N-glycosylation is a common protein modification process, which affects a number of properties of proteins. Little is known about the distribution of N-glycosylation sequons, for example, the distance between glycosylated sites and their position in the protein primary sequence. Using a large set of experimentally ...

The gene encoding protein p32, the most abundant and immunogenic protein induced by African swine fever virus at early times of infection, has been mapped in the EcoRI C' fragment of the genome of the Vero cell-adapted virus strain BA71V. Sequencing analysis has shown the existence of an open reading frame, named C'204L, encoding 204 ...

ADP (also known as E3-11.6K protein) is synthesized abundantly in late adenovirus infection and is required for efficient lysis of infected cells and release of viral progeny at the end of the viral replication cycle. ADP is a type III bitopic N_endoC_exo nuclear membrane and Golgi glycoprotein that is produced ...

We describe a novel method for producing homogeneous eukaryotic N-glycoproteins. The method involves the engineering and functional transfer of the C. jejuni glycosylation machinery in E. coli to express glycosylated proteins with the key GlcNAc-Asn linkage. The bacterial glycans were then trimmed and remodeled in vitro by enzymatic transglycosylation to fulfill a eukaryotic ...

NCBE (SLC4A10) is a member of the SLC4 family of bicarbonate transporters, several of which play important roles in intracellular-pH regulation and transepithelial HCO(3)(-) transport. Here we characterize a new antibody that was generated in rabbit against a fusion protein consisting of maltose-binding protein and the first 135 amino acids (aa) of the ...

Recent years have witnessed a rapid growth in the number and diversity of prokaryotic proteins shown to carry N- and/or O-glycans, with protein glycosylation now considered as fundamental to the biology of these organisms as it is in eukaryotic systems. This article overviews the major glycosylation pathways that are known to exist in eukarya, bacteria and ...

Autoimmune diseases are among the most prevalent of afflictions, yet the genetic factors responsible are largely undefined. Protein glycosylation in the Golgi apparatus produces structural variation at the cell surface and contributes to immune self-recognition. Altered protein glycosylation and antibodies that recognize endogenous glycans have been ...

Bm95 is an antigen isolated from Boophilus microplus strains with low susceptibility to antibodies developed in cattle vaccinated with the recombinant Bm86 antigen (Gavac, HeberBiotec S.A., Cuba). It is a Bm86-like surface protein, which by similarity contains seven EGF-like domains and a lipid-binding GPI-anchor site at the C-terminal region. The primary structure of the ...

Conservation of Pax gene expression in ectodermal placodes of the lamprey ... Location of 6 cysteine and 4 consensus N-glycosylation sites in the bovine sequence were identical to those in ... Evolution of calcium modulated proteins ...

Protein post-translational modifications (PTMs) are regulated separately from protein expression levels. Thus, simultaneous characterization of the proteome and its PTMs is pivotal to an understanding of protein regulation, function and activity. However, concurrent analysis of the proteome and its PTMs by mass spectrometry is a ...

The enormous complexity, wide dynamic range of relative protein abundance of interest (over 10 orders of magnitude), and tremendous heterogeneity (due to post-translational modifications, such as glycosylation) of the human blood plasma proteome severely challenges the capabilities of existing analytical methodologies. We describe here the comprehensive ...

In most polarized cells, the Na,K-ATPase is localized on the basolateral plasma membrane. However, an unusual location of the Na,K-ATPase was detected in polarized HGT-1 cells (a human gastric adenocarcinoma cell line). The Na,K-ATPase alpha1 subunit was detected along with the beta2 subunit predominantly on the apical membrane, whereas the Na,K-ATPase beta1 subunit was not found in HGT-1 cells. ...

N-Glycans attached to the ectodomains of plasma membrane pattern recognition receptors constitute likely initial contact sites between plant cells and invading pathogens. To assess the role of N-glycans in receptor-mediated immune responses, we investigated the functionality of Arabidopsis receptor kinases EFR and FLS2, sensing bacterial translation elongation factor Tu (elf18) and flagellin ...

N-Glycans attached to the ectodomains of plasma membrane pattern recognition receptors constitute likely initial contact sites between plant cells and invading pathogens. To assess the role of N-glycans in receptor-mediated immune responses, we investigated the functionality of Arabidopsis receptor kinases EFR and FLS2, sensing bacterial translation elongation factor Tu (elf18) and flagellin ...

N-glycosylation of E-cadherin has been shown to inhibit cell�cell adhesion. Specifically, our recent studies have provided evidence that the reduction of E-cadherin N-glycosylation promoted the recruitment of stabilizing components, vinculin and serine/threonine protein phosphatase 2A (PP2A), to adherens junctions (AJs) and enhanced ...

