as acetylation and phosphorylation. Finally, we demonstrate the applicability of these assays to natural-product screening through the identification of marine sponge extracts exhibiting genotype-specific inhibition of 5-bromodeoxyuridine incorporation and suppression of the anti-proliferative effect of rapamycin. We show that cytoblots can be used for high-throughput screening of small molecules in cell-based assays. Together with small-molecule libraries, the cytoblot assay can be used to perform chemical genetic screens analogous to those used in classical genetics and thus should be applicable to understanding a wide variety of cellular processes, especially those involving post-transitional modifications.

In Silico Analysis of the Small Molecule Content of Outer Membrane Vesicles Produced by Bacteroides thetaiotaomicron Indicates an Extensive Metabolic Link between Microbe and Host

PubMed Central

Bryant, William A.; Stentz, Régis; Le Gall, Gwenaelle; Sternberg, Michael J. E.; Carding, Simon R.; Wilhelm, Thomas

2017-01-01

The interactions between the gut microbiota and its host are of central importance to the health of the host. Outer membrane vesicles (OMVs) are produced ubiquitously by Gram-negative bacteria including the gut commensal Bacteroides thetaiotaomicron. These vesicles can interact with the host in various ways but until now their complement of small molecules has not been investigated in this context. Using an untargeted high-coverage metabolomic approach we have measured the small molecule content of these vesicles in contrasting in vitro conditions to establish what role these metabolites could perform when packed into these vesicles. B. thetaiotaomicron packs OMVs with a highly conserved core set of small molecules which are strikingly enriched with mouse-digestible metabolites and with metabolites previously shown to be associated with colonization of the murine GIT. By use of an expanded genome-scale metabolic model of B. thetaiotaomicron and a potential host (the mouse) we have established many possible metabolic pathways between the two organisms that were previously unknown, and have found several putative novel metabolic functions for mouse that are supported by gene annotations, but that do not currently appear in existing mouse metabolic networks. The lipidome of these OMVs bears no relation to the mouse lipidome, so the purpose of this particular composition of lipids remains unclear. We conclude from this analysis that through intimate symbiotic evolution OMVs produced by B. thetaiotaomicron are likely to have been adopted as a conduit for small molecules bound for the mammalian host in vivo. PMID:29276507

In Silico Analysis of the Small Molecule Content of Outer Membrane Vesicles Produced by Bacteroides thetaiotaomicron Indicates an Extensive Metabolic Link between Microbe and Host.

PubMed

Bryant, William A; Stentz, Régis; Le Gall, Gwenaelle; Sternberg, Michael J E; Carding, Simon R; Wilhelm, Thomas

2017-01-01

The interactions between the gut microbiota and its host are of central importance to the health of the host. Outer membrane vesicles (OMVs) are produced ubiquitously by Gram-negative bacteria including the gut commensal Bacteroides thetaiotaomicron . These vesicles can interact with the host in various ways but until now their complement of small molecules has not been investigated in this context. Using an untargeted high-coverage metabolomic approach we have measured the small molecule content of these vesicles in contrasting in vitro conditions to establish what role these metabolites could perform when packed into these vesicles. B. thetaiotaomicron packs OMVs with a highly conserved core set of small molecules which are strikingly enriched with mouse-digestible metabolites and with metabolites previously shown to be associated with colonization of the murine GIT. By use of an expanded genome-scale metabolic model of B. thetaiotaomicron and a potential host (the mouse) we have established many possible metabolic pathways between the two organisms that were previously unknown, and have found several putative novel metabolic functions for mouse that are supported by gene annotations, but that do not currently appear in existing mouse metabolic networks. The lipidome of these OMVs bears no relation to the mouse lipidome, so the purpose of this particular composition of lipids remains unclear. We conclude from this analysis that through intimate symbiotic evolution OMVs produced by B. thetaiotaomicron are likely to have been adopted as a conduit for small molecules bound for the mammalian host in vivo .

HC StratoMineR: A Web-Based Tool for the Rapid Analysis of High-Content Datasets.

PubMed

Omta, Wienand A; van Heesbeen, Roy G; Pagliero, Romina J; van der Velden, Lieke M; Lelieveld, Daphne; Nellen, Mehdi; Kramer, Maik; Yeong, Marley; Saeidi, Amir M; Medema, Rene H; Spruit, Marco; Brinkkemper, Sjaak; Klumperman, Judith; Egan, David A

2016-10-01

High-content screening (HCS) can generate large multidimensional datasets and when aligned with the appropriate data mining tools, it can yield valuable insights into the mechanism of action of bioactive molecules. However, easy-to-use data mining tools are not widely available, with the result that these datasets are frequently underutilized. Here, we present HC StratoMineR, a web-based tool for high-content data analysis. It is a decision-supportive platform that guides even non-expert users through a high-content data analysis workflow. HC StratoMineR is built by using My Structured Query Language for storage and querying, PHP: Hypertext Preprocessor as the main programming language, and jQuery for additional user interface functionality. R is used for statistical calculations, logic and data visualizations. Furthermore, C++ and graphical processor unit power is diffusely embedded in R by using the rcpp and rpud libraries for operations that are computationally highly intensive. We show that we can use HC StratoMineR for the analysis of multivariate data from a high-content siRNA knock-down screen and a small-molecule screen. It can be used to rapidly filter out undesirable data; to select relevant data; and to perform quality control, data reduction, data exploration, morphological hit picking, and data clustering. Our results demonstrate that HC StratoMineR can be used to functionally categorize HCS hits and, thus, provide valuable information for hit prioritization.

Therapeutic potential of abalone and status of bioactive molecules: A comprehensive review.

PubMed

Suleria, H A R; Masci, P P; Gobe, G C; Osborne, S A

2017-05-24

Marine organisms are increasingly being investigated as sources of bioactive molecules with therapeutic applications as nutraceuticals and pharmaceuticals. In particular, nutraceuticals are gaining popularity worldwide owing to their therapeutic potential and incorporation in functional foods and dietary supplements. Abalone, a marine gastropod, contains a variety of bioactive compounds with anti-oxidant, anti-thrombotic, anti-inflammatory, anti-microbial, and anti-cancer activities. For thousands of years different cultures have used abalone as a traditional functional food believing consumption provides health benefits. Abalone meat is one of the most precious commodities in Asian markets where it is considered a culinary delicacy. Recent research has revealed that abalone is composed of many vital moieties like polysaccharides, proteins, and fatty acids that provide health benefits beyond basic nutrition. A review of past and present research is presented with relevance to the therapeutic potential of bioactive molecules from abalone.

Regioselective synthesis of C3 alkylated and arylated benzothiophenes

NASA Astrophysics Data System (ADS)

Shrives, Harry J.; Fernández-Salas, José A.; Hedtke, Christin; Pulis, Alexander P.; Procter, David J.

2017-03-01

Benzothiophenes are heterocyclic constituents of important molecules relevant to society, including those with the potential to meet modern medical challenges. The construction of molecules would be vastly more efficient if carbon-hydrogen bonds, found in all organic molecules, can be directly converted into carbon-carbon bonds. In the case of elaborating benzothiophenes, functionalization of carbon-hydrogen bonds at carbon-number 3 (C3) is markedly more demanding than at C2 due to issues of regioselectivity (C3 versus C2), and the requirement of high temperatures, precious metals and the installation of superfluous directing groups. Herein, we demonstrate that synthetically unexplored but readily accessible benzothiophene S-oxides serve as novel precursors for C3-functionalized benzothiophenes. Employing an interrupted Pummerer reaction to capture and then deliver phenol and silane coupling partners, we have discovered a directing group-free method that delivers C3-arylated and -alkylated benzothiophenes with complete regioselectivity, under metal-free and mild conditions.

Graphane versus graphene: a computational investigation of the interaction of nucleobases, aminoacids, heterocycles, small molecules (CO2, H2O, NH3, CH4, H2), metal ions and onium ions.

PubMed

Umadevi, Deivasigamani; Narahari Sastry, G

2015-11-11

Graphane has emerged as a two-dimensional hydrocarbon with interesting physical properties and potential applications. Understanding the interaction of graphane with various molecules and ions is crucial to appreciate its potential applications. We investigated the interaction of nucleobases, aminoacids, saturated and unsaturated heterocycles, small molecules, metal ions and onium ions with graphane by using density functional theory calculations. The preferred orientations of these molecules and ions on the graphane surface have been analysed. The binding energies of graphane with these molecules have been compared with the corresponding binding energies of graphene. Our results reveal that graphane forms stable complexes with all the molecules and ions yet showing lesser binding affinity when compared to graphene. As an exemption, the preferential strong binding of H2O with graphane than graphene reveals the fact that graphane is more hydrophilic than graphene. Charge transfer between graphane and the molecules and ions have been found to be an important factor in determining the binding strength of the complexes. The effect of the interaction of these molecules and ions on the HOMO-LUMO energy gap of graphane has also been investigated.

Cost-effective differentiation of hepatocyte-like cells from human pluripotent stem cells using small molecules.

PubMed

Tasnim, Farah; Phan, Derek; Toh, Yi-Chin; Yu, Hanry

2015-11-01

Significant efforts have been invested into the differentiation of stem cells into functional hepatocyte-like cells that can be used for cell therapy, disease modeling and drug screening. Most of these efforts have been concentrated on the use of growth factors to recapitulate developmental signals under in vitro conditions. Using small molecules instead of growth factors would provide an attractive alternative since small molecules are cell-permeable and cheaper than growth factors. We have developed a protocol for the differentiation of human embryonic stem cells into hepatocyte-like cells using a predominantly small molecule-based approach (SM-Hep). This 3 step differentiation strategy involves the use of optimized concentrations of LY294002 and bromo-indirubin-3'-oxime (BIO) for the generation of definitive endoderm; sodium butyrate and dimethyl sulfoxide (DMSO) for the generation of hepatoblasts and SB431542 for differentiation into hepatocyte-like cells. Activin A is the only growth factor required in this protocol. Our results showed that SM-Hep were morphologically and functionally similar or better compared to the hepatocytes derived from the growth-factor induced differentiation (GF-Hep) in terms of expression of hepatic markers, urea and albumin production and cytochrome P450 (CYP1A2 and CYP3A4) activities. Cell viability assays following treatment with paradigm hepatotoxicants Acetaminophen, Chlorpromazine, Diclofenac, Digoxin, Quinidine and Troglitazone showed that their sensitivity to these drugs was similar to human primary hepatocytes (PHHs). Using SM-Hep would result in 67% and 81% cost reduction compared to GF-Hep and PHHs respectively. Therefore, SM-Hep can serve as a robust and cost effective replacement for PHHs for drug screening and development. Copyright © 2015 Elsevier Ltd. All rights reserved.

Molecular Probing of the HPV-16 E6 Protein Alpha Helix Binding Groove with Small Molecule Inhibitors

PubMed Central

Rietz, Anne; Petrov, Dino P.; Bartolowits, Matthew; DeSmet, Marsha; Davisson, V. Jo; Androphy, Elliot J.

