SEARCH · PubMed Health

Results for “posttranslational modification”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 145 records · Page 8Linked to original sources

PubMed1991

Characterization of posttranslational modifications in neuron-specific class III beta-tubulin by mass spectrometry.

Class III beta-tubulin, isolated from adult bovine brain, is resolved into at least seven charge variants on isoelectric focusing gels. To identify the posttranslational modifications responsible for this heterogeneity, a mixture of brain tubulins was treated with cyanogen bromide and the C-terminal fragments from the class III beta-tubulin isoforms were then isolated by binding them to the monoclonal antibody TuJ1. Combined use of tandem mass spectrometry and both subtractive and automated Edman degradation chemistry on the isolated peptides indicates that many of the isoforms differ by phosphorylation at Ser-444 plus attachment of one to six glutamic acid molecules to the side chain of the first glutamate residue, Glu-438, in the C-terminal sequence Tyr-Glu-Asp-Asp-Glu-Glu-Glu-Ser-glu-Ala-Gln-Gly-Pro-Lys.

Amino Acid Sequence↗

PubMed2006

Nitric oxide and posttranslational modification of the vascular proteome: S-nitrosation of reactive thiols.

Nitric oxide (NO*) is known to exert its effects via guanylyl cyclase and cyclic GMP-dependent pathways and by cyclic GMP-independent pathways, including the posttranslational modification of proteins. Much ongoing research is focused on defining the mechanisms of NO*-mediated protein modification, the identity and function of the modified proteins, and the significance of these changes in health and disease. S-nitrosation or thionitrite formation has only been found on a limited number of residues in a subset of proteins in in vitro and in vivo studies. Protein S-nitrosation also appears to be reversible. There are several theories about the in vivo S-nitrosating agent, and most suggest a role for oxidation products of NO* in this process. Flux in cellular S-nitrosoprotein pools appears to be regulated by NO* availability and is redox-sensitive. An analysis of S-nitrosation in candidate proteins has clarified the mechanism by which NO* regulates enzymatic and cellular functions. These findings suggest the utility of using proteomic methods to identify unique targets for protein S-nitrosation to understand further the molecular mechanisms of the effects of NO*.

Animals↗

PubMed2003

Posttranslational modification of MDM2.

The functions of the MDM2 protein, in particular its E3 ubiquitin ligase activity and its ability to interact with a number of cellular proteins intimately involved in growth regulation, are modulated by sumoylation and multisite phosphorylation. These posttranslational mechanisms not only regulate the intrinsic activity of MDM2 in response to cellular stresses, but also govern its subcellular localization, differentiate between MDM2-mediated ubiquitination of p53 and autoubiquitination, integrate the stress response with mechanisms that mediate cell survival, and modulate the interaction of MDM2 with cellular and viral proteins. In this review, we summarize our current knowledge of the role of posttranslational modifications of MDM2 and their functional relevance.

Humans↗

PubMed1986

Posttranslational modification at the N terminus of the human adenovirus type 12 E1A 235R tumor antigen.

The adenovirus E1A transforming region, which encodes immortalization, partial cell transformation, and gene activation functions, expresses two early mRNAs, 13S and 12S. Multiple-T antigen species with different electrophoretic mobilities are formed from each mRNA, presumably by unknown posttranslational modifications. The adenovirus type 12 (Ad12) 13S and 12S mRNAs encode E1A T antigens of 266 and 235 amino acid residues (266R and 235R), respectively. To study possible posttranslational processing at the N and C termini and to distinguish between the Ad12 266R and 235R T antigens, we prepared antibodies targeted to synthetic peptides encoded at the common C (peptide 204) and N (peptide 202) termini of the 266R and 235R T antigens and at the unique internal domain of the 266R T antigen (peptide 206). The specificity of each anti-peptide antibody was confirmed by immunoprecipitation of the 266R and 235R T antigens produced in Escherichia coli. Immunoprecipitation analysis of the E1A T antigens synthesized in Ad12-infected KB cells revealed the following. Antibody to the common C terminus recognized three T antigens with apparent Mrs of 43,000, 42,000, and 39,000 (43K, 42K, and 39K). All three forms were phosphorylated and were present in both the nucleus and the cytoplasm. The 43K and 42K T antigens were rapidly synthesized during a 10-min pulse with [35S]methionine in Ad12-infected cells. The 43K T antigen had a half-life of 20 min, the 42K T antigen had a longer half-life of about 40 min, and the 39K T antigen became the predominant E1A T antigen. Antibodies to the unique region immunoprecipitated the 43K T antigen but not the 42K and 39K T antigens. Antibody to the N terminus immunoprecipitated the 43K and 42K T antigens but not the 39K T antigen, suggesting that the 39K T antigen possessed a modified N terminus. Partial N-terminal amino acid sequence analysis showed that the 43K and 42K T antigens contain methionine at residues 1 and 5, as predicted from the DNA sequence, whereas no methionine was released from the 39K T antigen during the first six cycles of Edman degradation. We propose that the short-lived 43K T antigen is the primary product of the 13S mRNA, the 266R T antigen; the somewhat more stable 42K T antigen is the primary product of the 12S mRNA, the 235R T antigen.(ABSTRACT TRUNCATED AT 400 WORDS)