The human sodium-dependent vitamin C transporters (hSVCT1 and hSVCT2) mediate cellular uptake of ascorbic acid. Both these transporters contain potential sites for N-glycosylation in their extracellular domains (Asn-138, Asn-144 [hSVCT1]; Asn-188, Asn-196 [hSVCT2]), however the role of N-glycosylation in transporter function is unexplored. On the basis of the result that tunicamycin decreased {sup ...

This study illustrates multifunctionality of proteins of honeybee royal jelly (RJ) and how their neofunctionalization result from various PTMs of maternal proteins. Major proteins of RJ, designated as apalbumins belong to a protein family consisting of nine members with M(r) of 49-87 kDa and they are accompanied by ...

While Eukarya, Bacteria, and Archaea are all capable of protein N glycosylation, the archaeal version of this posttranslational modification is the least understood. To redress this imbalance, recent studies of the halophilic archaeon Haloferax volcanii have identified a gene cluster encoding the Agl proteins ...

Germin-like proteins (GLPs) ionically bound to the walls of preglobular somatic embryos of Pinus caribaea Morelet are markers of this early developmental stage. In order to reveal the physiological implications of such markers during early embryo development, we isolated a cDNA clone from somatic embryos predicted to encode a protein with sequence ...

The chitin elicitor-binding protein (CEBiP) from rice was the first plant lysin motif (LysM) protein for which the biological and biochemical function had been established. It belongs to a plant-specific family of extracellular LysM proteins (LYMs) for which we analyzed the phylogeny. LYMs are present in vascular plants only, where an ...

BackgroundThe addition of N-linked glycans to proteins is normally a cotranslational process that occurs during translocation of the nascent protein to the endoplasmic reticulum. Here, we report on an exception to this rule occurring on the hepatitis B virus (HBV) large L envelope protein that is a subject to co-plus posttranslational ...

Reserved. Posttranslational Protein Modification in Archaea Jerry Eichler1 * and Michael W. W. Adams2.....................................................................................................................................................397 Process of Protein N-Glycosylation in ...

ABSTRACT Glypicans are cell-surface heparan sulfate proteoglycans that regulate developmental signaling pathways by binding growth factors to their heparan sulfate chains. The primary structures of glypican core proteins contain potential N-glycosylation sites but the importance of N-glycosylation in glypicans has never been investigated in detail. Here, ...

N-Glycosylation of membrane proteins is critical for their proper folding, co-assembly and subsequent matriculation through the secretory pathway. Here, we examine the kinetics of N-glycan addition to type I transmembrane KCNE1 K(+) channel ?-subunits, where point mutations that prevent N-glycosylation at one consensus site give rise ...

N-glycosylation, a common cotranslational modification, is thought to be critical for plasma membrane expression of glycoproteins by enhancing protein folding, trafficking, and stability through targeting them to the ER folding cycles via lectin-like chaperones. In this study, we show that N-glycans, specifically core glycans, enhance the productive ...

Although many eukaryotic proteins have been secreted by transfected bacterial cells, little is known about how a bacterial protein is treated as it passes through the secretory pathway when expressed in a eukaryotic cell. The eukaryotic N-glycosylation system could interfere with folding and secretion of prokaryotic ...

AbstractAlthough many eukaryotic proteins have been secreted by transfected bacterial cells, little is known about how a bacterial protein is treated as it passes through the secretory pathway when expressed in a eukaryotic cell. The eukaryotic N-glycosylation system could interfere with folding and secretion of prokaryotic ...

Recently, the prospect of using Escherichia coli as a host for human glycoprotein production has increased due to detailed characterization of the prokaryotic N-glycosylation process and the ability to transfer the system into this bacterium. Although functionality of the native Campylobacter jejuni N-glycosylation system in E. coli has been demonstrated, ...

Recently, the prospect of using Escherichia coli as a host for human glycoprotein production has increased due to detailed characterization of the prokaryotic N-glycosylation process and the ability to transfer the system into this bacterium. Although functionality of the native Campylobacter jejuni N-glycosylation system in E. coli has been demonstrated, ...

We describe the identification of a widely expressed receptor-type (transmembrane) protein tyrosine phosphatase (PTPase; EC 3.1.3.48). Screening of a mouse brain cDNA library under low-stringency conditions with a probe encompassing the intracellular (phosphatase) domain of the CD45 lymphocyte antigen yielded cDNA clones coding for a 794-amino acid transmembrane ...

