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The Journal of Biological Chemistry May 1999Human C4b-binding protein (C4BP) is a regulator of the complement system and plays an important role in the regulation of the anticoagulant protein C pathway. C4BP can...
Human C4b-binding protein (C4BP) is a regulator of the complement system and plays an important role in the regulation of the anticoagulant protein C pathway. C4BP can bind anticoagulant protein S, resulting in a decreased cofactor function of protein S for activated protein C. C4BP is a multimeric protein containing several identical alpha-chains and a single beta-chain (C4BPbeta), each chain being composed of short consensus repeats (SCRs). Previous studies have localized the protein S binding site to the NH2-terminal SCR (SCR-1) of C4BPbeta. To further localize the protein S binding site, we constructed chimeras containing C4BPbeta SCR-1, SCR-2, SCR-3, SCR-1+2, SCR-1+3, and SCR-2+3 fused to tissue-type plasminogen activator. Binding assays of protein S with these chimeras indicated that SCR-2 contributes to the interaction of protein S with SCR-1, since the affinity of protein S for SCR-1+2 was up to 5-fold higher compared with SCR-1 and SCR-1+3. Using an assay that measures protein S cofactor activity, we showed that cofactor activity was decreased due to binding to constructs that contain SCR-1. SCR-1+2 inhibited more potently than SCR-1 and SCR-1+3. SCR-3 had no additional effect on SCR-1, and therefore the effect of SCR-2 was specific. In conclusion, beta-chain SCR-2 contributes to the interaction of C4BP with protein S.
Topics: Complement C4b; Complement Inactivator Proteins; Consensus Sequence; Glycoproteins; Humans; Microsatellite Repeats; Protein S; Receptors, Complement; Recombinant Fusion Proteins
PubMed: 10329721
DOI: 10.1074/jbc.274.21.15144 -
Developmental Cell Aug 2001Most current models of spindle assembly checkpoint signaling involve inhibition of the Cdc20-APC by Mad2 protein. Interestingly, a paper from Hongtao Yu and colleagues... (Review)
Review
Most current models of spindle assembly checkpoint signaling involve inhibition of the Cdc20-APC by Mad2 protein. Interestingly, a paper from Hongtao Yu and colleagues in this issue of Developmental Cell suggests that the Cdc20/APC can also be inhibited in a Mad2-independent manner by a complex of proteins that includes BubR1.
Topics: Animals; Calcium-Binding Proteins; Carrier Proteins; Cdc20 Proteins; Cell Cycle; Cell Cycle Proteins; Fungal Proteins; Humans; Kinetochores; Mad2 Proteins; Models, Biological; Nuclear Proteins; Protein Binding; Protein Kinases; Protein Serine-Threonine Kinases; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Signal Transduction
PubMed: 11702776
DOI: 10.1016/s1534-5807(01)00032-6 -
Prion May 2017Yeast prions are protein-based genetic elements that propagate through cell populations via cytosolic transfer from mother to daughter cell. Molecular chaperone proteins...
Yeast prions are protein-based genetic elements that propagate through cell populations via cytosolic transfer from mother to daughter cell. Molecular chaperone proteins including Hsp70, the Hsp40/J-protein Sis1, and Hsp104 are required for continued prion propagation, however the specific requirements of chaperone proteins differ for various prions. We recently reported that Swa2, the yeast homolog of the mammalian protein auxilin, is specifically required for the propagation of the prion [URE3]. [URE3] propagation requires both a functional J-domain and the tetratricopeptide repeat (TPR) domain of Swa2, but does not require Swa2 clathrin binding. We concluded that the TPR domain determines the specificity of the genetic interaction between Swa2 and [URE3], and that this domain likely interacts with one or more proteins with a C-terminal EEVD motif. Here we extend that analysis to incorporate additional data that supports this hypothesis. We also present new data eliminating Hsp104 as the relevant Swa2 binding partner and discuss our findings in the context of other recent work involving Hsp90. Based on these findings, we propose a new model for Swa2's involvement in [URE3] propagation in which Swa2 and Hsp90 mediate the formation of a multi-protein complex that increases the number of sites available for Hsp104 disaggregation.
