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1.
Phosphorylation of proteins on serine and threonine residues has traditionally been viewed as a means to allosterically regulate catalytic activity. Research within the past five years, however, has revealed that serine/threonine phosphorylation can also directly result in the formation of multimolecular signaling complexes through specific interactions between phosphoserine/threonine (pSer/Thr)-binding modules and phosphorylated sequence motifs. pSer/Thr-binding proteins and domains currently include 14-3-3, WW domains, forkhead-associated domains, and, tentatively, WD40 repeats and leucine-rich regions. It seems likely that additional modules will be found in the future. The amino acid sequences recognized by these pSer/Thr-binding modules show partial overlap with the optimal phosphorylation motifs for different protein kinase subfamilies, allowing the formation of specific signaling complexes to be controlled through combinatorial interactions between particular upstream kinases and a particular binding module. The structural basis for pSer/Thr binding differs dramatically between 14-3-3 proteins, WW domains and forkhead-associated domains, suggesting that their pSer/Thr binding function was acquired through convergent evolution. 相似文献
2.
Phosphopeptide binding domains mediate the directed and localized assembly of protein complexes essential to intracellular kinase signaling. To identify phosphopeptide binding proteins, we developed a proteomic screening method using immobilized partially degenerate phosphopeptide mixtures combined with SILAC and microcapillary LC-MS/MS. The method was used to identify proteins that specifically bound to phosphorylated peptide library affinity matrices, including pTyr, and the motifs pSer/pThr-Pro, pSer/pThr-X-X-X-pSer/pThr, pSer/pThr-Glu/Asp, or pSer/pThr-pSer/pThr in degenerate sequence contexts. Heavy and light SILAC lysates were applied to columns containing these phosphorylated and nonphosphorylated (control) peptide libraries respectively, and bound proteins were eluted, combined, digested, and analyzed by LC-MS/MS using a hybrid quadrupole-TOF mass spectrometer. Heavy/light peptide ion ratios were calculated, and peptides that yielded ratios greater than ~3:1 were considered as being from potential phosphopeptide binding proteins since this ratio represents the lowest ratio from a known positive control. Many of those identified were known phosphopeptide-binding proteins, including the SH2 domain containing p85 subunit of PI3K bound to pTyr, 14-3-3 bound to pSer/pThr-Asp/Glu, polo-box domain containing PLK1 and Pin1 bound to pSer/pThr-Pro, and pyruvate kinase M2 binding to pTyr. Approximately half of the hits identified by the peptide library screens were novel. Protein domain enrichment analysis revealed that most pTyr hits contain SH2 domains, as expected, and to a lesser extent SH3, C1, STAT, Tyr phosphatase, Pkinase, C2, and PH domains; however, pSer/pThr motifs did not reveal enriched domains across hits. 相似文献
3.
Phosphotyrosine-binding (PTB) domains were originally identified as modular domains that recognize phosphorylated Asn-Pro-Xxx-p Tyr-containing proteins. Recent binding and structural studies of PTB domain complexes with target peptides have revealed a number of deviations from the previously described mode of interaction, with respect to both the sequences of possible targets and their structures within the complexes. This diversity of recognition by PTB domains extends and strengthens our general understanding of modular binding domain recognition. 相似文献
4.
The forkhead-associated (FHA) domain is a 55-75 amino acid residue module found in >20 proteins from yeast to human. It has been suggested to participate in signal transduction pathways, perhaps via protein-protein interactions involving recognition of phosphopeptides. Neither the structure nor the ligand of FHA is known. Yeast Rad53, a checkpoint protein involved in DNA damage response, contains two FHA domains, FHA1 (residues 66-116) and FHA2 (residues 601-664), the second of which recognizes phosphorylated Rad9. We herein report the solution structure of an "FHA2-containing domain" of Rad53 (residues 573-730). The structure consists of a beta-sandwich containing two antiparallel beta-sheets and a short, C-terminal alpha-helix. Binding experiments suggested that the FHA2-containing domain specifically recognizes pTyr and a pTyr-containing peptide from Rad9, and that the binding site involves residues highly conserved across FHA domains. The results, along with other recent reports, suggest that FHA domains could have pTyr and pSer/Thr dual specificity. 相似文献
5.