BackgroundYeast expression systems with altered N-glycosylation are now available to produce glycoproteins with homogenous, defined N-glycans. However, data on the behaviour of these strains in high cell density cultivation are scarce.ResultsHere, we report on cultivations under controlled specific growth rate of a GlycoSwitch-Man5 Pichia pastoris strain producing ...

Earlier the authors have shown that ..beta..-adrenoreceptor stimulation increases protein N-glycosylation in parotid acinar cells via a cAMP-mediated process, by activating key glycosyltransferases of the Dol-P linked pathway. Also, the authors have observed approx.50% increase in Man-P-Dol synthase activity in membranes phosphorylated in vitro with ...

Numerous studies have recently focused on the identification of specific glycan biomarkers; given the important roles that protein linked glycans play, for example, during development and disease progression. The identification of protein glycobiomarkers, which are part of a very complex proteome, has involved the use of fractionation techniques such as ...

Page 1. Influence of Protein Abundance on High-Throughput Protein-Protein Interaction Detection ... The previously noted trend between protein ...

Human {beta}1,3-N-acetylglucosaminyltransferase 2 ({beta}3GnT2) is thought to be an enzyme that extends the polylactosamine acceptor chains, but its function and structure analysis are unknown. To obtain insight into the structure of {beta}3GnT2, the effects of N-glycosylation on its biological function were evaluated using the addition of inhibitors, site-directed mutagenesis of potential ...

N-Glycosylation is normally a co-translational process that occurs as soon as a nascent and unfolded polypeptide chain has emerged ?12 residues into the lumen of the endoplasmic reticulum (ER). Here, we describe the efficient utilization of an N-glycosylation site engineered within the luminal extreme C-terminal residues of distinct integral membrane glycoproteins, a native ER ...

The ascomycete fungus Mycosphaerella graminicola is the causal agent of Septoria Tritici Blotch disease of wheat and can grow as yeast-like cells or as hyphae depending on environmental conditions. Hyphal growth is however essential for successful leaf infection. A T-DNA mutagenesis screen performed on haploid spores identified a mutant, which can undergo yeast-like growth but cannot switch to ...

Intracellular trafficking and subcellular deposition are critical factors influencing the accumulation and posttranslational modifications of proteins. In seeds, these processes are not yet fully understood. In this study, we set out to investigate the intracellular transport, final destination, N-glycosylation status, and stability of the fusion of ...

Intracellular trafficking and subcellular deposition are critical factors influencing the accumulation and posttranslational modifications of proteins. In seeds, these processes are not yet fully understood. In this study, we set out to investigate the intracellular transport, final destination, N-glycosylation status, and stability of the fusion of ...

Prostate-specific membrane antigen (PSMA) is a membrane protein that is over-expressed manifold in prostate cancer and provides an attractive target for therapy. PSMA ADC is an antibody-drug conjugate (ADC) that consists of a fully human anti-PSMA monoclonal antibody conjugated to monomethylauristatin E through a valine-citrulline linker. In this study, the antitumor activity ...

A cDNA (Cel1) encoding an endo-1,4-beta-glucanase (EGase) was isolated from ripe fruit of strawberry (Fragaria x ananassa). The deduced protein of 496 amino acids contains a presumptive signal sequence, a common feature of cell wall-localized EGases, and one potential N-glycosylation site. Southern- blot analysis of genomic DNA from F. x ananassa, an ...

A cDNA (Cel1) encoding an endo-1,4-?-glucanase (EGase) was isolated from ripe fruit of strawberry (Fragaria � ananassa). The deduced protein of 496 amino acids contains a presumptive signal sequence, a common feature of cell wall-localized EGases, and one potential N-glycosylation site. Southern- blot analysis of genomic DNA from F. � ananassa, ...

Vitellogenin, an egg-yolk protein precursor common to oviparous animals, is found abundantly in honeybee workers - a caste of helpers that do not usually lay eggs. Instead, honeybee vitellogenin (180 kDa) participates in processes other than reproduction: it influences hormone signaling, food-related behavior, immunity, stress resistance and longevity. The ...

Recently, significant attention has been drawn to the biology of small leucine-rich repeat proteoglycans (SLRPs) due to their multiple functionalities in various cell types and tissues. Here, we characterize a novel SLRP member, "Podocan-like (Podnl) protein" identified by a bioinformatics approach. The Podnl protein has a signal peptide, a unique ...

of the association of a 110-kDa peripheral membrane protein with the Golgi apparatus, J. Cell Biol. 112 (1991) 579 is known to induce an effective collapse of the Golgi complex into the ER, the hypothesis that a prolonged by the coalescence of the Golgi into the ER caused by BFA. � 2002 Elsevier Science (USA). All rights reserved

Plant-specific N-glycosylation can represent an important limitation for the use of recombinant glycoproteins of mammalian origin produced by transgenic plants. Comparison of plant and mammalian N-glycan biosynthesis indicates that beta1,4-galactosyltransferase is the most important enzyme that is missing for conversion of typical plant N-glycans into mammalian-like N-glycans. ...