2016-01-01

The human papillomavirus (HPV) HPV E6 protein has emerged as a central oncoprotein in HPV-associated cancers in which sustained expression is required for tumor progression. A majority of the E6 protein interactions within the human proteome use an alpha-helix groove interface for binding. The UBE3A/E6AP HECT domain ubiquitin ligase binds E6 at this helix-groove interface. This enables formation of a trimeric complex with p53, resulting in destruction of this tumor suppressor. While recent x-ray crystal structures are useful, examples of small molecule probes that can modulate protein interactions at this interface are limited. To develop insights useful for potential structure-based design of ligands for HPV E6, a series of 2,6-disubstituted benzopyranones were prepared and tested as competitive antagonists of E6-E6AP helix-groove interactions. These small molecule probes were used in both binding and functional assays to evaluate recognition features of the E6 protein. Evidence for an ionic functional group interaction within the helix groove was implicated by the structure-activity among the highest affinity ligands. The molecular topographies of these protein-ligand interactions were evaluated by comparing the binding and activities of single amino acid E6 mutants with the results of molecular dynamic simulations. A group of arginine residues that form a rim-cap over the E6 helix groove offer compensatory roles in binding and recognition of the small molecule probes. The flexibility and impact on the overall helix-groove shape dictated by these residues offer new insights for structure-based targeting of HPV E6. PMID:26915086

Targeting of the MYCN Protein with Small Molecule c-MYC Inhibitors

PubMed Central

Müller, Inga; Larsson, Karin; Frenzel, Anna; Oliynyk, Ganna; Zirath, Hanna; Prochownik, Edward V.; Westwood, Nicholas J.; Henriksson, Marie Arsenian

2014-01-01

Members of the MYC family are the most frequently deregulated oncogenes in human cancer and are often correlated with aggressive disease and/or poorly differentiated tumors. Since patients with MYCN-amplified neuroblastoma have a poor prognosis, targeting MYCN using small molecule inhibitors could represent a promising therapeutic approach. We have previously demonstrated that the small molecule 10058-F4, known to bind to the c-MYC bHLHZip dimerization domain and inhibiting the c-MYC/MAX interaction, also interferes with the MYCN/MAX dimerization in vitro and imparts anti-tumorigenic effects in neuroblastoma tumor models with MYCN overexpression. Our previous work also revealed that MYCN-inhibition leads to mitochondrial dysfunction resulting in accumulation of lipid droplets in neuroblastoma cells. To expand our understanding of how small molecules interfere with MYCN, we have now analyzed the direct binding of 10058-F4, as well as three of its analogs; #474, #764 and 10058-F4(7RH), one metabolite C-m/z 232, and a structurally unrelated c-MYC inhibitor 10074-G5, to the bHLHZip domain of MYCN. We also assessed their ability to induce apoptosis, neurite outgrowth and lipid accumulation in neuroblastoma cells. Interestingly, all c-MYC binding molecules tested also bind MYCN as assayed by surface plasmon resonance. Using a proximity ligation assay, we found reduced interaction between MYCN and MAX after treatment with all molecules except for the 10058-F4 metabolite C-m/z 232 and the non-binder 10058-F4(7RH). Importantly, 10074-G5 and 10058-F4 were the most efficient in inducing neuronal differentiation and lipid accumulation in MYCN-amplified neuroblastoma cells. Together our data demonstrate MYCN-binding properties for a selection of small molecules, and provide functional information that could be of importance for future development of targeted therapies against MYCN-amplified neuroblastoma. PMID:24859015

On-Demand Targeting: Investigating Biology with Proximity-Directed Chemistry

PubMed Central

2016-01-01

Proximity enhancement is a central chemical tenet underpinning an exciting suite of small-molecule toolsets that have allowed us to unravel many biological complexities. The leitmotif of this opus is “tethering”—a strategy in which a multifunctional small molecule serves as a template to bring proteins/biomolecules together. Scaffolding approaches have been powerfully applied to control diverse biological outcomes such as protein–protein association, protein stability, activity, and improve imaging capabilities. A new twist on this strategy has recently appeared, in which the small-molecule probe is engineered to unleash controlled amounts of reactive chemical signals within the microenvironment of a target protein. Modification of a specific target elicits a precisely timed and spatially controlled gain-of-function (or dominant loss-of-function) signaling response. Presented herein is a unique personal outlook conceptualizing the powerful proximity-enhanced chemical biology toolsets into two paradigms: “multifunctional scaffolding” versus “on-demand targeting”. By addressing the latest advances and challenges in the established yet constantly evolving multifunctional scaffolding strategies as well as in the emerging on-demand precision targeting (and related) systems, this Perspective is aimed at choosing when it is best to employ each of the two strategies, with an emphasis toward further promoting novel applications and discoveries stemming from these innovative chemical biology platforms. PMID:26907082

On-Demand Targeting: Investigating Biology with Proximity-Directed Chemistry.

PubMed

Long, Marcus J C; Poganik, Jesse R; Aye, Yimon

2016-03-23

Proximity enhancement is a central chemical tenet underpinning an exciting suite of small-molecule toolsets that have allowed us to unravel many biological complexities. The leitmotif of this opus is "tethering"-a strategy in which a multifunctional small molecule serves as a template to bring proteins/biomolecules together. Scaffolding approaches have been powerfully applied to control diverse biological outcomes such as protein-protein association, protein stability, activity, and improve imaging capabilities. A new twist on this strategy has recently appeared, in which the small-molecule probe is engineered to unleash controlled amounts of reactive chemical signals within the microenvironment of a target protein. Modification of a specific target elicits a precisely timed and spatially controlled gain-of-function (or dominant loss-of-function) signaling response. Presented herein is a unique personal outlook conceptualizing the powerful proximity-enhanced chemical biology toolsets into two paradigms: "multifunctional scaffolding" versus "on-demand targeting". By addressing the latest advances and challenges in the established yet constantly evolving multifunctional scaffolding strategies as well as in the emerging on-demand precision targeting (and related) systems, this Perspective is aimed at choosing when it is best to employ each of the two strategies, with an emphasis toward further promoting novel applications and discoveries stemming from these innovative chemical biology platforms.

Promiscuous Actions of Small Molecule Inhibitors of the Protein Kinase D-Class IIa HDAC Axis in Striated Muscle

PubMed Central

Lemon, Douglas D.; Harrison, Brooke C.; Horn, Todd R.; Stratton, Matthew S.; Ferguson, Bradley S.; Wempe, Michael F.; McKinsey, Timothy A.

2015-01-01

PKD-mediated phosphorylation of class IIa HDACs frees the MEF2 transcription factor to activate genes that govern muscle differentiation and growth. Studies of the regulation and function of this signaling axis have involved MC1568 and Gö-6976, which are small molecule inhibitors of class IIa HDAC and PKD catalytic activity, respectively. We describe unanticipated effects of these compounds. MC1568 failed to inhibit class IIa HDAC catalytic activity in vitro, and exerted divergent effects on skeletal muscle differentiation compared to a bona fide inhibitor of these HDACs. In cardiomyocytes, Gö-6976 triggered calcium signaling and activated stress-inducible kinases. Based on these findings, caution is warranted when employing MC1568 and Gö-6976 as pharmacological tool compounds to assess functions of class IIa HDACs and PKD. PMID:25816750

Ras activation by SOS: Allosteric regulation by altered fluctuation dynamics

PubMed Central

Iversen, Lars; Tu, Hsiung-Lin; Lin, Wan-Chen; Christensen, Sune M.; Abel, Steven M.; Iwig, Jeff; Wu, Hung-Jen; Gureasko, Jodi; Rhodes, Christopher; Petit, Rebecca S.; Hansen, Scott D.; Thill, Peter; Yu, Cheng-Han; Stamou, Dimitrios; Chakraborty, Arup K.; Kuriyan, John; Groves, Jay T.

2014-01-01

Activation of the small guanosine triphosphatase H-Ras by the exchange factor Son of Sevenless (SOS) is an important hub for signal transduction. Multiple layers of regulation, through protein and membrane interactions, govern activity of SOS. We characterized the specific activity of individual SOS molecules catalyzing nucleotide exchange in H-Ras. Single-molecule kinetic traces revealed that SOS samples a broad distribution of turnover rates through stochastic fluctuations between distinct, long-lived (more than 100 seconds), functional states. The expected allosteric activation of SOS by Ras–guanosine triphosphate (GTP) was conspicuously absent in the mean rate. However, fluctuations into highly active states were modulated by Ras-GTP. This reveals a mechanism in which functional output may be determined by the dynamical spectrum of rates sampled by a small number of enzymes, rather than the ensemble average. PMID:24994643

Small Molecule Ligands of Methyl-Lysine Binding Proteins

PubMed Central

Herold, J. Martin; Wigle, Tim J.; Norris, Jacqueline L.; Lam, Robert; Korboukh, Victoria K.; Gao, Cen; Ingerman, Lindsey A.; Kireev, Dmitri B.; Senisterra, Guillermo; Vedadi, Masoud; Tripathy, Ashutosh; Brown, Peter J.; Arrowsmith, Cheryl H.; Jin, Jian; Janzen, William P.; Frye, Stephen V.

2011-01-01

Proteins which bind methylated lysines (“readers” of the histone code) are important components in the epigenetic regulation of gene expression and can also modulate other proteins that contain methyl-lysine such as p53 and Rb. Recognition of methyl-lysine marks by MBT domains leads to compaction of chromatin and a repressed transcriptional state. Antagonists of MBT domains would serve as probes to interrogate the functional role of these proteins and initiate the chemical biology of methyl-lysine readers as a target class. Small molecule MBT antagonists were designed based on the structure of histone peptide-MBT complexes and their interaction with MBT domains determined using a chemiluminescent assay and ITC. The ligands discovered antagonize native histone peptide binding, exhibiting 5-fold stronger binding affinity to L3MBTL1 than its preferred histone peptide. The first co-crystal structure of a small molecule bound to L3MBTL1 was determined and provides new insights into binding requirements for further ligand design. PMID:21417280

Probing a 2-aminobenzimidazole library for binding to RNA internal loops via two-dimensional combinatorial screening.

PubMed

Velagapudi, Sai Pradeep; Pushechnikov, Alexei; Labuda, Lucas P; French, Jonathan M; Disney, Matthew D

2012-11-16

There are many potential RNA drug targets in bacterial, viral, and human transcriptomes. However, there are few small molecules that modulate RNA function. This is due, in part, to a lack of fundamental understanding about RNA-ligand interactions including the types of small molecules that bind to RNA structural elements and the RNA structural elements that bind to small molecules. In an effort to better understand RNA-ligand interactions, we diversified the 2-aminobenzimidazole core (2AB) and probed the resulting library for binding to a library of RNA internal loops. We chose the 2AB core for these studies because it is a privileged scaffold for binding RNA based on previous reports. These studies identified that N-methyl pyrrolidine, imidazole, and propylamine diversity elements at the R1 position increase binding to internal loops; variability at the R2 position is well tolerated. The preferred RNA loop space was also determined for five ligands using a statistical approach and identified trends that lead to selective recognition.

Single molecule optical measurements of orientation and rotations of biological macromolecules

PubMed Central

Shroder, Deborah Y; Lippert, Lisa G; Goldman, Yale E

2016-01-01

The subdomains of macromolecules often undergo large orientation changes during their catalytic cycles that are essential for their activity. Tracking these rearrangements in real time opens a powerful window into the link between protein structure and functional output. Site-specific labeling of individual molecules with polarized optical probes and measuring their spatial orientation can give insight into the crucial conformational changes, dynamics, and fluctuations of macromolecules. Here we describe the range of single molecule optical technologies that can extract orientation information from these probes, we review the relevant types of probes and labeling techniques, and we highlight the advantages and disadvantages of these technologies for addressing specific inquiries. PMID:28192292

The condensation and vaporization behavior of ices containing SO2, H2S, and CO2 - Implications for Io

NASA Astrophysics Data System (ADS)

Sandford, Scott A.; Allamandola, Louis J.

1993-12-01

The present compilation of measurements of the physical and IR spectral properties of ices whose molecular compositions are relevant to the case of Io encompasses ice systems containing SO2, H2S, and CO2. Surface-binding energies used to calculate the residence times of molecules on a surface as a function of temperature furnish crucially important parameters for models attending to the transport of such molecules to the surface of Io. The values thus derived show that SO2 frosts anneal rapidly.

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

Ren, Chunyan; Morohashi, Keita; Plotnikov, Alexander N.

Chromobox homolog 7 (CBX7) plays an important role in gene transcription in a wide array of cellular processes, ranging from stem cell self-renewal and differentiation to tumor progression. CBX7 functions through its N-terminal chromodomain (ChD), which recognizes tri-methylated lysine 27 of histone 3 (H3K27me3), a conserved epigenetic mark that signifies gene transcriptional repression. Here in this study, we report discovery of small molecules that inhibit CBX7ChD binding to H3K27me3. Our crystal structures reveal the binding modes of these molecules that compete against H3K27me3 binding through interactions with key residues in the methyl-lysine binding pocket of CBX7ChD. We further show thatmore » a lead compound MS37452, derepresses transcription of Polycomb repressive complex target gene p16/CDKN2A by displacing CBX7 binding to the INK4A/ARF locus in prostate cancer cells. Ultimately, these small molecules have the potential to be developed into high-potency chemical modulators that target CBX7 functions in gene transcription in different disease pathways.« less

Systematic mapping of contact sites reveals tethers and a function for the peroxisome-mitochondria contact.

PubMed

Shai, Nadav; Yifrach, Eden; van Roermund, Carlo W T; Cohen, Nir; Bibi, Chen; IJlst, Lodewijk; Cavellini, Laetitia; Meurisse, Julie; Schuster, Ramona; Zada, Lior; Mari, Muriel C; Reggiori, Fulvio M; Hughes, Adam L; Escobar-Henriques, Mafalda; Cohen, Mickael M; Waterham, Hans R; Wanders, Ronald J A; Schuldiner, Maya; Zalckvar, Einat

2018-05-02

The understanding that organelles are not floating in the cytosol, but rather held in an organized yet dynamic interplay through membrane contact sites, is altering the way we grasp cell biological phenomena. However, we still have not identified the entire repertoire of contact sites, their tethering molecules and functions. To systematically characterize contact sites and their tethering molecules here we employ a proximity detection method based on split fluorophores and discover four potential new yeast contact sites. We then focus on a little-studied yet highly disease-relevant contact, the Peroxisome-Mitochondria (PerMit) proximity, and uncover and characterize two tether proteins: Fzo1 and Pex34. We genetically expand the PerMit contact site and demonstrate a physiological function in β-oxidation of fatty acids. Our work showcases how systematic analysis of contact site machinery and functions can deepen our understanding of these structures in health and disease.