Adenovirus Early Proteins↗

PubMed1997

Tissue transglutaminase-dependent posttranslational modification of the retinoblastoma gene product in promonocytic cells undergoing apoptosis.

The retinoblastoma gene product (pRB) plays an important role in controlling both cell release from the G1 phase and apoptosis. We show here that in the early phases of apoptosis, pRB is posttranslationally modified by a tissue transglutaminase (tTG)-catalyzed reaction. In fact, by employing a novel haptenized lysis synthetic substrate which allows the isolation of glutaminyl-tTG substrates in vivo, we identified pRB as a potential tTG substrate in U937 cells undergoing apoptosis. In keeping with this finding, we showed that apoptosis of U937 cells is characterized by the rapid disappearance of the 105,000- to 110,000-molecular-weight pRB forms concomitantly with the appearance of a smear of immunoreactive products with a molecular weight of greater than 250,000. The shift in pRB molecular weight was reproduced by adding exogenous purified tTG to extracts obtained from viable U937 cells and was prevented by dansylcadaverine, a potent enzyme inhibitor. The effect of the pRB posttranslational modification during apoptosis was investigated by determining the E2F-1 levels and by isolating and characterizing pRB-null clones from U937 cells. Notably, the lack of pRB in these U937-derived clones renders these p53-null cells highly resistant to apoptosis induced by serum withdrawal, calphostin C, and ceramide. Taken together, these data suggest that tTG, acting on the pRB protein, might play an important role in the cell progression through the death program.

Apoptosis↗

PubMed2000

Posttranslational modifications of p53 in replicative senescence overlapping but distinct from those induced by DNA damage.

Replicative senescence in human fibroblasts is absolutely dependent on the function of the phosphoprotein p53 and correlates with activation of p53-dependent transcription. However, no evidence for posttranslational modification of p53 in senescence has been presented, raising the possibility that changes in transcriptional activity result from upregulation of a coactivator. Using a series of antibodies with phosphorylation-sensitive epitopes, we now show that senescence is associated with major changes at putative regulatory sites in the N and C termini of p53 consistent with increased phosphorylation at serine-15, threonine-18, and serine-376 and decreased phosphorylation at serine-392. Ionizing and UV radiation generated overlapping but distinct profiles of response, with increased serine-15 phosphorylation being the only common change. These results support a direct role for p53 in signaling replicative senescence and are consistent with the generation by telomere erosion of a signal which shares some but not all of the features of DNA double-strand breaks.

Cell Line↗

PubMed1998

Posttranslational modifications of the C-terminus of alpha-tubulin in adult rat brain: alpha 4 is glutamylated at two residues.