Plant-specific N-glycosylation can represent an important limitation for the use of recombinant glycoproteins of mammalian origin produced by transgenic plants. Comparison of plant and mammalian N-glycan biosynthesis indicates that ?1,4-galactosyltransferase is the most important enzyme that is missing for conversion of typical plant N-glycans into mammalian-like ...

Glycosylation is one of the most important posttranslational modifications affecting the functions of proteins and cell activities. Mass spectrometry (MS) has proven to be an effective tool for structural glycobiology and has helped gain an understanding of glycoprotein-mediated diseases. Although electro-spray ionization-tandem MS remains widely recognized as an effective ...

Pregnancy-associated glycoproteins (PAGs) are major secretory proteins of trophoblast cells in ruminants. Binucleate trophoblast giant cells (BNCs) store these proteins in secretory granules and release them into the maternal organism after fusion with maternal uterine epithelial cells. By matrix assisted laser desorption ionisation-mass spectrometry ...

Ebola virus (EBOV), an enveloped, single-stranded, negative-sense RNA virus, causes severe hemorrhagic fever in humans and nonhuman primates. The EBOV glycoprotein (GP) gene encodes the nonstructural soluble glycoprotein (sGP) but also produces the transmembrane glycoprotein (GP?,?) through transcriptional editing. A third GP gene product, a small soluble glycoprotein (ssGP), has long been ...

The open reading frame III of Borna disease virus (BDV) codes for a protein with a mass of 16 kDa, named p16 or BDV-M. p16 was described as an N-glycosylated protein in several previous publications and therefore was termed gp18, although the amino acid sequence of p16 does not contain any regular consensus sequence for N ...

N-glycosylated proteins were isolated from Arabidopsis thaliana mature stems using affinity chromatography on Concanavalin A Sepharose, separated by 2D-electrophoresis and identified using nanoHPLC-MS/MS and MALDI-TOF MS. 102 glycoproteins were identified. 94% of these proteins were predicted by bioinformatics to be targeted to the ...

The plasma phospholipid transfer protein (PLTP) plays a key role in lipid and lipoprotein metabolism. It has six potential N-glycosylation sites. To study the impact of these sites on PLTP secretion and activity, six variants containing serine to alanine point mutations were prepared by site-directed mutagenesis and expressed in Chinese hamster ovary ...

Recent insight into the N-glycosylation pathway of the haloarchaeon, Haloferax volcanii, is helping to bridge the gap between our limited understanding of the archaeal version of this universal post-translational modification and the better-described eukaryal and bacterial processes. To delineate as yet undefined steps of the Hfx.?volcanii N-glycosylation pathway, a ...

A commercially available polyclonal antibody against a rClC-3/GST fusion protein was used in order to investigate the tissue distribution of the ClC-3 chloride channel protein. The antibody appeared to be specific to rClC-3 since no cross-reaction could be observed with rClC-4 or rClC-5 proteins when overexpressed in Xenopus oocytes. ...

... biases that arise from the varying cellular abundances of the proteins. ... and Escherichia coli incorporate a correlation between protein degree, or ...

Human complement factor H (FH), an abundant 155-kDa plasma glycoprotein with 40 disulphide bonds, regulates the alternative-pathway complement cascade. Mutations and single nucleotide polymorphisms in the FH gene predispose to development of age-related macular degeneration, atypical haemolytic uraemic syndrome and dense deposit disease. Supplementation with FH variants ...

SummaryNatural killer (NK) cell-mediated resistance to murine cytomegalovirus (MCMV) is controlled by allelic Ly49 receptors, including activating Ly49H (C57BL/6 strain) and inhibitory Ly49I (129 strain), which specifically recognize MCMV m157, a glycosylphosphatidylinositol-linked protein with homology to MHC class I. Although the Ly49 receptors retain significant homology to ...

In eukaryotic cells, N-glycosylation has been recognized as one of the most common and functionally important co- or post-translational modifications of proteins. �Free� forms of N-glycans accumulate in the cytosol of mammalian cells, but the precise mechanism for their formation and degradation remains unknown. Here, we report a method for the ...