Members of the LanL family of lanthionine synthetases consist of three catalytic domains, an N-terminal pSer/pThr lyase domain, a central Ser/Thr kinase domain, and a C-terminal lanthionine cyclase domain. The N-terminal lyase domain has sequence homology with members of the OspF family of effector proteins. In this study, the residues in the lyase domain of VenL that are conserved in the active site of OspF proteins were mutated to evaluate their importance for catalysis. In addition, residues that are fully conserved in the LanL family but not in the OspF family were mutated. Activity assays with these mutant proteins are consistent with a model in which Lys80 in VenL deprotonates the α-proton of pSer/pThr residues to initiate the elimination reaction. Lys51 is proposed to activate this proton by coordination to the carbonyl of the pSer/pThr, and His53 is believed to protonate the phosphate leaving group. These functions are very similar to the corresponding homologous residues in OspF proteins. On the other hand, recognition of the phosphate group of pSer/pThr appears to be achieved differently in VenL than in the OspF proteins. Arg156 and Lys103 are thought to interact with the phosphate group on the basis of a structural homology model. 相似文献
6.
Thy-1, a glycosylphosphatidylinositol (GPI)-anchored glycoprotein expressed at high levels on thymocytes, has been implicated in positive and negative signal transduction. We show that Thy-1 associates with a protein of 85--90 kDa, which is prominently phosphorylated in vitro as well as in vivo following the stimulation of thymocytes with pervanadate. pp85--90 is not identical to known proteins that are phosphorylated following T cell activation. The SH2 domains of fyn, csk, phosphatidylinositol 3'-kinase, rasGAP, vav and lck bind to pp85--90 with varying affinities. The SH2 domains of ZAP70, SHP-1 and PLC gamma 1 and the SH3 domains of lck, vav and HS1 did not bind to pp85--90. The molecular weight, iso-electric point, efficient phosphorylation by fyn and lck and preferential binding to the SH2 domain of fyn compared to that of lck indicate that Thy-1-associated pp85-90 may be identical to a recently cloned, fyn-associated transmembrane adaptor protein, PAG-85. 相似文献
7.
8.
Continuous and discontinuous domains: an algorithm for the automatic generation of reliable protein domain definitions. 总被引:9,自引:5,他引:4
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A. S. Siddiqui G. J. Barton 《Protein science : a publication of the Protein Society》1995,4(5):872-884
An algorithm is presented for the fast and accurate definition of protein structural domains from coordinate data without prior knowledge of the number or type of domains. The algorithm explicitly locates domains that comprise one or two continuous segments of protein chain. Domains that include more than two segments are also located. The algorithm was applied to a nonredundant database of 230 protein structures and the results compared to domain definitions obtained from the literature, or by inspection of the coordinates on molecular graphics. For 70% of the proteins, the derived domains agree with the reference definitions, 18% show minor differences and only 12% (28 proteins) show very different definitions. Three screens were applied to identify the derived domains least likely to agree with the subjective definition set. These screens revealed a set of 173 proteins, 97% of which agree well with the subjective definitions. The algorithm represents a practical domain identification tool that can be run routinely on the entire structural database. Adjustment of parameters also allows smaller compact units to be identified in proteins. 相似文献
9.
Won Byoung Chae Youn-Je Park Kyung Sun Lee Ill-Sup Nou Man-Ho Oh 《Genes & genomics.》2016,38(11):1111-1119
14-3-3 proteins are pSer/pThr-binding proteins that interact with a wide array of cellular ‘client’ proteins. The plant brassinosteroids (BRs) receptor, BRASSINOSTEROID INSENSITIVE 1 (BRI1), is a member of the large family of leucine-rich repeat receptor-like kinases (LRR-RLKs) that contain cytoplasmic protein kinase domains. At least two LRR-RLKs are involved in BR perception and signal transduction: BRI1 and BRI1-associated receptor kinase 1 (BAK1). We determined that several 14-3-3 proteins bind to BRI1-CD and are phosphorylated by BRI1, BAK1 and At3g21430 receptor kinases in vitro. Moreover, we observed14-3-3 s are phosphorylated on threonine residue(s) with BR-dependent manner. To reveal the function of 14-3-3 proteins interacting with LRR-RLKs, we treated tyrosine phosphatase (PTP1B) to the BRI1-CD recombinant protein, which is autophosphorylated on tyrosine residue(s). Tyrosine autophosphorylation signal was disappeared, suggesting that 14-3-3 proteins cannot protect BRI1 tyrosine phosphorylation from PTP1B phosphatase. Our study suggests that 14-3-3 proteins may be important for plant growth and development through BR signaling. 相似文献
10.