Topics: Auxilins; Glutathione Peroxidase; HSP40 Heat-Shock Proteins; HSP90 Heat-Shock Proteins; Heat-Shock Proteins; Phosphoproteins; Prions; Protein Binding; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Vesicular Transport Proteins
PubMed: 28574745
DOI: 10.1080/19336896.2017.1331810 -
FEBS Letters Aug 1998Activated protein C (APC) regulates blood coagulation by degrading factor Va (FVa) and factor VIIIa (FVIIIa). Protein S is a cofactor to APC in the FVa degradation,...
Activated protein C (APC) regulates blood coagulation by degrading factor Va (FVa) and factor VIIIa (FVIIIa). Protein S is a cofactor to APC in the FVa degradation, whereas FVIIIa degradation is potentiated by the synergistic APC-cofactor activity of protein S and factor V (FV). To elucidate the importance of the sex-hormone-binding globulin (SHBG)-like region in protein S for expression of anticoagulant activity, a recombinant protein S/Gas6 chimera was constructed. It comprised the amino-terminal half of protein S and the SHBG-like region of Gas6, a structurally similar protein having no known anticoagulant properties. The protein S/Gas6 chimera expressed 40-50%, APC-cofactor activity in plasma as compared to wild-type protein S. In the degradation of FVa by APC, the protein S/Gas6 chimera was only slightly less efficient than wild-type protein S. In contrast, the protein S/Gas6 chimera expressed no FV-dependent APC-cofactor activity in a FVIIIa-degradation system. This demonstrates the SHBG-like region to be important for expression of APC-cofactor activity of protein S and suggests that the SHBG-like region of protein S interacts with FV during the APC-mediated inactivation of FVIIIa.
Topics: Factor V; Factor VIIIa; Factor Va; Humans; Intercellular Signaling Peptides and Proteins; Partial Thromboplastin Time; Protein C; Protein S; Proteins; Recombinant Fusion Proteins; Sex Hormone-Binding Globulin; Structure-Activity Relationship
PubMed: 9738926
DOI: 10.1016/s0014-5793(98)00877-1 -
Genes To Cells : Devoted To Molecular &... Jan 2000The correct transmission of chromosomes from mother to daughter cells is fundamental for genetic inheritance. Separation and segregation of sister chromatids in growing... (Review)
Review
The correct transmission of chromosomes from mother to daughter cells is fundamental for genetic inheritance. Separation and segregation of sister chromatids in growing cells occurs in the cell cycle stage called 'anaphase'. The basic process of sister chromatid separation is similar in all eukaryotes: many gene products required are conserved. In this review, the roles of two proteins essential for the onset of anaphase in fission yeast, Cut2/securin and Cut1/separin, are discussed with regard to cell cycle regulation, and compared with the postulated roles of homologous proteins in other organisms. Securin, like mitotic cyclins, is the target of the anaphase promoting complex (APC)/cyclosome and is polyubiquitinated before destruction in a manner dependent upon the destruction sequence. The anaphase never occurs properly in the absence of securin destruction. In human cells, securin is an oncogene. Separin is a large protein (MW approximately 180 kDa), the C-terminus of which is conserved, and is thought to be inhibited by association with securin at the nonconserved N-terminus. In the budding yeast, Esp1/separin is thought to be a component of proteolysis against Scc1, an essential subunit of cohesin which is thought to link duplicated sister chromatids up to the anaphase. Whether fission yeast Cut1/separin is also implicated in proteolysis of cohesin is discussed.