Trypanosoma cruzi epimastigotes (insect gut stage) incubated with (U- UC)glucose synthesized Man9GlcNAc2-P-P-dolichol as practically the sole dolichol-P-P derivative. On the other hand, amastigotes (intracellular stage) of the same parasite synthesized four to five times more Man7GlcNAc2-P-P-dolichol than Man9GlcNAc2-P-P-dolichol. Evidence is presented indicating that, whereas in epimastigotes ...

Dolichol monophosphate (Dol-P) functions as an obligate glycosyl carrier lipid in protein glycosylation reactions. Dol-P is synthesized by the successive condensation of isopentenyl diphosphate (IPP), with farnesyl diphosphate catalysed by a cis-isoprenyltransferase (cis-IPTase) activity. Despite the recognition of cis-IPTase activity 40 years ago and the molecular cloning of ...

Although several lysosomal membrane glycoproteins have been characterized by using specific antibodies, none of the studies so far elucidated the amino acid sequence of a lysosomal membrane glycoprotein. Here we describe cDNA clones encoding for one of the lysosome-associated membrane proteins with apparent M{sub r} {approx} 120,000, lamp A. The amino acid sequence based on ...

Two experiments were conducted to determine the effects of ruminal protein degradability, supplementation frequency, and increasing dietary protein on the expression and distribution of urea transporter-B (UT-B) in lambs fed low-quality forage (mature crested wheatgrass hay; 4.2 to 4.7% CP). In Exp. 1, 15 Dorset wether lambs (initial BW=45.8+/-1.3 kg) were ...

Hepatitis delta virus (HDV) particles are coated with the large (L), middle (M), and small (S) hepatitis B virus envelope proteins. In the present study, we constructed glycosylation-defective envelope protein mutants and evaluated their capacity to assist in the maturation of infectious HDV in vitro. We observed that the removal of N-linked carbohydrates ...

Seed-specific expression is an appealing alternative technology for the production of recombinant proteins in transgenic plants. Whereas attractive yields of recombinant proteins have been achieved by this method, little attention has been paid to the intracellular deposition and the quality of such products. Here, we demonstrate a comparative study of two ...

The core oligosaccharide Glc(3)Man(9)GlcNAc(2) is assembled by a series of membrane-bound glycosyltransferases as the lipid carrier dolichylpyrophosphate-linked glycan in the endoplasmic reticulum (ER). The first step of this assembly pathway on the ER luminal side is mediated by ALG3 (asparagine-linked glycosylation 3), which is a highly conserved reaction among eukaryotic cells. Complementary ...

The prM protein of Japanese encephalitis virus (JEV) contains a single potential N-linked glycosylation site, N(15)-X(16)-T(17), which is highly conserved among JEV strains and closely related flaviviruses. To investigate the role of this site in JEV replication and pathogenesis, we manipulated the RNA genome by using infectious JEV cDNA to generate three prM mutants (N15A, ...

The prM protein of Japanese encephalitis virus (JEV) contains a single potential N-linked glycosylation site, N¹⁵-X¹⁶-T¹⁷, which is highly conserved among JEV strains and closely related flaviviruses. To investigate the role of this site in JEV replication and pathogenesis, we manipulated the RNA genome by ...

Protein N-glycosylation requires the presence of asparagine (N) in the consensus tri-peptide NXS/T (where X is any amino acid, S is serine and T is threonine). Several factors affect the glycosylation potential of NXS/T sequons and one such factor is the type of amino acid at position X. While proline was shown to negatively affect ...

PI3K and PTEN lipid phosphatase control the level of cellular phosphatidylinositol (3,4,5)-trisphosphate, an activator of AKT kinases that promotes cell growth and survival. Mutations activating AKT are commonly observed in human cancers. We report here that ENTPD5, an endoplasmic reticulum (ER) enzyme, is upregulated in cell lines and primary human tumor samples with active AKT. ENTPD5 hydrolyzes ...

Misfolding and subsequent aggregation of endogeneous proteins constitute essential steps in many human disorders, including Alzheimer and prion diseases. In most prion protein-folding studies, the posttranslational modifications, the lipid anchor in particular, were lacking. Here, we studied a fully posttranslationally modified cellular prion ...

The selectivity and individual roles of the N-linked oligosaccharide chains of Sendai virus fusion protein (F protein) in the interaction with endoplasmic reticulum molecular chaperones were investigated by analyses of transient expression of single N-glycosylation mutants and sequential immunoprecipitation. We demonstrated ...

Among the Dengue virus structural proteins, the Envelope glycoprotein is the most important because of its antigenic characteristics. In this work, the E protein from Dengue-2 virus truncated at the C-terminus region was successfully expressed in Pichia pastoris. The E2trunc gene was cloned under the AOX1 promoter from P. pastoris and the signal peptide of ...