Two phosphoserine tetradecapeptides corresponding to sequences 987-1000 (peptide pSer994) and 1017-1030 (peptide pSer1023/1025) from the human insulin receptor involved in the regulation of its activity were successfully synthesized using Fmoc-based chemistry. Phosphorylation was performed by post-assembly phosphitylation followed by oxidation. The selective phosphorylation of Ser residues was achieved incorporating into the peptide chain the Ser (Trt) derivative and t-Bu blocking groups at sites other than those intended to be phosphorylated. The Trt group was selectively removed with dichloroacetic acid while under this condition t-Bu protecting groups remained unaltered. Following conjugation to keyhole limpet hemocyanin phosphopeptides were used as immunogens to generate sequence-specific phosphoserine antibodies. Peptide pSer994 induced antibodies in New Zealand white rabbits which discriminated between the phosphorylated and nonphosphorylated forms of the peptide, thus representing promising candidates to recognize signaling pathways associated to the regulation of the human insulin receptor. 相似文献
11.
The N-terminal RNA-binding domain (RBD1) of the human U1A protein is evolutionarily designed to bind its RNA targets with great affinity and specificity. The physical mechanisms that modulate the coupling (local cooperativity) among amino acid residues on the extensive binding surface of RBD1 are investigated here, using mutants that replace a highly conserved glycine residue. This glycine residue, at the strand/loop junction of beta3/loop3, is found in U1A RBD1, and in most RBD domains, suggesting it has a specific role in modulation of RNA binding. Here, two RBD1 proteins are constructed in which that residue (Gly53) is replaced by either alanine or valine. These new proteins are shown by NMR methods and molecular dynamics simulations to be very similar to the wild-type RBD1, both in structure and in their backbone dynamics. However, RNA-binding assays show that affinity for the U1 snRNA stem-loop II RNA target is reduced by nearly 200-fold for the RBD1-G53A protein, and by 1.6 x 10(4)-fold for RBD1-G53V. The mode of RNA binding by RBD1-G53A is similar to that of RBD1-WT, displaying its characteristic non-additive free energies of base recognition and its salt-dependence. The binding mode of RBD1-G53V is altered, having lost its salt-dependence and displaying site-independence of base recognition. The molecular basis for this alteration in RNA-binding properties is proposed to result from the inability of the RNA to induce a change in the structure of the free protein to produce a high-affinity complex. 相似文献
12.
真核生物中锌指蛋白的结构与功能 总被引:3,自引:0,他引:3
真核生物中的许多蛋白质分子包含锌指结构区,这类蛋白称为锌指蛋白.锌指蛋白因其包含特殊的指状结构,在对DNA、蛋白质和RNA的识别和结合中起重要作用.许多锌指蛋白的锌指结构域包含能与DNA特异结合的区域,并与某些效应结构域(如KRAB、SCAN、BTB/POZ、SNAG、SANT和PLAG等)相连,这类锌指蛋白常作为转录因子起作用,可调控靶基因的转录.一些锌指蛋白包含蛋白质识别结构域(如LIM锌指、MYND锌指、PHD锌指和RING锌指等),它们能够特异地介导蛋白质之间的相互作用,因此被称作蛋白适配器.此外,某些锌指蛋白还可以结合RNA,起转录后调控作用.本文就锌指蛋白与DNA、RNA以及蛋白质分子间的相互作用作一综述. 相似文献
13.