Antivirulence Activity of the Human Gut Metabolome

PubMed Central

Antunes, L. Caetano M.; McDonald, Julie A. K.; Schroeter, Kathleen; Carlucci, Christian; Ferreira, Rosana B. R.; Wang, Melody; Yurist-Doutsch, Sophie; Hira, Gill; Jacobson, Kevan; Davies, Julian; Allen-Vercoe, Emma

2014-01-01

ABSTRACT The mammalian gut contains a complex assembly of commensal microbes termed microbiota. Although much has been learned about the role of these microbes in health, the mechanisms underlying these functions are ill defined. We have recently shown that the mammalian gut contains thousands of small molecules, most of which are currently unidentified. Therefore, we hypothesized that these molecules function as chemical cues used by hosts and microbes during their interactions in health and disease. Thus, a search was initiated to identify molecules produced by the microbiota that are sensed by pathogens. We found that a secreted molecule produced by clostridia acts as a strong repressor of Salmonella virulence, obliterating expression of the Salmonella pathogenicity island 1 as well as host cell invasion. It has been known for decades that the microbiota protects its hosts from invading pathogens, and these data suggest that chemical sensing may be involved in this phenomenon. Further investigations should reveal the exact biological role of this molecule as well as its therapeutic potential. PMID:25073640

'Systems toxicology' approach identifies coordinated metabolic responses to copper in a terrestrial non-model invertebrate, the earthworm Lumbricus rubellus.

PubMed

Bundy, Jacob G; Sidhu, Jasmin K; Rana, Faisal; Spurgeon, David J; Svendsen, Claus; Wren, Jodie F; Stürzenbaum, Stephen R; Morgan, A John; Kille, Peter

2008-06-03

New methods are needed for research into non-model organisms, to monitor the effects of toxic disruption at both the molecular and functional organism level. We exposed earthworms (Lumbricus rubellus Hoffmeister) to sub-lethal levels of copper (10-480 mg/kg soil) for 70 days as a real-world situation, and monitored both molecular (cDNA transcript microarrays and nuclear magnetic resonance-based metabolic profiling: metabolomics) and ecological/functional endpoints (reproduction rate and weight change, which have direct relevance to population-level impacts). Both of the molecular endpoints, metabolomics and transcriptomics, were highly sensitive, with clear copper-induced differences even at levels below those that caused a reduction in reproductive parameters. The microarray and metabolomic data provided evidence that the copper exposure led to a disruption of energy metabolism: transcripts of enzymes from oxidative phosphorylation were significantly over-represented, and increases in transcripts of carbohydrate metabolising enzymes (maltase-glucoamylase, mannosidase) had corresponding decreases in small-molecule metabolites (glucose, mannose). Treating both enzymes and metabolites as functional cohorts led to clear inferences about changes in energetic metabolism (carbohydrate use and oxidative phosphorylation), which would not have been possible by taking a 'biomarker' approach to data analysis. Multiple post-genomic techniques can be combined to provide mechanistic information about the toxic effects of chemical contaminants, even for non-model organisms with few additional mechanistic toxicological data. With 70-day no-observed-effect and lowest-observed-effect concentrations (NOEC and LOEC) of 10 and 40 mg kg-1 for metabolomic and microarray profiles, copper is shown to interfere with energy metabolism in an important soil organism at an ecologically and functionally relevant level.

High-Density Lipoprotein-Targeted Therapy and Apolipoprotein A-I Mimetic Peptides.

PubMed

Uehara, Yoshinari; Chiesa, Giulia; Saku, Keijiro

2015-01-01

Numerous randomized clinical trials have established statins as the major standard therapy for atherosclerotic diseases because these molecules decrease the plasma level of low-density lipoprotein (LDL) cholesterol and moderately increase that of plasma high-density lipoprotein (HDL) cholesterol. The reverse cholesterol transport pathway, mediated by HDL particles, has a relevant antiatherogenic potential. An important approach to HDL-targeted therapy is optimization of the HDL-cholesterol level and enhanced removal of plasma cholesterol, together with the prevention and mitigation of inflammation related to atherosclerosis. Small-molecule inhibitors of cholesteryl ester transfer protein (CETP) increase the HDL-cholesterol level in subjects with normal or low HDL-cholesterol. However, CETP inhibitors do not seem to reduce the risk of atherosclerotic diseases. HDL therapies using reconstituted HDL, including apolipoprotein (Apo) A-I Milano, ApoA-I mimetics, or full-length ApoA-I, are dramatically effective in animal models. Of those, the ApoA-I-mimetic peptide called FAMP effectively removes cholesterol via the ABCA1 transporter and acts as an antiatherosclerotic agent by enhancing the biological functions of HDL without elevating the HDL-cholesterol level. Our review of the literature leads us to conclude that HDL-targeted therapies have significant atheroprotective potential and thus may effectively treat patients with cardiovascular diseases.

A theoretical study of potentially observable chirality-sensitive NMR effects in molecules.

PubMed

Garbacz, Piotr; Cukras, Janusz; Jaszuński, Michał

2015-09-21

Two recently predicted nuclear magnetic resonance effects, the chirality-induced rotating electric polarization and the oscillating magnetization, are examined for several experimentally available chiral molecules. We discuss in detail the requirements for experimental detection of chirality-sensitive NMR effects of the studied molecules. These requirements are related to two parameters: the shielding polarizability and the antisymmetric part of the nuclear magnetic shielding tensor. The dominant second contribution has been computed for small molecules at the coupled cluster and density functional theory levels. It was found that DFT calculations using the KT2 functional and the aug-cc-pCVTZ basis set adequately reproduce the CCSD(T) values obtained with the same basis set. The largest values of parameters, thus most promising from the experimental point of view, were obtained for the fluorine nuclei in 1,3-difluorocyclopropene and 1,3-diphenyl-2-fluoro-3-trifluoromethylcyclopropene.

On the accuracy of density functional theory and wave function methods for calculating vertical ionization energies

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

McKechnie, Scott; Booth, George H.; Cohen, Aron J.

The best practice in computational methods for determining vertical ionization energies (VIEs) is assessed, via reference to experimentally determined VIEs that are corroborated by highly accurate coupled-cluster calculations. These reference values are used to benchmark the performance of density-functional theory (DFT) and wave function methods: Hartree-Fock theory (HF), second-order Møller-Plesset perturbation theory (MP2) and Electron Propagator Theory (EPT). The core test set consists of 147 small molecules. An extended set of six larger molecules, from benzene to hexacene, is also considered to investigate the dependence of the results on molecule size. The closest agreement with experiment is found for ionizationmore » energies obtained from total energy diff calculations. In particular, DFT calculations using exchange-correlation functionals with either a large amount of exact exchange or long-range correction perform best. The results from these functionals are also the least sensitive to an increase in molecule size. In general, ionization energies calculated directly from the orbital energies of the neutral species are less accurate and more sensitive to an increase in molecule size. For the single-calculation approach, the EPT calculations are in closest agreement for both sets of molecules. For the orbital energies from DFT functionals, only those with long-range correction give quantitative agreement with dramatic failing for all other functionals considered. The results offer a practical hierarchy of approximations for the calculation of vertical ionization energies. In addition, the experimental and computational reference values can be used as a standardized set of benchmarks, against which other approximate methods can be compared.« less

Beta carbonic anhydrases: novel targets for pesticides and anti-parasitic agents in agriculture and livestock husbandry.

PubMed

Zolfaghari Emameh, Reza; Barker, Harlan; Hytönen, Vesa P; Tolvanen, Martti E E; Parkkila, Seppo

2014-08-29

The genomes of many insect and parasite species contain beta carbonic anhydrase (β-CA) protein coding sequences. The lack of β-CA proteins in mammals makes them interesting target proteins for inhibition in treatment of some infectious diseases and pests. Many insects and parasites represent important pests for agriculture and cause enormous economic damage worldwide. Meanwhile, pollution of the environment by old pesticides, emergence of strains resistant to them, and their off-target effects are major challenges for agriculture and society. In this study, we analyzed a multiple sequence alignment of 31 β-CAs from insects, some parasites, and selected plant species relevant to agriculture and livestock husbandry. Using bioinformatics tools a phylogenetic tree was generated and the subcellular localizations and antigenic sites of each protein were predicted. Structural models for β-CAs of Ancylostoma caninum, Ascaris suum, Trichinella spiralis, and Entamoeba histolytica, were built using Pisum sativum and Mycobacterium tuberculosis β-CAs as templates. Six β-CAs of insects and parasites and six β-CAs of plants are predicted to be mitochondrial and chloroplastic, respectively, and thus may be involved in important metabolic functions. All 31 sequences showed the presence of the highly conserved β-CA active site sequence motifs, CXDXR and HXXC (C: cysteine, D: aspartic acid, R: arginine, H: histidine, X: any residue). We discovered that these two motifs are more antigenic than others. Homology models suggested that these motifs are mostly buried and thus not well accessible for recognition by antibodies. The predicted mitochondrial localization of several β-CAs and hidden antigenic epitopes within the protein molecule, suggest that they may not be considered major targets for vaccines. Instead, they are promising candidate enzymes for small-molecule inhibitors which can easily penetrate the cell membrane. Based on current knowledge, we conclude that β-CAs are potential targets for development of small molecule pesticides or anti-parasitic agents with minimal side effects on vertebrates.

Accurate high-speed liquid handling of very small biological samples.

PubMed

Schober, A; Günther, R; Schwienhorst, A; Döring, M; Lindemann, B F

1993-08-01

Molecular biology techniques require the accurate pipetting of buffers and solutions with volumes in the microliter range. Traditionally, hand-held pipetting devices are used to fulfill these requirements, but many laboratories have also introduced robotic workstations for the handling of liquids. Piston-operated pumps are commonly used in manually as well as automatically operated pipettors. These devices cannot meet the demands for extremely accurate pipetting of very small volumes at the high speed that would be necessary for certain applications (e.g., in sequencing projects with high throughput). In this paper we describe a technique for the accurate microdispensation of biochemically relevant solutions and suspensions with the aid of a piezoelectric transducer. It is suitable for liquids of a viscosity between 0.5 and 500 milliPascals. The obtainable drop sizes range from 5 picoliters to a few nanoliters with up to 10,000 drops per second. Liquids can be dispensed in single or accumulated drops to handle a wide volume range. The system proved to be excellently suitable for the handling of biological samples. It did not show any detectable negative impact on the biological function of dissolved or suspended molecules or particles.

Targeting RNA Splicing for Disease Therapy

PubMed Central

Havens, Mallory A.; Duelli, Dominik M.

2013-01-01

Splicing of pre-messenger RNA into mature messenger RNA is an essential step for expression of most genes in higher eukaryotes. Defects in this process typically affect cellular function and can have pathological consequences. Many human genetic diseases are caused by mutations that cause splicing defects. Furthermore, a number of diseases are associated with splicing defects that are not attributed to overt mutations. Targeting splicing directly to correct disease-associated aberrant splicing is a logical approach to therapy. Splicing is a favorable intervention point for disease therapeutics, because it is an early step in gene expression and does not alter the genome. Significant advances have been made in the development of approaches to manipulate splicing for therapy. Splicing can be manipulated with a number of tools including antisense oligonucleotides, modified small nuclear RNAs (snRNAs), trans-splicing, and small molecule compounds, all of which have been used to increase specific alternatively spliced isoforms or to correct aberrant gene expression resulting from gene mutations that alter splicing. Here we describe clinically relevant splicing defects in disease states, the current tools used to target and alter splicing, specific mutations and diseases that are being targeted using splice-modulating approaches, and emerging therapeutics. PMID:23512601

Targeting RNA splicing for disease therapy.

PubMed

Havens, Mallory A; Duelli, Dominik M; Hastings, Michelle L

2013-01-01

Splicing of pre-messenger RNA into mature messenger RNA is an essential step for the expression of most genes in higher eukaryotes. Defects in this process typically affect cellular function and can have pathological consequences. Many human genetic diseases are caused by mutations that cause splicing defects. Furthermore, a number of diseases are associated with splicing defects that are not attributed to overt mutations. Targeting splicing directly to correct disease-associated aberrant splicing is a logical approach to therapy. Splicing is a favorable intervention point for disease therapeutics, because it is an early step in gene expression and does not alter the genome. Significant advances have been made in the development of approaches to manipulate splicing for therapy. Splicing can be manipulated with a number of tools including antisense oligonucleotides, modified small nuclear RNAs (snRNAs), trans-splicing, and small molecule compounds, all of which have been used to increase specific alternatively spliced isoforms or to correct aberrant gene expression resulting from gene mutations that alter splicing. Here we describe clinically relevant splicing defects in disease states, the current tools used to target and alter splicing, specific mutations and diseases that are being targeted using splice-modulating approaches, and emerging therapeutics. Copyright © 2013 John Wiley & Sons, Ltd.