Topics: Anaphase; Animals; Cell Cycle; Cell Cycle Proteins; Chromatids; Chromosomal Proteins, Non-Histone; Endopeptidases; Fungal Proteins; Humans; Nuclear Proteins; Phosphoproteins; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Schizosaccharomyces; Schizosaccharomyces pombe Proteins; Securin; Separase; Signal Transduction
PubMed: 10651900
DOI: 10.1046/j.1365-2443.2000.00306.x -
Genes & Development Mar 2004
Review
Topics: Amyloid; Animals; Fungal Proteins; Glutathione Peroxidase; Humans; Peptide Termination Factors; Prion Diseases; Prions; Saccharomyces cerevisiae Proteins
PubMed: 15037545
DOI: 10.1101/gad.1177104 -
Proceedings of the National Academy of... Jun 1983C4b-binding protein (C4bp) participates in the regulation of the C3 convertase of the classical pathway of complement. By binding to C4b, which is one of the structural...
C4b-binding protein (C4bp) participates in the regulation of the C3 convertase of the classical pathway of complement. By binding to C4b, which is one of the structural subunits of this enzyme, C4bp accelerates the decay-dissociation of the enzyme and renders C4b susceptible to degradation by factor I (C3b inactivator). C4bp is a high molecular weight plasma protein (Mr = 570,000) composed of apparently identical subunits (Mr = 70,000) linked by disulfide bonds. In plasma and in purified form C4bp also forms a bimolecular complex (Kd = 0.9 X 10(-7) M) with protein S, a recently identified vitamin K-dependent plasma protein. The binding sites on C4bp for protein S and C4b are distinct and noncompetitive and protein S does not influence the function of C4bp as a regulator of the C3 convertase. C4bp, C4b, and protein S were visualized by electron microscopy by negative staining. C4bp was found to have an unusual spider-like structure. It is composed of seven thin (30 A), elongated (330 A), and flexible subunits that are linked to a small central body. Protein S exhibited two globular domains of equal size with a center-to-center distance of approximately equal to 50 A. Protein S was found to bind to the C4bp through only one of its domains by attaching to a short subunit that is distinct from the other seven subunits. C4b imaged as an irregular, relatively compact molecule. It was found to interact with the peripheral ends of the elongated subunits, suggesting seven C4b-binding sites per molecule of C4bp.
Topics: Carrier Proteins; Complement C4; Complement C4b; Complement Inactivator Proteins; Glycoproteins; Humans; Macromolecular Substances; Microscopy, Electron; Molecular Weight; Protein S
PubMed: 6222381
DOI: 10.1073/pnas.80.11.3461 -
The Biochemical Journal Mar 1983C4b-binding protein was purified from human plasma in high yield by a simple procedure involving barium citrate adsorption and two subsequent chromatographic steps....
C4b-binding protein was purified from human plasma in high yield by a simple procedure involving barium citrate adsorption and two subsequent chromatographic steps. Approx. 80% of plasma C4b-binding protein was adsorbed on the barium citrate, presumably because of its complex-formation with vitamin K-dependent protein S. The purified C4b-binding protein had a molecular weight of 570 000, as determined by ultracentrifugation, and was composed of about eight subunits (Mr approx. 70 000). Uncomplexed plasma C4b-binding protein was purified from the supernatant after barium citrate adsorption. On sodium dodecyl sulphate/polyacrylamide-gel electrophoresis in non-reducing conditions and on agarose-gel electrophoresis it appeared as a doublet, indicating two forms differing slightly from each other in molecular weight and net charge. The protein band with the higher molecular weight in the doublet corresponded to the C4b-binding protein purified from the barium citrate eluate. Complex-formation between protein S and C4b-binding protein was studied in plasma, and in a system with purified components, by an agarose-gel electrophoresis technique. Protein S was found to form a 1:1 complex with the higher-molecular-weight form of C4b-binding protein, whereas the lower-molecular-weight form of C4b-binding protein did not bind protein S. The KD for the C4b-binding protein-protein S interaction in a system with purified components was approx. 0.9 X 10(-7) M. Rates of association and dissociation at 37 degrees C were low, namely about 1 X 10(3) M-1 . S-1 and 1.8 X 10(-4)-4.5 X 10(-4) S-1 respectively. In human plasma free protein S and free higher-molecular-weight C4b-binding protein were in equilibrium with the C4b-binding protein-protein S complex. Approx. 40% of both proteins existed as free proteins. From equilibrium data in plasma a KD of about 0.7 X 10(-7) M was calculated for the C4b-binding protein-protein S interaction.
