Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.21.4 (trypsin)
 42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The complement regulatory enzyme, C3b inactivator (C3bINA), has been purified from human serum by affinity chromatography on an anti-C3bINA Sepharose column. Subsequent chromatography on DEAE-cellulose and removal of IgG with anti-IgG Sepharose resulted in a product which was found to be homogeneous by polyacrylamide gel electrophoresis at pH 8.9 and by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The molecule is composed of two disulfide bonded polypeptide chains with mol wt of 50,000 and 38,000 daltons. Human CobINA was found to be a glycoprotein containing at least 10.7% carbohydrate and to have a normal serum concentration of 34 +/- 7 mug/ml (mean +/- 1 SD). Highly purified C3bINA cleaved neither free C3b nor free C4b if trace amounts of contaminating beta1H were removed from these proteins with anti-beta1H Sepharose. However, in the presence of highly purified beta1H and C3bINA, both C3bIna, both C3b and C4b were cleaved. Incubation of native C3 or C4 with C3bINA and beta1H had no effect on their cleaved. Incubation of native C3 or C4 with C3bINA and beta1H had no effect on their structure. The action of C3bINA and beta1H on C3b produced two fragments of the alpha1-chain which did not dissociate without reduction of the molecule. These fragments have mol wt of 67,000 and 40,000 daltons. The action of C3bINA and beta1H on C4b resulted in cleavage of the alpha'-chain giving rise to the 150,000-dalton C4c and the 49,000-dalton C4d fragments which dissociated without reduction. To produce from C3b the immunochemically defined C3c and C3d, fragments, the action of an additional serum enzyme appears to be required, the effect of which can be mimicked by trypsin.
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PMID:Human complement C3b inactivator: isolation, characterization, and demonstration of an absolute requirement for the serum protein beta1H for cleavage of C3b and C4b in solution. 30 46

The ability of lymphocytes to lyse human red cells coated with anti-D antibody was assessed by measuring 51 Cr release from labeled red cells incubated with peripheral blood leukocyte suspensions from 12 normal donors. Mixed mononuclear cell suspensions (containing monocytes and lymphocytes) from all donors produced lysis of sensitized red cells. Treatment with carbonyl iron reduced monocyte concentration to less than 1.2% in all donors, as measured by morphologic criteria, esterase staining and ingestion of latex particles. Lysis of red cells following monocyte depletion was markedly reduced in 8 of the 12 donors. Despite depletion of monocytes, unchanged or increased lysis was noticed with the leukocytes of the remaining 4 donors. This lysis was due to lymphocytes, not to residual monocytes. If target red cells were treated with papain or trypsin prior to sensitization, marked lysis occurred with lymphocytes of all donors, including those which did not lyse unmodified red cells. Direct cytolysis of sensitized red cells during contact with small lymphocytes was recorded using microcinematography, which confirmed the role of lymphocytes in mediating lysis. Lymphocyte-mediated lysis of red cells increased with mounting levels of antibody sensitization regardless to prior treatment with papain. Papain increased antibody coating per red cell, yet lysis per molecule of antibody bound was also increased. Lysis was inhibited by IgG1 and IgG3 in the fluid phase but not by IgG2 or IgG4. At an equivalent level of antibody sensitization lysis was augmented by concurrent coating of the red cells with C3b, C3d and/or C4b, though these components could not produce lysis in the absence of antibody coating.
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PMID:Quantitative evaluation of antibody-dependent lymphocyte-mediated lysis of human red cells. 53 2

The human regulatory complement component C4b-binding protein (C4BP) is a multimeric plasma protein, which regulates the classical pathway of the complement system. C4BP functions as a cofactor to factor 1 in the degradation of C4b and accelerates the decay rate of the C4b2a complex. Previously, we have demonstrated that monoclonal antibodies (C4-2 and 9) directed against the alpha'-chain of C4b inhibit the binding of C4b to C4BP. In order to identify the structural domain of C4b that binds C4BP, proteolytic fragments of C4 were generated with trypsin and Staphylococcus aureus V8 protease. Sodium dodecyl sulfate polyacrylamide gel electrophoresis, immunoblotting and amino acid sequence analysis of the proteolytic fragments reactive with the anti-C4 mAb's revealed that the residues Ala738-Arg826 of the alpha 3-fragment of C4b are important for the interaction with C4BP.
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PMID:Importance of the alpha 3-fragment of complement C4 for the binding with C4b-binding protein. 169 96