During the generation of Expressed Sequence Tags (ESTs) from the Fer de Lance viper (Bothrops jararaca) venom glands, a partial cDNA (clone H8) coding for a protein with all the features of a paramyxovirus fusion protein was characterized. It has 920 bp and codes for a partial protein of 279 amino acids. Two potential ...

Genetic information flows from DNA to macromolecular structures�the dominant force in the molecular organization of life. However, recent work suggests that metabolite availability to the hexosamine and Golgi N-glycosylation pathways exerts control over the assembly of macromolecular complexes on the cell surface, and in this capacity, acts upstream of signaling and gene ...

Ovomucin is a bioactive egg white glycoprotein responsible for the gel properties of fresh egg white and is believed to be involved in egg white thinning, a natural process that occurs during storage. Ovomucin is composed of two subunits: a carbohydrate-rich ?-ovomucin with molecular weight of 400-610�KDa and a carbohydrate-poor ?-ovomucin with molecular mass of 254�KDa. In addition to limited ...

Toxoplasma gondii motility, which is essential for host cell entry, migration through host tissues and invasion, is a unique form of actin-dependent gliding. It is powered by a motor complex mainly composed of myosin heavy chain A (MyoA), myosin light chain 1 (MLC1), gliding associated proteins GAP45 and GAP50, the only integral membrane anchor so far described. In the present ...

The cellular prion protein (PrP^C) plays a fundamental role in prion disease. PrP^C is a glycosylphosphatidylinositol (GPI)-anchored protein with two variably occupied N-glycosylation sites. In general, GPI-anchor and N-glycosylation direct proteins to apical membranes in ...

Lipopolysaccharide (LPS) is a major component on the surface of Gram negative bacteria and is composed of lipid A-core and the O antigen polysaccharide. O polysaccharides of the gastric pathogen Helicobacter pylori contain Lewis antigens, mimicking glycan structures produced by human cells. The interaction of Lewis antigens with human dendritic cells induces a modulation of the immune response, ...

Glycoengineering is increasingly being recognized as a powerful tool to generate recombinant glycoproteins with a customized N-glycosylation pattern. Here, we demonstrate the modulation of the plant glycosylation pathway toward the formation of human-type bisected and branched complex N-glycans. Glycoengineered Nicotiana benthamiana lacking plant-specific ...

We sequenced the genome of LATC06 generated by in vitro passage in Vero cells of the lapinized-avianized (LA) strain and compared its sequence to those of other rinderpest viruses. The LATC06 genome consists of 15882 nucleotides. Its transcriptional regulatory control sequences (TRSs) at gene boundaries are identical to those of the Kabete O strain. Cleavage sites for generating F1/F2 ...

We performed here MS-based cell surface proteome profiling of HCT-116 cells by two distinct methods based on biotin labeling and glycoprotein capturing. In total, 742 biotinylated and 219 glycosylated proteins were identified by the biotin labeling and glycoprotein capturing, of which 224 and 138 proteins known to be located on plasma membrane were ...

Polyprenoids, polymers containing varied numbers of isoprene subunits, serve numerous roles in biology. In Eukarya, dolichyl phosphate, a phosphorylated polyprenol bearing a saturated ?-end isoprene subunit, serves as the glycan carrier during N-glycosylation, namely that post-translational modification whereby glycans are covalently linked to select asparagine residues of a ...

Protein modifications of death receptor pathways play a central role in the regulation of apoptosis. It has been demonstrated that O-glycosylation of TRAIL-receptor (R) is essential for sensitivity and resistance towards TRAIL-mediated apoptosis. In this study we ask whether and how glycosylation of CD95 (Fas/APO-1), another death receptor, influences DISC formation and ...

Ansamitocins are potent antitumor maytansinoids produced by Actinosynnema pretiosum. Their biosynthesis involves the initial assembly of a macrolactam polyketide, followed by a series of postpolyketide synthase (PKS) modifications. Three ansamitocin glycosides were isolated from A. pretiosum and fully characterized structurally as novel ansamitocin derivatives, carrying a beta-D-glucosyl group ...

Chitin deacetylase (CDA), the enzyme that catalyzes the hydrolysis of acetamido groups of GlcNAc in chitin, was purified from culture filtrate of the fungus Mortierella sp. DY-52 and characterized. The extracellular enzyme is likely to be a highly N-glycosylated protein with a pI of 4.2-4.8. Its apparent molecular weight was determined to be about 52 kDa ...