Membrane-associated guanylate kinases (MAGUKs) are a large family of scaffold proteins that play essential roles in tissue developments, cell-cell communications, cell polarity control, and cellular signal transductions. Despite extensive studies over the past two decades, the functions of the signature guanylate kinase domain (GK) of MAGUKs are poorly understood. Here we show that the GK domain of DLG1/SAP97 binds to asymmetric cell division regulatory protein LGN in a phosphorylation-dependent manner. The structure of the DLG1 SH3-GK tandem in complex with a phospho-LGN peptide reveals that the GMP-binding site of GK has evolved into a specific pSer/pThr-binding pocket. Residues both N- and C-terminal to the pSer are also critical for the specific binding of the phospho-LGN peptide to GK. We further demonstrate that the previously reported GK domain-mediated interactions of DLGs with other targets, such as GKAP/DLGAP1/SAPAP1 and SPAR, are also phosphorylation dependent. Finally, we provide evidence that other MAGUK GKs also function as phospho-peptide-binding modules. The discovery of the phosphorylation-dependent MAGUK GK/target interactions indicates that MAGUK scaffold-mediated signalling complex organizations are dynamically regulated. 相似文献
14.
The core histone tail domains are "master control switches" that help define the structural and functional characteristics of chromatin at many levels. The tails modulate DNA accessibility within the nucleosome, are essential for stable folding of oligonucleosome arrays into condensed chromatin fibers, and are important for fiber-fiber interactions involved in higher order structures. Many nuclear signaling pathways impinge upon the tail domains, resulting in posttranslational modifications that are likely to alter the charge, structure, and/or interactions of the core histone tails or to serve as targets for the binding of ancillary proteins or other enzymatic functions. However, currently we have only a marginal understanding of the molecular details of core histone tail conformations and contacts. Here we review data related to the structures and interactions of the core histone tail domains and how these domains and posttranslational modifications therein may define the structure and function of chromatin. 相似文献
15.
Yan-Jing Kang Li-Min Zuo Shi-Zhong Luo 《International journal of peptide research and therapeutics》2010,16(2):87-93
Reversible protein phosphorylation is the most common and important regulatory modification of proteins. Litter is known about
exactly how protein phosphorylation affects protein local conformation. This study explores the effect of phosphorylation
on the local secondary structure of the peptide, which would have implications for understanding the structural properties
of kinase targets since these consist of the amino acid sequences immediately adjacent to the phosphorylated site as well
as the phosphorylated amino acid itself. To this end ten pentapeptides (five phosphorylated), which represent variations of
the consensus sequence for the cyclic-AMP dependent protein kinase, were synthesized and studied using NMR spectroscopy. The
NMR experiments with downfield shifts of these protons upon peptide phosphorylation as well as relatively low shifts upon
a temperature change, are consistent with the existence of transient hydrogen bonds between the phosphate group on pSer4 and
both the Arg1 εH and the pSer4 NH in a peptide of the sequence Ac-RKGpSS-NH2. Furthermore, pH titrations, which would be expected to deprotonate the phosphate, result in the expected upfield shift of
pSer4 side chain resonances and also show a striking downfield shift of pSer4 NH compared with other NHs and also a downfield
shift of Arg1 εH. Similar experimental observations were identified on Ac-RKGpTS-NH2 peptide, but not on Ac-RKGpYS-NH2 peptide. Conformational searches using a MCMM conformational modeling program resulted in peptide conformations within which
these hydrogen bonds were formed. These findings point to a possible structural explanation for the effect of preceding Arg
residues on the Ser and Thr phosphorylation specificity of protein kinases. 相似文献
16.
Proteins destined for secretion or membrane compartments possess signal peptides for insertion into the membrane. The signal peptide is therefore critical for localization and function of cell surface receptors and ligands that mediate cell-cell communication. About 4% of all human proteins listed in UniProt database have signal peptide domains in their N terminals. A comprehensive literature survey was performed to retrieve functional and disease associated genetic variants in the signal peptide domains of human proteins. In 21 human proteins we have identified 26 disease associated mutations within their signal peptide domains, 14 mutations of which have been experimentally shown to impair the signal peptide function and thus influence protein transportation. We took advantage of SignalP 3.0 predictions to characterize the signal peptide prediction score differences between the mutant and the wild-type alleles of each mutation, as well as 189 previously uncharacterized single nucleotide polymorphisms (SNPs) found to be located in the signal peptide domains of 165 human proteins. Comparisons of signal peptide prediction outcomes of mutations and SNPs, have implicated SNPs potentially impacting the signal peptide function, and thus the cellular localization of the human proteins. The majority of the top candidate proteins represented membrane and secreted proteins that are associated with molecular transport, cell signaling and cell to cell interaction processes of the cell. This is the first study that systematically characterizes genetic variation occurring in the signal peptides of all human proteins. This study represents a useful strategy for prioritization of SNPs occurring within the signal peptide domains of human proteins. Functional evaluation of candidates identified herein may reveal effects on major cellular processes including immune cell function, cell recognition and adhesion, and signal transduction. 相似文献
17.