In adult mammalian brain, the C-terminus of alpha-tubulin exhibits a high degree of polymorphism due to a combination of four covalent posttranslational modifications: glutamylation, tyrosination, detyrosination, and removal of the penultimate glutamate residue (C-terminal deglutamylation). Glutamylation is the most abundant. To characterize the glutamylation of alpha-tubulin and its relationship with the other modifications, we developed a chromatographic procedure for purifying alpha-tubulin C-terminal peptides. The purified peptides were identified by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF) and amino acid sequencing. In this report, we provide a complete description of the glutamylation of tyrosinated, detyrosinated, and C-terminal deglutamylated isoforms of both alpha-tubulin isotypes (alpha1/2 and alpha4) expressed in adult rat brain. In particular, we describe for the first time the glutamylation of alpha4. More than 90% of the alpha-tubulin is glutamylated, and more than 75% of it is nontyrosinated. alpha4 is more extensively glutamylated than alpha1/2, containing as many as 11 posttranslationally added glutamate residues. The most abundant alpha4 isoform is nontyrosinated, containing five posttranslationally added glutamates, whereas the most abundant alpha1/2 isoforms are nontyrosinated, with only one or two posttranslationally added glutamates. In contrast to alpha1/2, alpha4 is glutamylated at two separate residues (Glu-443 and Glu-445) in the sequence 431DYEEVGIDSYEDEDEGEE448. This is the first evidence that glutamylation can occur on two different residues in the same mammalian tubulin isotype.

Amino Acid Sequence↗

PubMed2002

Posttranslational modifications in the CP43 subunit of photosystem II.

Photosystem II (PSII) catalyzes the light-driven oxidation of water and the reduction of plastoquinone; the oxidation of water occurs at a cluster of four manganese. The PSII CP43 subunit functions in light harvesting, and mutations in the fifth luminal loop (E) of CP43 have established its importance in PSII structure and/or assembly [Kuhn, M. G. & Vermaas, V. F. J. (1993) Plant Mol. Biol. 23, 123-133]. The sequence A(350)PWLEPLR(357) in luminal loop E is conserved in CP43 genes from 50 organisms. To map important posttranslational modifications in this sequence, tandem mass spectrometry (MS/MS) was used. These data show that the indole side chain of Trp-352 is posttranslationally modified to give mass shifts of +4, +16, and +18 daltons. The masses of the modifications suggest that the tryptophan is modified to kynurenine (+4), a keto-/amino-/hydroxy- (+16) derivative, and a dihydro-hydroxy- (+18) derivative of the indole side chain. Peptide synthesis and MS/MS confirmed the kynurenine assignment. The +16 and +18 tryptophan modifications may be intermediates formed during the oxidative cleavage of the indole ring to give kynurenine. The site-directed mutations, W352C, W352L, and W352A, exhibit an increased rate of photoinhibition relative to wild type. We hypothesize that Trp-352 oxidative modifications are a byproduct of PSII water-splitting or electron transfer reactions and that these modifications target PSII for turnover. As a step toward understanding the tertiary structure of this CP43 peptide, structural modeling was performed by using molecular dynamics.

Algorithms↗

PubMed2003

Prediction of possible sites for posttranslational modifications in human gamma crystallins: effect of glycation on the structure of human gamma-B-crystallin as analyzed by molecular modeling.

Crystallins are recognized as one of the long-lived proteins of lens tissue that might serve as the target for several posttranslational modifications leading to cataract development. We have studied several such sites present in the human gamma-crystallins based either on PROSITE pattern search results or earlier experimental evidences. Their probabilities were examined on the basis of the database analysis of the gamma-crystallin sequences and on their specific locations in the constructed homology models. An N-glycosylation site in human gammaD-crystallin and several phosphorylation sites in all four human gamma-crystallins were predicted by the PROSITE search. Some of these sites were found to be strongly conserved in the gamma-crystallin sequences from different sources. An extensive analysis of these sites was performed to predict their probabilities as potential sites for protein modifications. Glycation studies were performed separately by attaching sugars to the human gammaB-crystallin model, and the effect of binding was analyzed. The studies showed that the major effect of alphaD-glucose (alphaD-G) and alphaD-glucose-6-phosphate (alphaD-G6P) binding was the disruption of charges not only at the surface but also within the molecule. Only a minor alteration in the distances of sulfhydryl groups of cysteines and on their positions in the three-dimensional models were observed, leading us to assume that glycation alone is not responsible for intra- and intermolecular disulfide bond formation.

Amino Acid Sequence↗

PubMed1998

Posttranslational modifications of endothelin receptor type B.

Endothelin receptor type B (ETb) is one of the two ET receptor subtypes, both of which belong to the G protein-coupled receptor (GPCR) superfamily. The primary amino acid sequence of ETb, published in 1990, predicted potential posttranslational modifications of the receptor protein, and in the last couple of years, we and others presented direct experimental evidence for palmitoylation and phosphorylation of ETb. Functional evaluation of both substitution and deletion mutants indicated a negative role on the part of these modifications in the ligand binding capacities and cellular sequestrations of the receptors. At least one of them, palmitoylation, however, appears to be critically involved in the coupling with G proteins.