Topics: Amino Acids; Carrier Proteins; Complement Inactivator Proteins; Electrophoresis, Agar Gel; Glycoproteins; Humans; Kinetics; Macromolecular Substances; Molecular Weight; Protein Binding; Protein S; Temperature
PubMed: 6223625
DOI: 10.1042/bj2090847 -
Blood Oct 1986Protein S, is a natural anticoagulant protein which serves as a cofactor for activated protein C. During pregnancy and in the postpartum period, functional protein S...
Protein S, is a natural anticoagulant protein which serves as a cofactor for activated protein C. During pregnancy and in the postpartum period, functional protein S levels are significantly reduced (38% +/- 17.3%, mean +/- 1 SD) when compared to nonpregnant females (97% +/- 31.6%) (P less than 0.001). In plasma an equilibrium exists between functionally active free protein S and protein S complexed with C4b-binding protein, which is functionally inactive. As a result of this equilibrium either a decreased level of total protein S antigen or an elevation of C4b-binding protein could lead to reduced protein S activity. C4b-binding protein levels measured by enzyme-linked immunoassay are not significantly different in pregnant women versus nonpregnant controls (103.5% +/- 21.2% v 100% +/- 16.9%). However, during pregnancy and in the postpartum period, total protein S levels are reduced (68% +/- 10.7%) compared to nonpregnant controls (100% +/- 17.0%). This difference is significant at P less than 0.001. These data demonstrated that the reduction in protein S activity observed during pregnancy is a result of reduced total protein S antigen.
Topics: Carrier Proteins; Complement C4; Complement Inactivator Proteins; Female; Glycoproteins; Humans; Pregnancy; Protein S; Radioimmunoassay
PubMed: 2944555
DOI: No ID Found -
PLoS Genetics Oct 2014Hsp100 family chaperones of microorganisms and plants cooperate with the Hsp70/Hsp40/NEF system to resolubilize and reactivate stress-denatured proteins. In yeast this...
Hsp100 family chaperones of microorganisms and plants cooperate with the Hsp70/Hsp40/NEF system to resolubilize and reactivate stress-denatured proteins. In yeast this machinery also promotes propagation of prions by fragmenting prion polymers. We previously showed the bacterial Hsp100 machinery cooperates with the yeast Hsp40 Ydj1 to support yeast thermotolerance and with the yeast Hsp40 Sis1 to propagate [PSI+] prions. Here we find these Hsp40s similarly directed specific activities of the yeast Hsp104-based machinery. By assessing the ability of Ydj1-Sis1 hybrid proteins to complement Ydj1 and Sis1 functions we show their C-terminal substrate-binding domains determined distinctions in these and other cellular functions of Ydj1 and Sis1. We find propagation of [URE3] prions was acutely sensitive to alterations in Sis1 activity, while that of [PIN+] prions was less sensitive than [URE3], but more sensitive than [PSI+]. These findings support the ideas that overexpressing Ydj1 cures [URE3] by competing with Sis1 for interaction with the Hsp104-based disaggregation machine, and that different prions rely differently on activity of this machinery, which can explain the various ways they respond to alterations in chaperone function.
Topics: Binding Sites; Endopeptidase Clp; Escherichia coli Proteins; Glutathione Peroxidase; HSP40 Heat-Shock Proteins; HSP70 Heat-Shock Proteins; HSP90 Heat-Shock Proteins; Heat-Shock Proteins; Molecular Chaperones; Mutation; Peptide Termination Factors; Prions; Protein Structure, Tertiary; Saccharomyces cerevisiae Proteins
PubMed: 25329162
DOI: 10.1371/journal.pgen.1004720