Extracts from myelinated and unmyelinated nerves, prepared using Nonidet P-40, contained receptors for C3b/C4b (CR1). Extracts from myelinated nerves inhibited EAC3b rosette formation with peripheral blood leucocytes and agglutinated EAC3b, whereas extract from unmyelinated nerves did not. Rosette formation with EAC3bi or EAC3d was not affected. CR1 in extracts from myelinated nerves also expressed decay-accelerating activity of the alternative pathway C3 convertase and cofactor activity in factor I-mediated cleavage of C3b, whereas CR1 in extract from unmyelinated nerves did not. Monoclonal anti-CR1 antibodies, but not monoclonal anti-CR2 (C3d receptors) or anti-CR3 (C3bi receptors) antibodies inhibited the functional activities. Accordingly, CR1 are the only C3 receptor present in the extracts and only CR1 in myelinated nerve extracts are functionally active. CR1 in both myelinated and unmyelinated nerve extracts had a molecular weight of approximately 190 kDa. The electrophoretic mobility did not change after reduction and the 190 kDa band was stained by concanavalin A, indicating that the CR1 are single-chained glycoproteins. Binding to lentil lectin-Sepharose 4B further sustained the glycoprotein nature of the CR1. Periodic acid abolished functional activities of CR1, whereas trypsin and heat did not, indicating the functional significance of the carbohydrate moiety. That CR1 are functionally active in myelinated nerves, but not in unmyelinated nerves, may therefore be due to differences in the carbohydrate moiety. The cofactor and decay-accelerating activities of CR1 may be of significance in the pathogenesis of demyelinating polyneuropathies by limiting complement activation.
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PMID:Isolation and characterization of complement receptors CR1 from human peripheral nerves. 252 22

Protein S is unique among the vitamin K-dependent proteins found in blood plasma because it is a cofactor rather than a zymogen of a serine protease. Instead of a trypsin-like domain, protein S contains a domain that has sequence homology with steroid binding proteins. In order to understand the function of this structural domain, peptides have been synthesized with amino acid sequences that are homologous between human protein S and rat androgen binding protein. Two peptides, corresponding to amino acids 400-407 (PINPRLDG) and 605-614 (GVQLDLDEAI) of the protein S sequence have been tested for their effects on protein S function. Neither peptide altered the clotting of bovine or human plasma. The peptide GVQLDLDEAI enhanced the anticoagulant activity of human-activated protein C in human plasma while the peptide PINPRLDG had no effect. The peptide GVQLDLDEAI was observed to inhibit the binding of protein S to C4b-binding protein in plasma, resulting in increased concentrations of free protein S. GVQLDLDEAI was also observed to enhance the disassociation of the protein S.C4b-binding protein complex when purified complex was used. Finally, C4b-binding protein was observed to bind to GVQLDLDEAI. These results suggest that the carboxyl-terminal region of protein S, which contains the sequence GVQLDLDEAI, is involved in the interaction between protein S and C4b-binding protein.
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PMID:Characterization of a synthetic peptide that inhibits the interaction between protein S and C4b-binding protein. 253 Feb 13

A large-scale procedure for the isolation of complement receptor type 1 (CR1, the C3b receptor) from human erythrocytes is described. Two of the four known phenotypes of CR1 are detectable in the isolated material. Amino acid and hexosamine analysis of the A phenotype (Mr 240 000) indicates a polypeptide chain length of about 2030 amino acids and a carbohydrate content of 8%. Both N- and O-linked sugars appear to be present. Trypsin digestion of isolated CR1 shows that it is degraded rapidly and extensively, and no stable products of Mr greater than 25000 are found. The ability of the receptor to bind to solid-phase ligand is destroyed after a single cleavage by trypsin. The capacity of the receptor to act as a cofactor for Factor I-mediated cleavage of soluble C3b is, however, only gradually decreased by proteolysis, and 30% of this activity remains after extensive degradation. The same pattern of loss of binding to solid-phase ligand, with partial retention of interaction with soluble ligand, is also characteristic of the complement proteins Factor H and C4bp, which are functionally related to CR1.
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PMID:Large-scale isolation of complement receptor type 1 (CR1) from human erythrocytes. Proteolytic fragmentation studies. 293 34