Journal Article↗

PubMed2026

Microtubule posttranslational modifications provide unique recognition patterns for associated proteins.

Microtubules are key components of the eukaryotic cytoskeleton involved in vital functions in virtually every cell. Among the emerging molecular mechanisms to adapt microtubules to their diverse functions is the biochemical diversification of tubulin molecules by posttranslational modifications (PTMs) and differential gene expression, a concept known as the 'tubulin code'. A key question remains whether the tubulin code has the potential to selectively control microtubule interactions of different microtubule-associated proteins (MAPs) to act as a specific signalling system. To answer this question, we used a medium-throughput in vitro approach to screen 46 proteins for their binding preferences to microtubules with altered PTM or isotype composition. We demonstrate that subsets of these MAPs have unique sensitivities to PTMs, while other proteins are not affected. As a result, PTMs, or combinations of them, differentially attract or repulse individual MAPs to microtubules. Our findings offer mechanistic proof for a key hypothesis of the tubulin code-the capacity to selectively and differentially regulate MAP-microtubule interactions.

Journal Article↗

PubMed1993

Collagen II from articular cartilage and annulus fibrosus. Structural and functional implication of tissue specific posttranslational modifications of collagen molecules.

Collagen II was isolated and characterized from hyaline cartilage (articular cartilage) and fibro-cartilage (annulus fibrosus). Collagen II from the latter tissue has a substantially higher degree of hydroxylation and glycosylation than that isolated from articular cartilage. The higher degree of posttranslational modification was associated with a slower electrophoretic mobility, a greater resistance to mammalian collagenase digestion and a higher thermal stability. An increase of glycosylation accelerates the initial steps in fibril formation of collagen molecules but slows down the following lateral growth. The newly formed aggregates of collagen II from annulus fibrosus consisted of fibrils with a smaller diameter.

Amino Acid Sequence↗

PubMed1993

Antibiotics synthesized by posttranslational modification.

Peptides that have antimicrobial activity are synthesized by many prokaryotic and eukaryotic organisms. Antimicrobial peptides commonly contain unusual amino acids that contribute to their properties and functions. Although bacteria synthesize most of these peptides by nonribosomal mechanisms, this review focuses on those that are synthesized by pathways that involve posttranslational modification of ribosomally synthesized precursor peptides. A particularly interesting class of these antimicrobial peptides is the lantibiotics, of which nisin and subtilin are the longest-known examples, although nearly a dozen new lantibiotics have been discovered in recent years. The fact that the lantibiotic structures are derived from gene-encoded peptides means that structural analogs of natural lantibiotics can be constructed by mutagenesis of their structural genes. Recent advances in our understanding of the molecular genetics of lantibiotics has made the construction of novel lantibiotics with enhanced chemical and antimicrobial properties possible. This review describes these advances and proposes future trends of research, as well as potential application of engineered lantibiotics, in the context of the general field of antimicrobial peptides.

Amino Acid Sequence↗

PubMed1994

Posttranslational modification of glycine-extended substance P by an alpha-amidating enzyme in cultured sensory neurons of dorsal root ganglia.

The terminal step in the biosynthesis of substance P is the conversion of its glycine-extended precursor to the mature, amidated peptide by the alpha-amidating enzyme. This posttranslational modification was demonstrated in cultured, dissociated sensory neurons of dorsal root ganglia from neonatal rats. An assay was developed to quantitate both substance P and its precursor peptide in these cells. More than 90% of these two peptides was present as mature peptide in uncultured cells. In contrast, after 8 days in culture, about 85% of the peptides was the precursor, which increased 200-fold, whereas the level of substance P itself tripled during this culturing period. Since alpha-amidating enzyme requires ascorbate for activity, this reducing agent was added to the culture medium. Ascorbate induced a dose-dependent rise in the percentage of amidated peptide, with an apparent Km of 20 microM. After 5 days of culturing in the presence of 500 microM ascorbate, substance P increased 8-fold, constituting 70% of the total. The alpha-amidating enzyme also needs copper for activity. Even with 500 microM ascorbate in the culture medium, the copper chelator diethyldithiocarbamate dose-dependently reduced substance P synthesis by the sensory neurons, with a concomitant increase in its precursor peptide. These results suggest the presence of alpha-amidating enzyme in sensory neurons of dorsal root ganglia. It is likely that conversion of other glycine-extended precursors to their mature peptides in cell cultures would also require ascorbate and copper.