Citronellal assumes a folded conformation in solution due to dispersion interactions: A joint NMR-DFT analysis

NASA Astrophysics Data System (ADS)

Nardini, Viviani; Dias, Luis Gustavo; Palaretti, Vinicius; da Silva, Gil Valdo José

2018-04-01

Citronellal, an acyclic monoterpenoid, is a small molecule suitable for systematic scanning of its conformational geometric parameters in solution or in the gas phase. We have studied the conformational distribution of citronellal by correlating its structure and theoretical chemical shifts with nuclear magnetic resonance data. Interestingly, folded conformations were the most relevant, as confirmed by NOE experiments. We concluded that the conformational distribution is due to intramolecular dispersion interactions.

A credit-card library approach for disrupting protein-protein interactions.

PubMed

Xu, Yang; Shi, Jin; Yamamoto, Noboru; Moss, Jason A; Vogt, Peter K; Janda, Kim D

2006-04-15

Protein-protein interfaces are prominent in many therapeutically important targets. Using small organic molecules to disrupt protein-protein interactions is a current challenge in chemical biology. An important example of protein-protein interactions is provided by the Myc protein, which is frequently deregulated in human cancers. Myc belongs to the family of basic helix-loop-helix leucine zipper (bHLH-ZIP) transcription factors. It is biologically active only as heterodimer with the bHLH-ZIP protein Max. Herein, we report a new strategy for the disruption of protein-protein interactions that has been corroborated through the design and synthesis of a small parallel library composed of 'credit-card' compounds. These compounds are derived from a planar, aromatic scaffold and functionalized with four points of diversity. From a 285 membered library, several hits were obtained that disrupted the c-Myc-Max interaction and cellular functions of c-Myc. The IC50 values determined for this small focused library for the disruption of Myc-Max dimerization are quite potent, especially since small molecule antagonists of protein-protein interactions are notoriously difficult to find. Furthermore, several of the compounds were active at the cellular level as shown by their biological effects on Myc action in chicken embryo fibroblast assays. In light of our findings, this approach is considered a valuable addition to the armamentarium of new molecules being developed to interact with protein-protein interfaces. Finally, this strategy for disrupting protein-protein interactions should prove applicable to other families of proteins.

A chemical-genetic approach for functional analysis of plant protein kinases

PubMed Central

Salomon, Dor; Bonshtien, Arale

2009-01-01

Plant genomes encode hundreds of protein kinases, yet only for a small fraction of them precise functions and phosphorylation targets have been identified. Recently, we applied a chemical-genetic approach to sensitize the tomato serine/threonine kinase Pto to analogs of PP1, an ATP-competitive and cell-permeable small-molecule inhibitor. The Pto kinase confers resistance to Pst bacteria by activating immune responses upon specific recognition of bacterial effectors. By using PP1 analogs in combination with the analog-sensitive Pto, we shed new light on the role of Pto kinase activity in effector recognition and signal transduction. Here we broaden the use of this chemical-genetic approach to another defense-related plant protein kinase, the MAP kinase LeMPK3. In addition, we show that analog-sensitive but not wild-type kinases are able to use unnatural N6-modified ATP analogs as phosphodonors that can be exploited for tagging direct phosphorylation targets of the kinase of interest. Thus, sensitization of kinases to analogs of the small-molecule inhibitor PP1 and ATP can be an effective tool for the discovery of cellular functions and phosphorylation substrates of plant protein kinases. PMID:19820342

Conformation-controlled binding kinetics of antibodies

NASA Astrophysics Data System (ADS)

Galanti, Marta; Fanelli, Duccio; Piazza, Francesco

2016-01-01

Antibodies are large, extremely flexible molecules, whose internal dynamics is certainly key to their astounding ability to bind antigens of all sizes, from small hormones to giant viruses. In this paper, we build a shape-based coarse-grained model of IgG molecules and show that it can be used to generate 3D conformations in agreement with single-molecule Cryo-Electron Tomography data. Furthermore, we elaborate a theoretical model that can be solved exactly to compute the binding rate constant of a small antigen to an IgG in a prescribed 3D conformation. Our model shows that the antigen binding process is tightly related to the internal dynamics of the IgG. Our findings pave the way for further investigation of the subtle connection between the dynamics and the function of large, flexible multi-valent molecular machines.

Staphylococcal Immune Evasion Proteins: Structure, Function, and Host Adaptation.

PubMed

Koymans, Kirsten J; Vrieling, Manouk; Gorham, Ronald D; van Strijp, Jos A G

2017-01-01

Staphylococcus aureus is a successful human and animal pathogen. Its pathogenicity is linked to its ability to secrete a large amount of virulence factors. These secreted proteins interfere with many critical components of the immune system, both innate and adaptive, and hamper proper immune functioning. In recent years, numerous studies have been conducted in order to understand the molecular mechanism underlying the interaction of evasion molecules with the host immune system. Structural studies have fundamentally contributed to our understanding of the mechanisms of action of the individual factors. Furthermore, such studies revealed one of the most striking characteristics of the secreted immune evasion molecules: their conserved structure. Despite high-sequence variability, most immune evasion molecules belong to a small number of structural categories. Another remarkable characteristic is that S. aureus carries most of these virulence factors on mobile genetic elements (MGE) or ex-MGE in its accessory genome. Coevolution of pathogen and host has resulted in immune evasion molecules with a highly host-specific function and prevalence. In this review, we explore how these shared structures and genomic locations relate to function and host specificity. This is discussed in the context of therapeutic options for these immune evasion molecules in infectious as well as in inflammatory diseases.

Synthetic-Molecule/Protein Hybrid Probe with Fluorogenic Switch for Live-Cell Imaging of DNA Methylation.

PubMed

Hori, Yuichiro; Otomura, Norimichi; Nishida, Ayuko; Nishiura, Miyako; Umeno, Maho; Suetake, Isao; Kikuchi, Kazuya

2018-02-07

Hybrid probes consisting of synthetic molecules and proteins are powerful tools for detecting biological molecules and signals in living cells. To date, most targets of the hybrid probes have been limited to pH and small analytes. Although biomacromolecules are essential to the physiological function of cells, the hybrid-probe-based approach has been scarcely employed for live-cell detection of biomacromolecules. Here, we developed a hybrid probe with a chemical switch for live-cell imaging of methylated DNA, an important macromolecule in the repression of gene expression. Using a protein labeling technique, we created a hybrid probe containing a DNA-binding fluorogen and a methylated-DNA-binding domain. The hybrid probe enhanced fluorescence intensity upon binding to methylated DNA and successfully monitored methylated DNA during mitosis. The hybrid probe offers notable advantages absent from probes based on small molecules or fluorescent proteins and is useful for live-cell analyses of epigenetic phenomena and diseases related to DNA methylation.

Soluble HLA-G: Are They Clinically Relevant?

PubMed Central

Pistoia, Vito; Morandi, Fabio; Wang, Xinhui; Ferrone, Soldano

2007-01-01

HLA-G is a non-classical HLA-class Ib molecule with multiple immunoregulatory properties. Its main function in physiological conditions is to abrogate maternal NK cell activity against foetal tissue and to establish immune tolerance at maternal-foetal interface. HLA-G is expressed not only as a membrane bound molecule on the surface of cells, but also as a soluble moiety in body fluids. The major isoforms of HLA-G present in serum are soluble HLA-G1 and HLA-G5 which are generated by shedding or proteolytic cleavage of the membrane bound isoform and by secretion of a soluble isoform, respectively. Here we review the data about soluble HLA-G (sHLA-G) serum levels in different pathological conditions, including immune-mediated disorders, transplantation, and malignancies. In particular, we focus on sHLA-G expression and function in human neuroblastoma, a pediatric tumor, with special emphasis on a novel potential immuno escape mechanism utilized by NB to instruct monocytes to produce and release sHLA-G. Finally, the potential clinical relevance of sHLA-G serum levels is discussed. PMID:17825579

Guaiacol hydrodeoxygenation mechanism on Pt(111): Insights from density functional theory and linear free energy relations

USDA-ARS?s Scientific Manuscript database

In this study density functional theory (DFT) was used to study the adsorption of guaiacol and its initial hydrodeoxygenation (HDO) reactions on Pt(111). Previously reported Brønsted–Evans–Polanyi (BEP) correlations for small open chain molecules are found to be inadequate in estimating the reaction...

Microbiota and Neurological Disorders: A Gut Feeling

PubMed Central

Moos, Walter H.; Faller, Douglas V.; Harpp, David N.; Kanara, Iphigenia; Pernokas, Julie; Powers, Whitney R.; Steliou, Kosta

2016-01-01

Abstract In the past century, noncommunicable diseases have surpassed infectious diseases as the principal cause of sickness and death, worldwide. Trillions of commensal microbes live in and on our body, and constitute the human microbiome. The vast majority of these microorganisms are maternally derived and live in the gut, where they perform functions essential to our health and survival, including: digesting food, activating certain drugs, producing short-chain fatty acids (which help to modulate gene expression by inhibiting the deacetylation of histone proteins), generating anti-inflammatory substances, and playing a fundamental role in the induction, training, and function of our immune system. Among the many roles the microbiome ultimately plays, it mitigates against untoward effects from our exposure to the environment by forming a biotic shield between us and the outside world. The importance of physical activity coupled with a balanced and healthy diet in the maintenance of our well-being has been recognized since antiquity. However, it is only recently that characterization of the host–microbiome intermetabolic and crosstalk pathways has come to the forefront in studying therapeutic design. As reviewed in this report, synthetic biology shows potential in developing microorganisms for correcting pathogenic dysbiosis (gut microbiota–host maladaptation), although this has yet to be proven. However, the development and use of small molecule drugs have a long and successful history in the clinic, with small molecule histone deacetylase inhibitors representing one relevant example already approved to treat cancer and other disorders. Moreover, preclinical research suggests that epigenetic treatment of neurological conditions holds significant promise. With the mouth being an extension of the digestive tract, it presents a readily accessible diagnostic site for the early detection of potential unhealthy pathogens resident in the gut. Taken together, the data outlined herein provide an encouraging roadmap toward important new medicines and companion diagnostic platforms in a wide range of therapeutic indications. PMID:27274912

Microbiota and Neurological Disorders: A Gut Feeling.

PubMed

Moos, Walter H; Faller, Douglas V; Harpp, David N; Kanara, Iphigenia; Pernokas, Julie; Powers, Whitney R; Steliou, Kosta

2016-01-01

In the past century, noncommunicable diseases have surpassed infectious diseases as the principal cause of sickness and death, worldwide. Trillions of commensal microbes live in and on our body, and constitute the human microbiome. The vast majority of these microorganisms are maternally derived and live in the gut, where they perform functions essential to our health and survival, including: digesting food, activating certain drugs, producing short-chain fatty acids (which help to modulate gene expression by inhibiting the deacetylation of histone proteins), generating anti-inflammatory substances, and playing a fundamental role in the induction, training, and function of our immune system. Among the many roles the microbiome ultimately plays, it mitigates against untoward effects from our exposure to the environment by forming a biotic shield between us and the outside world. The importance of physical activity coupled with a balanced and healthy diet in the maintenance of our well-being has been recognized since antiquity. However, it is only recently that characterization of the host-microbiome intermetabolic and crosstalk pathways has come to the forefront in studying therapeutic design. As reviewed in this report, synthetic biology shows potential in developing microorganisms for correcting pathogenic dysbiosis (gut microbiota-host maladaptation), although this has yet to be proven. However, the development and use of small molecule drugs have a long and successful history in the clinic, with small molecule histone deacetylase inhibitors representing one relevant example already approved to treat cancer and other disorders. Moreover, preclinical research suggests that epigenetic treatment of neurological conditions holds significant promise. With the mouth being an extension of the digestive tract, it presents a readily accessible diagnostic site for the early detection of potential unhealthy pathogens resident in the gut. Taken together, the data outlined herein provide an encouraging roadmap toward important new medicines and companion diagnostic platforms in a wide range of therapeutic indications.

Two pore channels control Ebolavirus host cell entry and are drug targets for disease treatment

PubMed Central

Sakurai, Yasuteru; Kolokoltsov, Andrey A.; Chen, Cheng-Chang; Tidwell, Michael W.; Bauta, William E.; Klugbauer, Norbert; Grimm, Christian; Wahl-Schott, Christian; Biel, Martin; Davey, Robert A.

2015-01-01

Ebolavirus causes sporadic outbreaks of lethal hemorrhagic fever in humans with no currently approved therapy. Cells take up Ebolavirus by macropinocytosis, followed by trafficking through endosomal vesicles. However, few factors controlling endosomal virus movement are known. Here we find that Ebolavirus entry into host cells requires the endosomal calcium channels called two pore channels (TPCs). Disrupting TPC function by gene knockout, small interfering RNAs or small molecule inhibitors halted virus trafficking and prevented infection. Tetrandrine, the most potent small molecule we tested, inhibited infection of human macrophages, the primary target of Ebolavirus in vivo, and also showed therapeutic efficacy in mice. Therefore, TPC proteins play a key role in Ebolavirus infection and may be effective targets for antiviral therapy. PMID:25722412

Rescue of the apoptotic-inducing function of mutant p53 by small molecule RITA.