The 5-hydroxytryptamine 3 (5-HT(3)) receptor is a pentameric ligand-gated ion channel with potential molecular isoforms arising from different subunit combinations and/or different post-translational modifications of the individual subunits. Since N-glycosylation of the 5-HT3A subunit impacts cell surface trafficking, the presence of N-glycosylation of the human (h) 5-HT3B ...

A great majority of G protein-coupled receptors are modified by N-glycosylation, but the functional significance of this modification for receptor folding and intracellular transport has remained elusive. Here we studied these phenomena by mutating the two N-terminal N-glycosylation sites (Asn(18) and Asn(33)) of the human delta-opioid receptor, and ...

A great majority of G protein-coupled receptors are modified by N-glycosylation, but the functional significance of this modification for receptor folding and intracellular transport has remained elusive. Here we studied these phenomena by mutating the two N-terminal N-glycosylation sites (Asn¹⁸ and Asn³³) ...

GPCRs (G-protein-coupled receptors) are preferentially N-glycosylated on ECL2 (extracellular loop 2). We previously showed that N-glycosylation of ECL2 was crucial for cell-surface expression of the hAT₁ receptor (human angiotensin II receptor subtype 1). Here, we ask whether positioning of the N-glycosylation sites within the ...

XylT (?1,2-xylosyltransferase) is a unique Golgi-bound glycosyltransferase that is involved in the biosynthesis of glycoprotein-bound N-glycans in plants. To delineate the catalytic domain of XylT, a series of N-terminal deletion mutants was heterologously expressed in insect cells. Whereas the first 54 residues could be deleted without affecting the catalytic activity of the enzyme, removal of an ...

Nucleolin is a major nucleolar protein involved in fundamental processes of ribosome biogenesis, regulation of cell proliferation and growth. Nucleolin is known to shuttle between nucleus, cytoplasm and cell surface. We have previously found that nucleolin undergoes complex N- and O-glycosylations in extra-nuclear isoforms. We found that surface nucleolin is exclusively ...

The possible role of post-translational modifications such as subunit oligomerization, protein glycosylation and oligosaccharide processing on the circulatory life-time of proteins was studied using recombinant human acetylcholinesterase (rHuAChE). Different preparations of rHuAChE containing various amounts of tetramers, dimers and monomers are cleared at ...

BackgroundT cell immunoglobulin and mucin domain containing 3 protein (Tim-3) expressed on terminally differentiated Th1 cells plays a suppressive role in Th1-mediated immune responses. Recently, it has been shown that N-glycosylation affects the binding activity of the Tim-3-Ig fusion protein to its ligand, galectin-9, but the binding properties of ...

Two genes encoding the auxin-binding protein (ABP1) of tobacco (Nicotiana tabacum L.), both of which possess the characteristics of a luminal protein of the endoplasmic reticulum (ER), were isolated and sequenced. These genes were composed of at least five exons and four introns. The two coding exons showed 95% sequence homology and coded for two precursor ...

Mannose for mammalian glycan biosynthesis can be imported directly from the medium, derived from glucose or salvaged from endogenous or external glycans. All pathways must generate mannose 6-phosphate, the activated form of mannose. Imported or salvaged mannose is directly phosphorylated by hexokinase, whereas fructose 6-phosphate from glucose is converted to mannose 6-phosphate by phosphomannose ...

Glycosylation is one of the most common yet diverse post-translational modifications. Information on glycan heterogeneity and glycosite occupancy is increasingly recognized as crucial to understanding glycoprotein structure and function. Yet, no approach currently exists with which to holistically consider both the proteomic and glycomic aspects of a system. Here, we developed a novel method of ...

Three different analytical techniques (planar SDS-PAGE, CGE-on-a-chip and MALDI-TOF-MS) applied for determination of the molecular weight of intact and partly and completely de-N-glycosylated human serum glycoproteins (antithrombin III and coagulation factor IX) have been compared. N-Glycans were removed from the protein backbone of both complex ...

BackgroundSpecific immunological unresponsiveness to alloantigens can be induced in vivo by treating mice with a donor alloantigen in combination with a non-depleting anti-CD4 antibody. This tolerance induction protocol enriches for alloantigen reactive regulatory T cells (Treg). We previously demonstrated that alpha-1,2-mannosidase, an enzyme involved in the synthesis and processing of N-linked ...

CmCatB, a cowpea bruchid cathepsin B-like cysteine protease, facilitates insects coping with dietary protease inhibitor challenge. Expression of recombinant CmCatB using a Pichia pastoris system yielded an enzymatically active protein that was heterogeneously glycosylated, migrating as a smear of > or =50kDa on SDS-PAGE. Treatment with peptide:N-glycosidase F indicated that ...