The effect of complement fragments C3b and C4b, on the triggering of antigen-specific human T-cell clones by Epstein-Barr virus-transformed human lymphoblastoid B cells (LCL) when these fragments are covalently coupled to the antigen tetanus toxin (TT) is described. TT was chemically cross-linked to purified C3b [(TT-C3b)n], C4b [(TT-C4b)n] or bovine serum albumin [(TT-BSA)n] as a control. T-cell activation was quantified by tritiated thymidine incorporation and 51Cr release. (TT-C3b)n and (TT-C4b)n induced proliferative responses comparable to (TT-BSA)n but at 18-25 and 4-6 lower concentrations, respectively. This enhancing effect required the covalent cross-linking of the complement fragments to the antigen and involved intracellular processing of the latter by LCL. Antigen presentation was similarly enhanced when measuring the cytotoxic activity of a helper T-cell clone against LCL previously pulsed with (TT-C3b)n or (TT-C4b)n compared with (TT-BSA)n. Binding studies, carried out on LCL using TT radiolabelled with 125I before cross-linking, indicated that (TT-C3b)n and (TT-C4b)n gave three- to four-fold more binding than (TT-BSA)n. Addition of antibodies against CR1 and CR2 or proteolytic removal of these complement receptors with trypsin inhibited by about 60% the enhancing effect of TT-bound C3b and C4b in both binding and functional assays. These results indicate that binding of C3b or C4b to antigen enhances antigen-specific proliferative and cytotoxic responses of T cells by targeting opsonized antigen onto complement receptors CR1 and CR2 of LCL. The putative significance of these findings in terms of regulation of immune responses by complement is discussed.
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PMID:Antigen-bound C3b and C4b enhance antigen-presenting cell function in activation of human T-cell clones. 297 31

We investigated the binding of C4 and C3 to red cell surfaces by non-complement enzymes. Cell bound C components were quantitated by a radioimmunoassay, the chain structure of bound components was analyzed by Western blotting and the hemolytic activity of bound components was determined. Trypsin, chymotrypsin, plasmin, elastase, thrombin, kallikrein and enzymes from Bacillus subtilis, Staphylococcus aureus and Streptomyces griseus all were found capable of binding C4b and C3b to sheep red cells. C4b bound by any of these enzymes was hemolytically active; both classical and alternate pathway activity of C3 could be demonstrated for most enzymes except plasmin and thrombin. In addition, trypsin and the bacterial enzymes were also able to generate the classical pathway C3-convertase from C4b + C2. The hemolytic efficiency of enzyme bound C4b and C3b was about the same as for these molecules bound by complement enzymes. In contrast, the process of binding by the non-complement enzymes was several hundred-fold less efficient than by cell bound complement enzymes. The results demonstrate that several enzymes can replace the C1 and C42 enzymes in the classical pathway and are able to initiate the alternative pathway by activating C3 and binding C3b to the cell surface.
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PMID:Binding and activation of C4 and C3 on the red cell surface by non-complement enzymes. 341 32

The binding and cofactor activities of C4b-binding protein were examined before and after limited proteolysis by pepsin, trypsin and chymotrypsin. The major fragments generated were characterized by amino acid sequencing, thus establishing the precise points of limited proteolysis. These studies allow a tentative assignment of the cofactor activity site to the residues 177-322 of the 549 amino acid long chain of C4b-binding protein but indicated that residues in the region 332-395 are important in the binding activity.
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PMID:Structural and functional studies on C4b-binding protein, a regulatory component of the human complement system. 393 98

An efficient procedure for the isolation of the complement-system control protein beta 1H (Factor H) from human plasma was developed. The chemical composition and physical characteristics of the protein were studied, and a sequence of 17 amino acid residues at the N-terminus was determined. Factor H is a single-polypeptide-chain glycoprotein of mol.wt. 155 000 containing 9.3% carbohydrate. Factor H is cleaved by plasma proteinases to a two-chain form. This cleavage can be mimicked by trypsin, and the two-chain form retains fully the C3b-inactivator cofactor activity of Factor H. The proteolytic fragments of Factor H are compared with those of other proteins (C4b-binding protein and erythrocyte C3b-receptor) that act as cofactors for C3b-inactivator.
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PMID:Purification and structural studies on the complement-system control protein beta 1H (Factor H). 621 18


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