PubMed

Zhao, Carolyn Y; Grinkevich, Vera V; Nikulenkov, Fedor; Bao, Wenjie; Selivanova, Galina

2010-05-01

Expression of mutant p53 correlates with poor prognosis in many tumors, therefore strategies aimed at reactivation of mutant p53 are likely to provide important benefits for treatment of tumors that are resistant to chemotherapy and radiotherapy. We have previously identified and characterized a small molecule RITA which binds p53 and induces a conformational change which prevents the binding of p53 to several inhibitors, including its own destructor MDM2. In this way, RITA rescues the tumor suppression function of wild type p53. Here, we demonstrate that RITA suppressed the growth and induced apoptosis in human tumor cell lines of a diverse origin carrying mutant p53 proteins. RITA restored transcriptional transactivation and transrepression function of several hot spot p53 mutants. The ability of RITA to rescue the activity of different p53 mutants suggests its generic mechanism of action. Thus, RITA is a promising lead for the development of anti-cancer drugs that reactivate the tumor suppressor function of p53 in cancer cells irrespective whether they express mutant or wild type p53.

Personalizing Group Environments: A Conceptual Approach Toward More Effective Small Group Functioning.

ERIC Educational Resources Information Center

Beach, Wayne A.

A conceptual approach toward more effective small group functioning is undertaken in this paper to provide a basis from which empirically relevant hypotheses can be drawn and tested. This analysis views actualizing individuals as possessing the unique ability to perceive and utilize the types of behaviors which are conducive to personalizing group…

Two-photon absorption in conjugated energetic molecule

DOE PAGES

Bjorgaard, Josiah August; Sifain, Andrew; Nelson, Tammie Renee; ...

2016-06-03

Time-dependent density functional theory (TD-DFT) is used to investigate the relationship between molecular structure and one- and two-photon absorption (OPA and TPA, respectively) properties in novel and recently synthesized conjugated energetic molecules (CEMs). The molecular structure of CEMs can be strategically altered to influence the heat of formation and oxygen balance, two factors that can contribute to the sensitivity and strength of an explosive material. OPA and TPA are sensitive to changes in molecular structure as well, influencing optical range of excitation. We find calculated vertical excitation energies in good agreement with experiment for most molecules. Peak TPA intensities aremore » significant and on the order of 102 GM. Natural transition orbitals for essential electronic states defining TPA peaks of relatively large intensity to examine the character of relevant transitions. Minor modification of molecular substituents, such as additional oxygen and other functional groups, produces significant changes in electronic structure, OPA, TPA, and improves the oxygen balance. Results show that select molecules are apt to nonlinear absorption, opening the possibility for controlled, direct optical initiation of CEMs through photochemical pathways.« less

Amide-Directed Photoredox Catalyzed C-C Bond Formation at Unactivated sp3 C-H Bonds

PubMed Central

Chu, John C. K.; Rovis, Tomislav

2017-01-01

Carbon-carbon (C-C) bond formation is paramount in the synthesis of biologically relevant molecules, modern synthetic materials and commodity chemicals such as fuels and lubricants. Traditionally, the presence of a functional group is required at the site of C-C bond formation. Strategies that allow C-C bond formation at inert carbon-hydrogen (C-H) bonds allow scientists to access molecules which would otherwise be inaccessible and to develop more efficient syntheses of complex molecules.1,2 Herein we report a method for the formation of C-C bonds by directed cleavage of traditionally non-reactive C-H bonds and their subsequent coupling with readily available alkenes. Our methodology allows for the selective C-C bond formation at single C-H bonds in molecules that contain a multitude of seemingly indifferentiable such bonds. Selectivity arises through a relayed photoredox catalyzed oxidation of an N-H bond. We anticipate our findings to serve as a starting point for functionalization at inert C-H bonds through a hydrogen atom transfer strategy. PMID:27732580

Selective staining of proteins with hydrophobic surface sites on a native electrophoretic gel.

PubMed

Bertsch, Martina; Kassner, Richard J

2003-01-01

Chemical proteomics aims to characterize all of the proteins in the proteome with respect to their function, which is associated with their interaction with other molecules. We propose the identification of a subproteomic library of expressed proteins whose native structures are typified by the presence of hydrophobic surface sites, which are often involved in interactions with small molecules, membrane lipids, and other proteins, pertaining to their functions. We demonstrate that soluble globular proteins with hydrophobic surface sites can be detected selectively by staining on an electrophoretic gel run under nondenaturing conditions. The application of these staining techniques may help elucidate new catalytic, transport, and regulatory functionalities in complex proteomic screenings.

Mass spectrometry-based metabolomics: Targeting the crosstalk between gut microbiota and brain in neurodegenerative disorders.

PubMed

Luan, Hemi; Wang, Xian; Cai, Zongwei

2017-11-12

Metabolomics seeks to take a "snapshot" in a time of the levels, activities, regulation and interactions of all small molecule metabolites in response to a biological system with genetic or environmental changes. The emerging development in mass spectrometry technologies has shown promise in the discovery and quantitation of neuroactive small molecule metabolites associated with gut microbiota and brain. Significant progress has been made recently in the characterization of intermediate role of small molecule metabolites linked to neural development and neurodegenerative disorder, showing its potential in understanding the crosstalk between gut microbiota and the host brain. More evidence reveals that small molecule metabolites may play a critical role in mediating microbial effects on neurotransmission and disease development. Mass spectrometry-based metabolomics is uniquely suitable for obtaining the metabolic signals in bidirectional communication between gut microbiota and brain. In this review, we summarized major mass spectrometry technologies including liquid chromatography-mass spectrometry, gas chromatography-mass spectrometry, and imaging mass spectrometry for metabolomics studies of neurodegenerative disorders. We also reviewed the recent advances in the identification of new metabolites by mass spectrometry and metabolic pathways involved in the connection of intestinal microbiota and brain. These metabolic pathways allowed the microbiota to impact the regular function of the brain, which can in turn affect the composition of microbiota via the neurotransmitter substances. The dysfunctional interaction of this crosstalk connects neurodegenerative diseases, including Parkinson's disease, Alzheimer's disease and Huntington's disease. The mass spectrometry-based metabolomics analysis provides information for targeting dysfunctional pathways of small molecule metabolites in the development of the neurodegenerative diseases, which may be valuable for the investigation of underlying mechanism of therapeutic strategies. © 2017 Wiley Periodicals, Inc.

Electron dynamics in complex environments with real-time time dependent density functional theory in a QM-MM framework

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

Morzan, Uriel N.; Ramírez, Francisco F.; Scherlis, Damián A., E-mail: damian@qi.fcen.uba.ar, E-mail: mcgl@qb.ffyb.uba.ar

2014-04-28

This article presents a time dependent density functional theory (TDDFT) implementation to propagate the Kohn-Sham equations in real time, including the effects of a molecular environment through a Quantum-Mechanics Molecular-Mechanics (QM-MM) hamiltonian. The code delivers an all-electron description employing Gaussian basis functions, and incorporates the Amber force-field in the QM-MM treatment. The most expensive parts of the computation, comprising the commutators between the hamiltonian and the density matrix—required to propagate the electron dynamics—, and the evaluation of the exchange-correlation energy, were migrated to the CUDA platform to run on graphics processing units, which remarkably accelerates the performance of the code.more » The method was validated by reproducing linear-response TDDFT results for the absorption spectra of several molecular species. Two different schemes were tested to propagate the quantum dynamics: (i) a leap-frog Verlet algorithm, and (ii) the Magnus expansion to first-order. These two approaches were confronted, to find that the Magnus scheme is more efficient by a factor of six in small molecules. Interestingly, the presence of iron was found to seriously limitate the length of the integration time step, due to the high frequencies associated with the core-electrons. This highlights the importance of pseudopotentials to alleviate the cost of the propagation of the inner states when heavy nuclei are present. Finally, the methodology was applied to investigate the shifts induced by the chemical environment on the most intense UV absorption bands of two model systems of general relevance: the formamide molecule in water solution, and the carboxy-heme group in Flavohemoglobin. In both cases, shifts of several nanometers are observed, consistently with the available experimental data.« less

Electron dynamics in complex environments with real-time time dependent density functional theory in a QM-MM framework

NASA Astrophysics Data System (ADS)

Morzan, Uriel N.; Ramírez, Francisco F.; Oviedo, M. Belén; Sánchez, Cristián G.; Scherlis, Damián A.; Lebrero, Mariano C. González

2014-04-01

This article presents a time dependent density functional theory (TDDFT) implementation to propagate the Kohn-Sham equations in real time, including the effects of a molecular environment through a Quantum-Mechanics Molecular-Mechanics (QM-MM) hamiltonian. The code delivers an all-electron description employing Gaussian basis functions, and incorporates the Amber force-field in the QM-MM treatment. The most expensive parts of the computation, comprising the commutators between the hamiltonian and the density matrix—required to propagate the electron dynamics—, and the evaluation of the exchange-correlation energy, were migrated to the CUDA platform to run on graphics processing units, which remarkably accelerates the performance of the code. The method was validated by reproducing linear-response TDDFT results for the absorption spectra of several molecular species. Two different schemes were tested to propagate the quantum dynamics: (i) a leap-frog Verlet algorithm, and (ii) the Magnus expansion to first-order. These two approaches were confronted, to find that the Magnus scheme is more efficient by a factor of six in small molecules. Interestingly, the presence of iron was found to seriously limitate the length of the integration time step, due to the high frequencies associated with the core-electrons. This highlights the importance of pseudopotentials to alleviate the cost of the propagation of the inner states when heavy nuclei are present. Finally, the methodology was applied to investigate the shifts induced by the chemical environment on the most intense UV absorption bands of two model systems of general relevance: the formamide molecule in water solution, and the carboxy-heme group in Flavohemoglobin. In both cases, shifts of several nanometers are observed, consistently with the available experimental data.

Electron dynamics in complex environments with real-time time dependent density functional theory in a QM-MM framework.

PubMed

Morzan, Uriel N; Ramírez, Francisco F; Oviedo, M Belén; Sánchez, Cristián G; Scherlis, Damián A; Lebrero, Mariano C González

2014-04-28

This article presents a time dependent density functional theory (TDDFT) implementation to propagate the Kohn-Sham equations in real time, including the effects of a molecular environment through a Quantum-Mechanics Molecular-Mechanics (QM-MM) hamiltonian. The code delivers an all-electron description employing Gaussian basis functions, and incorporates the Amber force-field in the QM-MM treatment. The most expensive parts of the computation, comprising the commutators between the hamiltonian and the density matrix-required to propagate the electron dynamics-, and the evaluation of the exchange-correlation energy, were migrated to the CUDA platform to run on graphics processing units, which remarkably accelerates the performance of the code. The method was validated by reproducing linear-response TDDFT results for the absorption spectra of several molecular species. Two different schemes were tested to propagate the quantum dynamics: (i) a leap-frog Verlet algorithm, and (ii) the Magnus expansion to first-order. These two approaches were confronted, to find that the Magnus scheme is more efficient by a factor of six in small molecules. Interestingly, the presence of iron was found to seriously limitate the length of the integration time step, due to the high frequencies associated with the core-electrons. This highlights the importance of pseudopotentials to alleviate the cost of the propagation of the inner states when heavy nuclei are present. Finally, the methodology was applied to investigate the shifts induced by the chemical environment on the most intense UV absorption bands of two model systems of general relevance: the formamide molecule in water solution, and the carboxy-heme group in Flavohemoglobin. In both cases, shifts of several nanometers are observed, consistently with the available experimental data.