Animals↗

PubMed2002

The assembly of Ebola virus nucleocapsid requires virion-associated proteins 35 and 24 and posttranslational modification of nucleoprotein.

Ebola virus encodes seven viral structural and regulatory proteins that support its high rates of replication, but little is known about nucleocapsid assembly of this virus in infected cells. We report here that three viral proteins are necessary and sufficient for formation of Ebola virus particles and that intracellular posttranslational modification regulates this process. Expression of the nucleoprotein (NP) and virion-associated proteins VP35 and VP24 led to spontaneous assembly of nucleocapsids in transfected 293T cells by transmission electron microscopy. A specific biochemical interaction of these three proteins was demonstrated, and, interestingly, O-glycosylation and sialation of NP were demonstrated and necessary for their association. This distinct mechanism of regulation for filovirus assembly suggests new approaches for viral therapies and vaccines for Ebola and related viruses.

Amino Acid Sequence↗

PubMed1999

Observation of Escherichia coli ribosomal proteins and their posttranslational modifications by mass spectrometry.

Ribosomes from the K-12 strain of Escherichia coli were analyzed with good sensitivity and high mass accuracy using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Fifty-five of the 56 subunit proteins were observable. Mass spectral peak locations were consistent with previously reported post-translational modifications involving N-terminal methionine loss, methylation, thiomethylation, and acetylation for all but one case. The speed and accuracy of mass spectrometry make it a good candidate for phylogenetic studies of ribosomes and the observation of posttranslational modifications in other organisms.

Escherichia coli↗

PubMed2005

Signalling via the hypoxia-inducible factor-1alpha requires multiple posttranslational modifications.

Cellular hypoxia, a local decrease in the oxygen concentration below normal (21%) atmospheric concentrations, occurs in both physiological and pathological situations. The transcriptional complex Hypoxia-Inducible Factor-1 (HIF-1) is the key player in the signalling pathway that controls the hypoxic response of mammalian cells. Tight regulation of this response involves posttranslational modification of the alpha subunit of HIF-1. Hydroxylation, ubiquitination, acetylation, S-nitrosation and phosphorylation have been shown to determine its half-life and/or transcriptional activity. The precise spatio-temporal occurrence of these multiple modifications is still not fully understood but is dependent on the microenvironment and determines the driving force of variable cellular responses.

Animals↗

PubMed2005

Posttranslational modifications of decidual IGFBP-1 by steroid hormones in vitro.

Insulin-like growth factor binding protein-1 (IGFBP-1) appears to regulate insulin-like growth factors (IGFs; IGF-I and IGF-II) biological activity within the local environment of human placenta by modulating IGFs interaction with their receptors. Considering that posttranslational modifications of IGFBP-1 such as phosphorylation and proteolysis affect its affinity for IGFs, this study was undertaken to identify the role of estrogen and progesterone in this regard. The conditioned media of steroid hormone-treated decidual cells were evaluated using different approaches using sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and non-denaturing PAGE following immunoblotting as well as zymographys that contained gelatin and IGFBP-1 as substrates. Our results demonstrated that medroxy progesterone acetate (MPA) treatment increased both phosphorylated and non-phosphorylated decidual-secreted IGFBP-1, whereas 17beta-estradiol (E2) treatment attenuated its phosphorylated forms. Furthermore, the results of zymography revealed that steroid hormones regulated the activity of decidual-secreted matrix metalloproteinases (MMP)-2 and -9, in which E2 treatment up-regulated the MMP-9 activity. Finally, it was demonstrated in our study that decidual-secreted MMP-9 was capable of degrading human amniotic fluid-derived IGFBP-1. In conclusion, our data implicate steroid hormones in the control of IGF system activities at the embryo-maternal interface, at least in part, through their effects on the post-translation changes of decidual-secreted IGFBP-1 such as its phosphorylation and/or proteolysis.

Adult↗