Optimizing the amounts of proteins required to separate and characterize both abundant and less abundant proteins by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) is critical for conducting proteomic research. In this study, we tested five different levels of soybean seed ...

BackgroundBiosynthesis of the dolichol linked oligosaccharide (DLO) required for protein N-glycosylation starts on the cytoplasmic face of the ER to give Man₅GlcNAc₂-PP-dolichol, which then flips into the ER for further glycosylation yielding mature DLO ...

Progress has been achieved in three areas. N-glycosylation has been shown not to be absolutely essential for maintenance of activity of cellulase enzymes through both inhibitor (Tunicamycin) and enzyme (Endo-H) degradative studies. Thus deglycosylated cellulase may be functional as a direct transcript from mRNA. However, initial studies of in vitro synthesis of cellulase, ...

The first gene coding for an amino-acid permease of Candida albicans was sequenced. The DNA fragment complementing the lysine-permease deficiency was 3385 bp long. An open reading frame of 1713 nucleotides was found encoding a protein of 571 amino acids, with a calculated molecular weight of 63,343. Analysis of the deduced primary structure revealed ten membrane spanning ...

Cotranslational N-glycosylation can accelerate protein folding, slow protein unfolding, and increase protein stability, but the molecular basis for these energetic effects is incompletely understood. N-glycosylation of proteins at na�ve sites could be a useful strategy ...

The Campylobacter jejuni pgl gene cluster encodes a complete N-linked protein glycosylation pathway that can be functionally transferred into Escherichia coli. In this system, we analyzed the interplay between N-linked glycosylation, membrane translocation and folding of acceptor proteins in bacteria. We developed a recombinant N-glycan acceptor peptide ...

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BackgroundHuman pathogenic coronavirus NL63 (hCoV-NL63) is a group 1 (alpha) coronavirus commonly associated with respiratory tract infections. In addition to known non-structural and structural proteins all coronaviruses have one or more accessory proteins whose functions are mostly unknown. Our study focuses on hCoV-NL63 open reading frame 3 (ORF 3) ...

BackgroundDentin sialophosphoprotein (Dspp) is a multidomain, secreted protein that is critical for the formation of tooth dentin. Mutations in DSPP cause inherited dentin defects categorized as dentin dysplasia type II and dentinogenesis imperfecta type II and type III. Dentin sialoprotein (Dsp), the N-terminal domain of dentin sialophosphoprotein (Dspp), is a highly ...

Protein translocation across the membrane of the endoplasmic reticulum (ER) proceeds through a proteinaceous translocation machinery, the translocon. To identify components that may regulate translocation by interacting with nascent polypeptides in the translocon, we used site-specific photo-crosslinking. We found that a region C-terminal of the two ...

Germline mutations may cause human disease by various mechanisms. Missense and other in-frame mutations may be deleterious because the mutant proteins are not correctly targeted, do not function correctly, or both. We studied a child with mycobacterial disease caused by homozygosity for a novel in-frame microinsertion in IFNGR2. In cells transfected with the mutant allele, ...

A live attenuated influenza A/Vietnam/1203/2004 (H5N1) vaccine virus (VN04 ca) has receptor binding specificity to ?2,3-linked sialosides (?2,3SAL), and a single dose induces a minimal serum antibody response in mice and ferrets. In contrast, A/Hong Kong/213/2003 (H5N1) vaccine virus (HK03 ca) binds to both ?2,6SAL and ?2,3SAL and generates a stronger serum antibody response in animals. Among the ...

... authored a paper titled "N-glycosylation and microtubule ... of the ECM thus modulating the motility ... specific membrane antigen expression in normal ...

Recent developments in genomics and proteomics provide evidence that yeast and other fungal cell walls share a common origin. The fibrous component of yeast cell walls usually consists of ?-glucan and/or chitin. N-glycosylated proteins form an amorphous, cross-linking matrix as well as fibres on the outer surfaces of the walls. While the enzymes ...

GP300 is a high molecular weight glycoprotein of the bovine lungworm Dictyocaulus viviparus. The N-linked glycans are substituted with phosphorylcholine (PC) giving it immunomodulatory potential. GP300 is highly immunogenic and its recognition by IgE antibodies is correlated with protection against infection. Here we identified and characterized the protein backbone of GP300. ...

Glycoproteins with asparagine-linked (N-linked) glycans occur in all eukaryotic cells. The function of their glycan moieties is one of the central problems in contemporary cell biology. N-glycosylation may modify physicochemical and biological protein properties such as conformation, degradation, intracellular sorting or secretion. We have isolated and ...