Development of novel small molecules for imaging and drug release

NASA Astrophysics Data System (ADS)

Cao, Yanting

Small organic molecules, including small molecule based fluorescent probes, small molecule based drugs or prodrugs, and smart multifunctional fluorescent drug delivery systems play important roles in biological research, drug discovery, and clinical practices. Despite the significant progress made in these fields, the development of novel and diverse small molecules is needed to meet various demands for research and clinical applications. My Ph.D study focuses on the development of novel functional molecules for recognition, imaging and drug release. In the first part, a turn-on fluorescent probe is developed for the detection of intracellular adenosine-5'-triphosphate (ATP) levels based on multiplexing recognitions. Considering the unique and complicated structure of ATP molecules, a fluorescent probe has been implemented with improved sensitivity and selectivity due to two synergistic binding recognitions by incorporating of 2, 2'-dipicolylamine (Dpa)-Zn(II) for targeting of phospho anions and phenylboronic acid group for cis-diol moiety. The novel probe is able to detect intracellular ATP levels in SH-SY5Y cells. Meanwhile, the advantages of multiplexing recognition design concept have been demonstrated using two control molecules. In the second part, a prodrug system is developed to deliver multiple drugs within one small molecule entity. The prodrug is designed by using 1-(2-nitrophenyl)ethyl (NPE) as phototrigger, and biphenol biquaternary ammonium as the prodrug. With controlled photo activation, both DNA cross-linking agents mechlorethamine and o-quinone methide are delivered and released at the preferred site, leading to efficient DNA cross-links formation and cell death. The prodrug shows negligible cytotoxicity towards normal skin cells (Hekn cells) with and without UV activation, but displays potent activity towards cancer cells (HeLa cells) upon UV activation. The multiple drug release system may hold a great potential for practical application. In the last part, a new photo-initiated fluorescent anticancer prodrug for DNA alkylating agent mechlorethamine releasing and monitoring has been developed. The theranostic prodrug consists a photolabile NPE group, an inactive form of mechlorethamine and a nonfluorescent coumarin in one small molecule. It is demonstrated that the prodrug shows negligible cytotoxicity towards normal skin cells (Hekn cells) with and without UV activation, while the original parent drug mechlorethamine can be photocontrol-released and induces effective DNA cross-linking activity. Importantly, the drug release progress can be conveniently monitored by the 'off-on' fluorescence enhancement in cells. Moreover, the selective prodrug is not only cell permeable but also nuclear permeable. Therefore, the prodrug serves as a promising drug delivery system for spatiotemporal control release and monitoring of an anticancer drug to obtain the optimal treatment efficacy.

Differential Toxicity of mDia Formin-Directed Functional Agonists and Antagonists in Developing Zebrafish.

PubMed

LeCorgne, Hunter; Tudosie, Andrew M; Lavik, Kari; Su, Robin; Becker, Kathryn N; Moore, Sara; Walia, Yashna; Wisner, Alexander; Koehler, Daniel; Alberts, Arthur S; Williams, Frederick E; Eisenmann, Kathryn M

2018-01-01

The mammalian Diaphanous-related (mDia) formins are cytoskeletal regulators that assemble and, in some cases, bundle filamentous actin (F-actin), as well as stabilize microtubules. The development of small molecule antagonists and agonists that interrogate mDia formin function has allowed us to investigate the roles of formins in disease states. A small molecule inhibitor of FH2 domain (SMIFH2) inhibits mDia-dependent actin dynamics and abrogates tumor cell migration and cell division in vitro and ex vivo tissue explants. mDia formin activation with small molecule intramimics IMM01/02 and mDia2-DAD peptides inhibited glioblastoma motility and invasion in vitro and ex vivo rat brain slices. However, SMIFH2, IMMs, and mDia2 DAD efficacy in vivo remains largely unexplored and potential toxicity across a range of developmental phenotypes has not been thoroughly characterized. In this study, we performed an in vivo screen of early life-stage toxicity in Danio rerio zebrafish embryos 2 days post-fertilization (dpf) in response to SMIFH2, IMM01/02, and mDia2 DAD. SMIFH2 at concentrations ≥5-10 μM induced significant defects in developing zebrafish, including shorter body lengths, tail curvature and defective tail cellularity, craniofacial malformations, pericardial edema, absent and/or compromised vasculature function and flow, depressed heart rates and increased mortality. Conversely, IMM and mDia2 DAD peptides were minimally toxic at concentrations up to 10-20 and 50 μM, respectively. SMIFH2's therapeutic potential may therefore be limited by its substantial in vivo toxicity at functional concentrations. mDia formin agonism with IMMs and mDia2 DADs may therefore be a more effective and less toxic anti-invasive therapeutic approach.

Differential Toxicity of mDia Formin-Directed Functional Agonists and Antagonists in Developing Zebrafish

PubMed Central

LeCorgne, Hunter; Tudosie, Andrew M.; Lavik, Kari; Su, Robin; Becker, Kathryn N.; Moore, Sara; Walia, Yashna; Wisner, Alexander; Koehler, Daniel; Alberts, Arthur S.; Williams, Frederick E.; Eisenmann, Kathryn M.

2018-01-01

The mammalian Diaphanous-related (mDia) formins are cytoskeletal regulators that assemble and, in some cases, bundle filamentous actin (F-actin), as well as stabilize microtubules. The development of small molecule antagonists and agonists that interrogate mDia formin function has allowed us to investigate the roles of formins in disease states. A small molecule inhibitor of FH2 domain (SMIFH2) inhibits mDia-dependent actin dynamics and abrogates tumor cell migration and cell division in vitro and ex vivo tissue explants. mDia formin activation with small molecule intramimics IMM01/02 and mDia2-DAD peptides inhibited glioblastoma motility and invasion in vitro and ex vivo rat brain slices. However, SMIFH2, IMMs, and mDia2 DAD efficacy in vivo remains largely unexplored and potential toxicity across a range of developmental phenotypes has not been thoroughly characterized. In this study, we performed an in vivo screen of early life-stage toxicity in Danio rerio zebrafish embryos 2 days post-fertilization (dpf) in response to SMIFH2, IMM01/02, and mDia2 DAD. SMIFH2 at concentrations ≥5–10 μM induced significant defects in developing zebrafish, including shorter body lengths, tail curvature and defective tail cellularity, craniofacial malformations, pericardial edema, absent and/or compromised vasculature function and flow, depressed heart rates and increased mortality. Conversely, IMM and mDia2 DAD peptides were minimally toxic at concentrations up to 10–20 and 50 μM, respectively. SMIFH2's therapeutic potential may therefore be limited by its substantial in vivo toxicity at functional concentrations. mDia formin agonism with IMMs and mDia2 DADs may therefore be a more effective and less toxic anti-invasive therapeutic approach. PMID:29692731

Meta-Analysis of Placental Transcriptome Data Identifies a Novel Molecular Pathway Related to Preeclampsia.

PubMed

van Uitert, Miranda; Moerland, Perry D; Enquobahrie, Daniel A; Laivuori, Hannele; van der Post, Joris A M; Ris-Stalpers, Carrie; Afink, Gijs B

2015-01-01

Studies using the placental transcriptome to identify key molecules relevant for preeclampsia are hampered by a relatively small sample size. In addition, they use a variety of bioinformatics and statistical methods, making comparison of findings challenging. To generate a more robust preeclampsia gene expression signature, we performed a meta-analysis on the original data of 11 placenta RNA microarray experiments, representing 139 normotensive and 116 preeclamptic pregnancies. Microarray data were pre-processed and analyzed using standardized bioinformatics and statistical procedures and the effect sizes were combined using an inverse-variance random-effects model. Interactions between genes in the resulting gene expression signature were identified by pathway analysis (Ingenuity Pathway Analysis, Gene Set Enrichment Analysis, Graphite) and protein-protein associations (STRING). This approach has resulted in a comprehensive list of differentially expressed genes that led to a 388-gene meta-signature of preeclamptic placenta. Pathway analysis highlights the involvement of the previously identified hypoxia/HIF1A pathway in the establishment of the preeclamptic gene expression profile, while analysis of protein interaction networks indicates CREBBP/EP300 as a novel element central to the preeclamptic placental transcriptome. In addition, there is an apparent high incidence of preeclampsia in women carrying a child with a mutation in CREBBP/EP300 (Rubinstein-Taybi Syndrome). The 388-gene preeclampsia meta-signature offers a vital starting point for further studies into the relevance of these genes (in particular CREBBP/EP300) and their concomitant pathways as biomarkers or functional molecules in preeclampsia. This will result in a better understanding of the molecular basis of this disease and opens up the opportunity to develop rational therapies targeting the placental dysfunction causal to preeclampsia.

Confined wormlike chains in external fields

NASA Astrophysics Data System (ADS)

Morrison, Greg

The confinement of biomolecules is ubiquitous in nature, such as the spatial constraints of viral encapsulation, histone binding, and chromosomal packing. Advances in microfluidics and nanopore fabrication have permitted powerful new tools in single molecule manipulation and gene sequencing through molecular confinement as well. In order to fully understand and exploit these systems, the ability to predict the structure of spatially confined molecules is essential. In this talk, I describe a mean field approach to determine the properties of stiff polymers confined to cylinders and slits, which is relevant for a variety of biological and experimental conditions. I show that this approach is able to not only reproduce known scaling laws for confined wormlike chains, but also provides an improvement over existing weakly bending rod approximations in determining the detailed chain properties (such as correlation functions). Using this approach, we also show that it is possible to study the effect of an externally applied tension or static electric field in a natural and analytically tractable way. These external perturbations can alter the scaling laws and introduce important new length scales into the system, relevant for histone unbinding and single-molecule analysis of DNA.

University of Texas Southwestern Medical Center (UTSW): Lung Cancer Oncogenotype-Selective Drug Target Discovery (Natural Products Focus) | Office of Cancer Genomics

Cancer.gov

The goal of this project is to use small molecules and RNAi to functionally define subtypes of non-small cell lung cancer (NSCLC) using a panel of cell lines prepared and molecularly annotated by Drs. John Minna and Adi Gazdar. Experimental Approaches Lung Cancer Natural Products Screening/Chemical Library Screening

University of Texas Southwestern Medical Center: Lung Cancer Oncogenotype-Selective Drug Target Discovery (Natural Products Focus) | Office of Cancer Genomics

Cancer.gov

The goal of this project is to use small molecules and RNAi to functionally define subtypes of non-small cell lung cancer (NSCLC) using a panel of cell lines prepared and molecularly annotated by Drs. John Minna and Adi Gazdar. Experimental Approaches Lung Cancer Natural Products Screening/Chemical Library Screening

Small non-coding RNAs (sncRNA) regulate gene silencing and modify homeostatic status in animals faced with porcine reproductive and respiratory syndrome virus (PRRSV)

USDA-ARS?s Scientific Manuscript database

It has been established that reduced susceptibility to porcine reproductive and respiratory syndrome virus (PRRSV) has a genetic component. This genetic component may take the form of small non-coding RNAs (sncRNA), which are molecules that function as regulators of gene expression. Various sncRNAs ...

Small Molecule Recognition and Tools to Study Modulation of r(CGG)(exp) in Fragile X-Associated Tremor Ataxia Syndrome.

PubMed

Yang, Wang-Yong; He, Fang; Strack, Rita L; Oh, Seok Yoon; Frazer, Michelle; Jaffrey, Samie R; Todd, Peter K; Disney, Matthew D

2016-09-16

RNA transcripts containing expanded nucleotide repeats cause many incurable diseases via various mechanisms. One such disorder, fragile X-associated tremor ataxia syndrome (FXTAS), is caused by a noncoding r(CGG) repeat expansion (r(CGG)(exp)) that (i) sequesters proteins involved in RNA metabolism in nuclear foci, causing dysregulation of alternative pre-mRNA splicing, and (ii) undergoes repeat associated non-ATG translation (RANT), which produces toxic homopolymeric proteins without using a start codon. Here, we describe the design of two small molecules that inhibit both modes of toxicity and the implementation of various tools to study perturbation of these cellular events. Competitive Chemical Cross Linking and Isolation by Pull Down (C-Chem-CLIP) established that compounds bind r(CGG)(exp) and defined small molecule occupancy of r(CGG)(exp) in cells, the first approach to do so. Using an RNA GFP mimic, r(CGG)(exp)-Spinach2, we observe that our optimal designed compound binds r(CGG)(exp) and affects RNA localization by disrupting preformed RNA foci. These events correlate with an improvement of pre-mRNA splicing defects caused by RNA gain of function. In addition, the compounds reduced levels of toxic homopolymeric proteins formed via RANT. Polysome profiling studies showed that small molecules decreased loading of polysomes onto r(CGG)(exp), explaining decreased translation.

Small Molecule Deubiquitinase Inhibitors Promote Macrophage Anti-Infective Capacity

PubMed Central

Charbonneau, Marie-Eve; Gonzalez-Hernandez, Marta J.; Showalter, Hollis D.; Donato, Nicholas J.; Wobus, Christiane E.; O’Riordan, Mary X. D.

2014-01-01

The global spread of anti-microbial resistance requires urgent attention, and diverse alternative strategies have been suggested to address this public health concern. Host-directed immunomodulatory therapies represent one approach that could reduce selection for resistant bacterial strains. Recently, the small molecule deubiquitinase inhibitor WP1130 was reported as a potential anti-infective drug against important human food-borne pathogens, notably Listeria monocytogenes and noroviruses. Utilization of WP1130 itself is limited due to poor solubility, but given the potential of this new compound, we initiated an iterative rational design approach to synthesize new derivatives with increased solubility that retained anti-infective activity. Here, we test a small library of novel synthetic molecules based on the structure of the parent compound, WP1130, for anti-infective activity in vitro. Our studies identify a promising candidate, compound 9, which reduced intracellular growth of L. monocytogenes at concentrations that caused minimal cellular toxicity. Compound 9 itself had no bactericidal activity and only modestly slowed Listeria growth rate in liquid broth culture, suggesting that this drug acts as an anti-infective compound by modulating host-cell function. Moreover, this new compound also showed anti-infective activity against murine norovirus (MNV-1) and human norovirus, using the Norwalk virus replicon system. This small molecule inhibitor may provide a chemical platform for further development of therapeutic deubiquitinase inhibitors with broad-spectrum anti-infective activity. PMID:25093325

Small molecule deubiquitinase inhibitors promote macrophage anti-infective capacity.