Wenjian Qian Jung-Eun Park Fa Liu Kyung S. Lee Terrence R. Burke 《Bioorganic & medicinal chemistry》2013,21(14):3996-4003
Protein–protein interactions (PPIs) mediated by the polo-box domain (PBD) of polo-like kinase 1 (Plk1) serve important roles in cell proliferation. Critical elements in the high affinity recognition of peptides and proteins by PBD are derived from pThr/pSer-residues in the binding ligands. However, there has been little examination of pThr/pSer mimetics within a PBD context. Our current paper compares the abilities of a variety of amino acid residues and derivatives to serve as pThr/pSer replacements by exploring the role of methyl functionality at the pThr β-position and by replacing the phosphoryl group by phosphonic acid, sulfonic acid and carboxylic acids. This work sheds new light on structure activity relationships for PBD recognition of phosphoamino acid mimetics. 相似文献
18.
J F Allen 《Trends in biochemical sciences》1992,17(1):12-17
Phosphorylation of light-harvesting antenna proteins redirects absorbed light energy between reaction centres of photosynthetic membranes. A generally accepted explanation for this is that electrostatic forces drive the more negatively charged, phosphorylated antenna proteins between membrane domains that differ in surface charge. However, structural studies on soluble phosphoproteins indicate that phosphorylated amino acid side chains have specific effects on molecular recognition, by ligand blocking or by intramolecular interactions which alter protein structure. These studies suggest alternative mechanisms for phosphorylation in control of pairwise protein-protein interactions in biological membranes. Thus, in photosynthesis, the surface charge model is only one possible interpretation. 相似文献
19.
Pinning down phosphorylated tau and tauopathies 总被引:4,自引:0,他引:4
Neurofibrillary tangles (NFTs) are prominent neuronal lesions in a large subset of neurodegenerative diseases, including Alzheimer's disease (AD). NFTs are mainly composed of insoluble Tau that is hyperphosphorylated on many serine or threonine residues preceding proline (pSer/Thr-Pro). Tau hyperphosphorylation abolishes its biological function to bind microtubules and promotes microtubule assembly and precedes neurodegeneration. Not much is known about how tau is further regulated following phosphorylation. Notably, we have recently shown that phosphorylated Ser/Thr-Pro motifs exist in two distinct conformations. The conversion between two conformations in some proteins is catalyzed by the prolyl isomerase Pin1. Pin1 binds to tau phosphorylated specifically on the Thr231-Pro site and probably catalyzes cis/trans isomerization of pSer/Thr-Pro motif(s), thereby inducing conformational changes in tau. Such conformational changes can directly restore the ability of phosphorylated Tau to bind microtubules and promote microtubule assembly and/or facilitate tau dephosphorylation by its phosphatase PP2A, as PP2A activity is conformation-specific. Furthermore, Pin1 expression inversely correlates with the predicted neuronal vulnerability in normally aged brain and also with actual neurofibrillary degeneration in AD brain. Moreover, deletion of the gene encoding Pin1 in mice causes progressive age-dependent neuropathy characterized by motor and behavioral deficits, tau hyperphosphorylation, tau filament formation and neuronal degeneration. Distinct from all other mouse models where transgenic overexpression of specific proteins elicits tau-related pathologies, Pin1 is the first protein whose depletion causes age-dependent neurodegeneration and tau pathologies. Thus, Pin1 is pivotal in maintaining normal neuronal function and preventing age-dependent neurodegeneration. This could represent a promising interventive target to prevent neurodegenerative diseases. 相似文献
20.
Lysine acetylation and the bromodomain: a new partnership for signaling 总被引:16,自引:0,他引:16
Yang XJ 《BioEssays : news and reviews in molecular, cellular and developmental biology》2004,26(10):1076-1087