PubMed

Charbonneau, Marie-Eve; Gonzalez-Hernandez, Marta J; Showalter, Hollis D; Donato, Nicholas J; Wobus, Christiane E; O'Riordan, Mary X D

2014-01-01

The global spread of anti-microbial resistance requires urgent attention, and diverse alternative strategies have been suggested to address this public health concern. Host-directed immunomodulatory therapies represent one approach that could reduce selection for resistant bacterial strains. Recently, the small molecule deubiquitinase inhibitor WP1130 was reported as a potential anti-infective drug against important human food-borne pathogens, notably Listeria monocytogenes and noroviruses. Utilization of WP1130 itself is limited due to poor solubility, but given the potential of this new compound, we initiated an iterative rational design approach to synthesize new derivatives with increased solubility that retained anti-infective activity. Here, we test a small library of novel synthetic molecules based on the structure of the parent compound, WP1130, for anti-infective activity in vitro. Our studies identify a promising candidate, compound 9, which reduced intracellular growth of L. monocytogenes at concentrations that caused minimal cellular toxicity. Compound 9 itself had no bactericidal activity and only modestly slowed Listeria growth rate in liquid broth culture, suggesting that this drug acts as an anti-infective compound by modulating host-cell function. Moreover, this new compound also showed anti-infective activity against murine norovirus (MNV-1) and human norovirus, using the Norwalk virus replicon system. This small molecule inhibitor may provide a chemical platform for further development of therapeutic deubiquitinase inhibitors with broad-spectrum anti-infective activity.

Hitting the sweet spot

NASA Astrophysics Data System (ADS)

Malins, Lara R.

2018-06-01

As the most abundant class of biomolecules on Earth, carbohydrates are implicated in a multitude of biological functions. Now, a simple chemical transformation has enabled the direct and selective installation of carbohydrates onto a diverse range of small molecules and peptides.

Non-Born-Oppenheimer self-consistent field calculations with cubic scaling

NASA Astrophysics Data System (ADS)

Moncada, Félix; Posada, Edwin; Flores-Moreno, Roberto; Reyes, Andrés

2012-05-01

An efficient nuclear molecular orbital methodology is presented. This approach combines an auxiliary density functional theory for electrons (ADFT) and a localized Hartree product (LHP) representation for the nuclear wave function. A series of test calculations conducted on small molecules exposed that energy and geometry errors introduced by the use of ADFT and LHP approximations are small and comparable to those obtained by the use of electronic ADFT. In addition, sample calculations performed on (HF)n chains disclosed that the combined ADFT/LHP approach scales cubically with system size (n) as opposed to the quartic scaling of Hartree-Fock/LHP or DFT/LHP methods. Even for medium size molecules the improved scaling of the ADFT/LHP approach resulted in speedups of at least 5x with respect to Hartree-Fock/LHP calculations. The ADFT/LHP method opens up the possibility of studying nuclear quantum effects on large size systems that otherwise would be impractical.

A human genome-wide loss-of-function screen identifies effective chikungunya antiviral drugs

PubMed Central

Karlas, Alexander; Berre, Stefano; Couderc, Thérèse; Varjak, Margus; Braun, Peter; Meyer, Michael; Gangneux, Nicolas; Karo-Astover, Liis; Weege, Friderike; Raftery, Martin; Schönrich, Günther; Klemm, Uwe; Wurzlbauer, Anne; Bracher, Franz; Merits, Andres; Meyer, Thomas F.; Lecuit, Marc

2016-01-01

Chikungunya virus (CHIKV) is a globally spreading alphavirus against which there is no commercially available vaccine or therapy. Here we use a genome-wide siRNA screen to identify 156 proviral and 41 antiviral host factors affecting CHIKV replication. We analyse the cellular pathways in which human proviral genes are involved and identify druggable targets. Twenty-one small-molecule inhibitors, some of which are FDA approved, targeting six proviral factors or pathways, have high antiviral activity in vitro, with low toxicity. Three identified inhibitors have prophylactic antiviral effects in mouse models of chikungunya infection. Two of them, the calmodulin inhibitor pimozide and the fatty acid synthesis inhibitor TOFA, have a therapeutic effect in vivo when combined. These results demonstrate the value of loss-of-function screening and pathway analysis for the rational identification of small molecules with therapeutic potential and pave the way for the development of new, host-directed, antiviral agents. PMID:27177310

A human genome-wide loss-of-function screen identifies effective chikungunya antiviral drugs.

PubMed

Karlas, Alexander; Berre, Stefano; Couderc, Thérèse; Varjak, Margus; Braun, Peter; Meyer, Michael; Gangneux, Nicolas; Karo-Astover, Liis; Weege, Friderike; Raftery, Martin; Schönrich, Günther; Klemm, Uwe; Wurzlbauer, Anne; Bracher, Franz; Merits, Andres; Meyer, Thomas F; Lecuit, Marc

2016-05-12

Chikungunya virus (CHIKV) is a globally spreading alphavirus against which there is no commercially available vaccine or therapy. Here we use a genome-wide siRNA screen to identify 156 proviral and 41 antiviral host factors affecting CHIKV replication. We analyse the cellular pathways in which human proviral genes are involved and identify druggable targets. Twenty-one small-molecule inhibitors, some of which are FDA approved, targeting six proviral factors or pathways, have high antiviral activity in vitro, with low toxicity. Three identified inhibitors have prophylactic antiviral effects in mouse models of chikungunya infection. Two of them, the calmodulin inhibitor pimozide and the fatty acid synthesis inhibitor TOFA, have a therapeutic effect in vivo when combined. These results demonstrate the value of loss-of-function screening and pathway analysis for the rational identification of small molecules with therapeutic potential and pave the way for the development of new, host-directed, antiviral agents.

Small Molecule Bcl2 BH4 Antagonist for Lung Cancer Therapy

PubMed Central

Han, Bingshe; Park, Dongkyoo; Li, Rui; Xie, Maohua; Owonikoko, Taofeek K.; Zhang, Guojing; Sica, Gabriel L.; Ding, Chunyong; Zhou, Jia; Magis, Andrew T.; Chen, Zhuo G.; Shin, Dong M.; Ramalingam, Suresh S.; Khuri, Fadlo R.; Curran, Walter J.; Deng, Xingming

2015-01-01

SUMMARY The BH4 domain of Bcl2 is required for its antiapoptotic function, thus constituting a promising anticancer target. We identified a small molecule Bcl2-BH4 domain-antagonist (BDA-366) that binds BH4 with high affinity and selectivity. BDA-366-Bcl2 binding induces conformational change in Bcl2 that abrogates its antiapoptotic function, converting it from a survival to a cell death inducer. BDA-366 suppresses growth of lung cancer xenografts derived from cell lines and patient without significant normal tissue toxicity at effective doses. mTOR inhibition up-regulates Bcl2 in lung cancer cells and tumor tissues from clinical trial patients. Combined BDA-366 and RAD001 treatment exhibits strong synergy against lung cancer in vivo. Development of this Bcl2-BH4 antagonist may provide a strategy to improve lung cancer outcome. PMID:26004684

STK33 kinase inhibitor BRD-8899 has no effect on KRAS-dependent cancer cell viability.

PubMed

Luo, Tuoping; Masson, Kristina; Jaffe, Jacob D; Silkworth, Whitney; Ross, Nathan T; Scherer, Christina A; Scholl, Claudia; Fröhling, Stefan; Carr, Steven A; Stern, Andrew M; Schreiber, Stuart L; Golub, Todd R

2012-02-21

Approximately 30% of human cancers harbor oncogenic gain-of-function mutations in KRAS. Despite interest in KRAS as a therapeutic target, direct blockade of KRAS function with small molecules has yet to be demonstrated. Based on experiments that lower mRNA levels of protein kinases, KRAS-dependent cancer cells were proposed to have a unique requirement for the serine/threonine kinase STK33. Thus, it was suggested that small-molecule inhibitors of STK33 might have therapeutic benefit in these cancers. Here, we describe the development of selective, low nanomolar inhibitors of STK33's kinase activity. The most potent and selective of these, BRD8899, failed to kill KRAS-dependent cells. While several explanations for this result exist, our data are most consistent with the view that inhibition of STK33's kinase activity does not represent a promising anti-KRAS therapeutic strategy.

STK33 kinase inhibitor BRD-8899 has no effect on KRAS-dependent cancer cell viability

PubMed Central

Luo, Tuoping; Masson, Kristina; Jaffe, Jacob D.; Silkworth, Whitney; Ross, Nathan T.; Scherer, Christina A.; Scholl, Claudia; Fröhling, Stefan; Carr, Steven A.; Stern, Andrew M.; Schreiber, Stuart L.; Golub, Todd R.

2012-01-01

Approximately 30% of human cancers harbor oncogenic gain-of-function mutations in KRAS. Despite interest in KRAS as a therapeutic target, direct blockade of KRAS function with small molecules has yet to be demonstrated. Based on experiments that lower mRNA levels of protein kinases, KRAS-dependent cancer cells were proposed to have a unique requirement for the serine/threonine kinase STK33. Thus, it was suggested that small-molecule inhibitors of STK33 might have therapeutic benefit in these cancers. Here, we describe the development of selective, low nanomolar inhibitors of STK33’s kinase activity. The most potent and selective of these, BRD8899, failed to kill KRAS-dependent cells. While several explanations for this result exist, our data are most consistent with the view that inhibition of STK33’s kinase activity does not represent a promising anti-KRAS therapeutic strategy. PMID:22323609

Molecular kinetics. Ras activation by SOS: allosteric regulation by altered fluctuation dynamics.

PubMed

Iversen, Lars; Tu, Hsiung-Lin; Lin, Wan-Chen; Christensen, Sune M; Abel, Steven M; Iwig, Jeff; Wu, Hung-Jen; Gureasko, Jodi; Rhodes, Christopher; Petit, Rebecca S; Hansen, Scott D; Thill, Peter; Yu, Cheng-Han; Stamou, Dimitrios; Chakraborty, Arup K; Kuriyan, John; Groves, Jay T

2014-07-04

Activation of the small guanosine triphosphatase H-Ras by the exchange factor Son of Sevenless (SOS) is an important hub for signal transduction. Multiple layers of regulation, through protein and membrane interactions, govern activity of SOS. We characterized the specific activity of individual SOS molecules catalyzing nucleotide exchange in H-Ras. Single-molecule kinetic traces revealed that SOS samples a broad distribution of turnover rates through stochastic fluctuations between distinct, long-lived (more than 100 seconds), functional states. The expected allosteric activation of SOS by Ras-guanosine triphosphate (GTP) was conspicuously absent in the mean rate. However, fluctuations into highly active states were modulated by Ras-GTP. This reveals a mechanism in which functional output may be determined by the dynamical spectrum of rates sampled by a small number of enzymes, rather than the ensemble average. Copyright © 2014, American Association for the Advancement of Science.

A Multiwell Platform for Studying Stiffness-Dependent Cell Biology

PubMed Central

Mih, Justin D.; Sharif, Asma S.; Liu, Fei; Marinkovic, Aleksandar; Symer, Matthew M.; Tschumperlin, Daniel J.

2011-01-01

Adherent cells are typically cultured on rigid substrates that are orders of magnitude stiffer than their tissue of origin. Here, we describe a method to rapidly fabricate 96 and 384 well platforms for routine screening of cells in tissue-relevant stiffness contexts. Briefly, polyacrylamide (PA) hydrogels are cast in glass-bottom plates, functionalized with collagen, and sterilized for cell culture. The Young's modulus of each substrate can be specified from 0.3 to 55 kPa, with collagen surface density held constant over the stiffness range. Using automated fluorescence microscopy, we captured the morphological variations of 7 cell types cultured across a physiological range of stiffness within a 384 well plate. We performed assays of cell number, proliferation, and apoptosis in 96 wells and resolved distinct profiles of cell growth as a function of stiffness among primary and immortalized cell lines. We found that the stiffness-dependent growth of normal human lung fibroblasts is largely invariant with collagen density, and that differences in their accumulation are amplified by increasing serum concentration. Further, we performed a screen of 18 bioactive small molecules and identified compounds with enhanced or reduced effects on soft versus rigid substrates, including blebbistatin, which abolished the suppression of lung fibroblast growth at 1 kPa. The ability to deploy PA gels in multiwell plates for high throughput analysis of cells in tissue-relevant environments opens new opportunities for the discovery of cellular responses that operate in specific stiffness regimes. PMID:21637769

A multiwell platform for studying stiffness-dependent cell biology.

PubMed

Mih, Justin D; Sharif, Asma S; Liu, Fei; Marinkovic, Aleksandar; Symer, Matthew M; Tschumperlin, Daniel J

2011-01-01

Adherent cells are typically cultured on rigid substrates that are orders of magnitude stiffer than their tissue of origin. Here, we describe a method to rapidly fabricate 96 and 384 well platforms for routine screening of cells in tissue-relevant stiffness contexts. Briefly, polyacrylamide (PA) hydrogels are cast in glass-bottom plates, functionalized with collagen, and sterilized for cell culture. The Young's modulus of each substrate can be specified from 0.3 to 55 kPa, with collagen surface density held constant over the stiffness range. Using automated fluorescence microscopy, we captured the morphological variations of 7 cell types cultured across a physiological range of stiffness within a 384 well plate. We performed assays of cell number, proliferation, and apoptosis in 96 wells and resolved distinct profiles of cell growth as a function of stiffness among primary and immortalized cell lines. We found that the stiffness-dependent growth of normal human lung fibroblasts is largely invariant with collagen density, and that differences in their accumulation are amplified by increasing serum concentration. Further, we performed a screen of 18 bioactive small molecules and identified compounds with enhanced or reduced effects on soft versus rigid substrates, including blebbistatin, which abolished the suppression of lung fibroblast growth at 1 kPa. The ability to deploy PA gels in multiwell plates for high throughput analysis of cells in tissue-relevant environments opens new opportunities for the discovery of cellular responses that operate in specific stiffness regimes.

Models of Protocellular Structure, Function and Evolution

NASA Technical Reports Server (NTRS)

New, Michael H.; Pohorille, Andrew; Szostak, Jack W.; Keefe, Tony; Lanyi, Janos K.; DeVincenzi, Donald L. (Technical Monitor)

2001-01-01

In the absence of any record of protocells, the most direct way to test our understanding, of the origin of cellular life is to construct laboratory models that capture important features of protocellular systems. Such efforts are currently underway in a collaborative project between NASA-Ames, Harvard Medical School and University of California. They are accompanied by computational studies aimed at explaining self-organization of simple molecules into ordered structures. The centerpiece of this project is a method for the in vitro evolution of protein enzymes toward arbitrary catalytic targets. A similar approach has already been developed for nucleic acids in which a small number of functional molecules are selected from a large, random population of candidates. The selected molecules are next vastly multiplied using the polymerase chain reaction.

MicroRNAs in islet immunobiology and transplantation.

PubMed

Pileggi, Antonello; Klein, Dagmar; Fotino, Carmen; Bravo-Egaña, Valia; Rosero, Samuel; Doni, Marco; Podetta, Michele; Ricordi, Camillo; Molano, R Damaris; Pastori, Ricardo L

2013-12-01

The ultimate goal of diabetes therapy is the restoration of physiologic metabolic control. For type 1 diabetes, research efforts are focused on the prevention or early intervention to halt the autoimmune process and preserve β cell function. Replacement of pancreatic β cells via islet transplantation reestablishes physiologic β cell function in patients with diabetes. Emerging research shows that microRNAs (miRNAs), noncoding small RNA molecules produced by a newly discovered class of genes, negatively regulate gene expression. MiRNAs recognize and bind to partially complementary sequences of target messenger RNA (mRNA), regulating mRNA translation and affecting gene expression. Correlation between miRNA signatures and genome-wide RNA expression allows identification of multiple miRNA-mRNA pairs in biological processes. Because miRNAs target functionally related genes, they represent an exciting and indispensable approach for biomarkers and drug discovery. We are studying the role of miRNA in the context of islet immunobiology. Our research aims at understanding the mechanisms underlying pancreatic β cell loss and developing clinically relevant approaches for preservation and restoration of β cell function to treat insulin-dependent diabetes. Herein, we discuss some of our recent efforts related to the study of miRNA in islet inflammation and islet engraftment. Our working hypothesis is that modulation of the expression of specific microRNAs in the transplant microenvironment will be of assistance in enhancing islet engraftment and promoting long-term function.

Small Molecule Organic Optoelectronic Devices

NASA Astrophysics Data System (ADS)

Bakken, Nathan

Organic optoelectronics include a class of devices synthesized from carbon containing 'small molecule' thin films without long range order crystalline or polymer structure. Novel properties such as low modulus and flexibility as well as excellent device performance such as photon emission approaching 100% internal quantum efficiency have accelerated research in this area substantially. While optoelectronic organic light emitting devices have already realized commercial application, challenges to obtain extended lifetime for the high energy visible spectrum and the ability to reproduce natural white light with a simple architecture have limited the value of this technology for some display and lighting applications. In this research, novel materials discovered from a systematic analysis of empirical device data are shown to produce high quality white light through combination of monomer and excimer emission from a single molecule: platinum(II) bis(methyl-imidazolyl)toluene chloride (Pt-17). Illumination quality achieved Commission Internationale de L'Eclairage (CIE) chromaticity coordinates (x = 0.31, y = 0.38) and color rendering index (CRI) > 75. Further optimization of a device containing Pt-17 resulted in a maximum forward viewing power efficiency of 37.8 lm/W on a plain glass substrate. In addition, accelerated aging tests suggest high energy blue emission from a halogen-free cyclometalated platinum complex could demonstrate degradation rates comparable to known stable emitters. Finally, a buckling based metrology is applied to characterize the mechanical properties of small molecule organic thin films towards understanding the deposition kinetics responsible for an elastic modulus that is both temperature and thickness dependent. These results could contribute to the viability of organic electronic technology in potentially flexible display and lighting applications. The results also provide insight to organic film growth kinetics responsible for optical, mechanical, and water uptake properties relevant to engineering the next generation of optoelectronic devices.

Spontaneous Fluctuations can Guide Drug Design Strategies for Structurally Disordered Proteins.

PubMed

Maity, Barun Kumar; Vishvakarma, Vicky; Surendran, Dayana; Rawat, Anoop; Das, Anirban; Pramanik, Shreya; Arfin, Najmul; Maiti, Sudipta

2018-06-21

Structure-based 'rational' drug-design strategies fail for diseases associated with intrinsically disordered proteins (IDPs). However, structural disorder allows large amplitude spontaneous intramolecular dynamics in a protein. We demonstrate a method that exploits this dynamics to provide quantitative information about the degree of interaction of an IDP with other molecules. A candidate ligand molecule may not bind strongly, but even momentary interactions can be expected to perturb the fluctuations. We measure the amplitude and frequency of the equilibrium fluctuations of fluorescently labeled small oligomers of hIAPP (an IDP associated with Type II diabetes) in a physiological solution, using nanosecond fluorescence cross-correlation spectroscopy. We show that the inter-terminal distance fluctuates at a characteristic timescale of 134 ± 10 ns, and 6.4 ± 0.2 % of the population is in the 'closed' (quenched) state at equilibrium. These fluctuations are affected in a dose-dependent manner by a series of small molecules known to reduce the toxicity of various amyloid peptides. The degree of interaction shows the following order: resveratrol < epicatechin ~ quercetin < congo red < epigallocatechin-3-gallate. Such ordering can provide a direction for exploring the chemical space for finding stronger-binding ligands. We test the biological relevance of these measurements by measuring the effect of these molecules on the affinity of hIAPP for lipid vesicles and cell membranes. We find that the ability of a molecule to modulate intramolecular fluctuations correlates well with its ability to lower membrane affinity. We conclude that structural disorder may provide new avenues for rational drug design for IDPs.

Small-molecule photostabilizing agents are modifiers of lipid bilayer properties.

PubMed

Alejo, Jose L; Blanchard, Scott C; Andersen, Olaf S

2013-06-04

Small-molecule photostabilizing or protective agents (PAs) provide essential support for the stability demands on fluorescent dyes in single-molecule spectroscopy and fluorescence microscopy. These agents are employed also in studies of cell membranes and model systems mimicking lipid bilayer environments, but there is little information about their possible effects on membrane structure and physical properties. Given the impact of amphipathic small molecules on bilayer properties such as elasticity and intrinsic curvature, we investigated the effects of six commonly used PAs--cyclooctatetraene (COT), para-nitrobenzyl alcohol (NBA), Trolox (TX), 1,4-diazabicyclo[2.2.2]octane (DABCO), para-nitrobenzoic acid (pNBA), and n-propyl gallate (nPG)--on bilayer properties using a gramicidin A (gA)-based fluorescence quench assay to probe for PA-induced changes in the gramicidin monomer↔dimer equilibrium. The experiments were done using fluorophore-loaded large unilamellar vesicles that had been doped with gA, and changes in the gA monomer↔dimer equilibrium were assayed using a gA channel-permeable fluorescence quencher (Tl⁺). Changes in bilayer properties caused by, e.g., PA adsorption at the bilayer/solution interface that alter the equilibrium constant for gA channel formation, and thus the number of conducting gA channels in the large unilamellar vesicle membrane, will be detectable as changes in the rate of Tl⁺ influx-the fluorescence quench rate. Over the experimentally relevant millimolar concentration range, TX, NBA, and pNBA, caused comparable increases in gA channel activity. COT, also in the millimolar range, caused a slight decrease in gA channel activity. nPG increased channel activity at submillimolar concentrations. DABCO did not alter gA activity. Five of the six tested PAs thus alter lipid bilayer properties at experimentally relevant concentrations, which becomes important for the design and analysis of fluorescence studies in cells and model membrane systems. We therefore tested combinations of COT, NBA, and TX; the combinations altered the fluorescence quench rate less than would be predicted assuming their effects on bilayer properties were additive. The combination of equimolar concentrations of COT and NBA caused minimal changes in the fluorescence quench rate. Copyright © 2013 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Lessons from isolable nickel(I) precursor complexes for small molecule activation.

PubMed

Yao, Shenglai; Driess, Matthias

2012-02-21

Small-molecule activation by transition metals is essential to numerous organic transformations, both biological and industrial. Creating useful metal-mediated activation systems often depends on stabilizing the metal with uncommon low oxidation states and low coordination numbers. This provides a redox-active metal center with vacant coordination sites well suited for interacting with small molecules. Monovalent nickel species, with their d(9) electronic configuration, are moderately strong one-electron reducing agents that are synthetically attractive if they can be isolated. They represent suitable reagents for closing the knowledge gap in nickel-mediated activation of small molecules. Recently, the first strikingly stable dinuclear β-diketiminate nickel(I) precursor complexes were synthesized, proving to be suitable promoters for small-molecule binding and activation. They have led to many unprecedented nickel complexes bearing activated small molecules in different reduction stages. In this Account, we describe selected achievements in the activation of nitrous oxide (N(2)O), O(2), the heavier chalcogens (S, Se, and Te), and white phosphorus (P(4)) through this β-diketiminatonickel(I) precursor species. We emphasize the reductive activation of O(2), owing to its promise in oxidation processes. The one-electron-reduced O(2) activation product, that is, the corresponding β-diketiminato-supported Ni-O(2) complex, is a genuine superoxonickel(II) complex, representing an important intermediate in the early stages of O(2) activation. It selectively acts as an oxygen-atom transfer agent, hydrogen-atom scavenger, or both towards exogenous organic substrates to yield oxidation products. The one-electron reduction of the superoxonickel(II) moiety was examined by using elemental potassium, β-diketiminatozinc(II) chloride, and β-diketiminatoiron(I) complexes, affording the first heterobimetallic complexes featuring a [NiO(2)M] subunit (M is K, Zn, or Fe). Through density functional theory (DFT) calculations, the geometric and electronic structures of these complexes were established and their distinctive reactivity, including the unprecedented monooxygenase-like activity of a bis(μ-oxo)nickel-iron complex, was studied. The studies have further led to other heterobimetallic complexes containing a [NiO(2)M] core, which are useful for understanding the influence of the heterometal on structure-reactivity relationships. The activation of N(2)O led directly to the hydrogen-atom abstraction product bis(μ-hydroxo)nickel(II) species and prevented isolation of any intermediate. In contrast, the activation of elemental S, Se, and Te with the same nickel(I) reagent furnished activation products with superchalcogenido E(2)(-) (E is S, Se, or Te) and dichalcogenido E(2)(2-) ligand in different activation stages. The isolable supersulfidonickel(II) subunit may serve as a versatile building block for the synthesis of heterobimetallic disulfidonickel(II) complexes with a [NiS(2)M] core. In the case of white phosphorus, the P(4) molecule has been coordinated to the nickel(I) center of dinuclear β-diketiminatonickel(I) precursor complexes; however, the whole P(4) subunit is a weaker electron acceptor than the dichalcogen ligands E(2), thus remaining unreduced. This P(4) binding mode is rare and could open new doors for subsequent functionalization of P(4). Our advances in understanding how these small molecules are bound to a nickel(I) center and are activated for further transformation offer promise for designing new catalysts. These nickel-containing complexes offer exceptional potential for nickel-mediated transformations of organic molecules and as model compounds for mimicking active sites of nickel-containing metalloenzymes.