Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.7 (plasmin)
9,023 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The potential cross-link acceptor sites of fibrin were specifically labeled with the fluorescent, substitute cross-link donor monodansyl cadaverine (MDC). Several fluorescent alpha-chain peptides generated from enzymatic and cyanogen bromide (CNBr) cleavage of the labeled fibrin were identified by sodium dodecyl sulfate disc gel electrophoresis; they were isolated and then characterized by amino acid analysis, NH2-terminal sequence analysis, and chromatographic and electrophoretic analyses of their digestion products. Ancrod cleavage of MDC-labeled fibrin produced a series of six alpha-chain peptides of molecular weights 34,000 to 12,000, each of which contained an MDC-labeled acceptor site, and an NH2-terminal alpha-chain derivative of molecular weight 37,500. The latter remains disulfide bound in the residual fibrin and has two MDC-labeled sit-s which are separable by CNBr cleavage. Mild plasmin digestion of MDC-labeled fibrin generated fluorescent alpha-chain peptides of molecular weights 45,000, 42,000, 35,000, 23,000, 21,000, and 2,500 in the supernatant and a nonfluorescent NH2-terminal alpha-chain derivative of molecular weight 25,000 which remained in the insoluble residual fibrin. The alignment of these plasmic supernatant peptides was determined from NH2-terminal sequence analyses which indicated that an MDC acceptor site was located at approximately residue 255 of the Aalpha-chain. Cleavage of the MDC-labeled alpha-chain by CNBr, however, localized most of its fluorescence (approximately 80%) to a fragment of molecular weight 29,000 which had the same NH2-terminal sequence as the labeled plasmic peptide of molecular weight 21,000. Both peptides were cleaved by ancrod into two acceptor site-containing peptides of approximately equal fluorescence. The preliminary NH2-terminal sequence analyses of these peptides, when combined with the above findings, indicated that these two other cross-link acceptor sites are in a peptide segment which comprises the middle 17% of the Aalpha-chain.
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PMID:Localization of the alpha-chain cross-link acceptor sites of human fibrin. 63 62

Factor XII was purified approximately 14 000-fold from bovine plasma by ammonium sulfate fractionation followed by heparin-agarose, DEAE-Sephadex, CM-cellulose, arginine-agarose, and benzamidine-agarose column chromatography. By this method, about 15 mg of protein was purified from 15 L of plasma with an overall yield of 18%. The purified protein was homogeneous as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and amino-terminal analysis. Bovine factor XII is a glycoprotein with a mol wt of 74 000 as determined by sedimentation equilibrium centrifugation. It contains 13.5% carbohydrate including 3.4% hexose, 4.7% N-acetylhexosamine, and 5.4% N-acetylneuraminic acid. Factor XII is a single polypeptide chain with an NH2-terminal sequence of Thr-Pro-Pro-Trp-Lys-Gly-Pro-?-Lys-His. This sequence is homologous to the reactive-site regions of a number of protease inhibitors. The amino acid sequence of a carboxyl-terminal fragments prepared by cyanogen bromide digestion was found to be Leu-Cys-Ala-Gly-Phe-Leu-Glu-Gly-Gly-Thr-Asp-Ala-Cys-Gln-Gly-Asp-SER-Gly-Gly-Pro-Leu-Val-Cys-Glu-Asp-Glu. This sequence is homologous with the active site of a number of plasma serine proteases including thrombin, factor IXa, factor Xa, and plasmin. These data indicate that bovine factor XII is a precursor to a serine enzyme with an inhibitor sequence and a catalytic site located in the same single polypeptide chain.
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PMID:Isolation and characterization of bovine factor XII (Hageman factor). 86 Dec 10

Characterization of the cyanogen bromide (CNBr) fragments of the beta chain of human haptoglobin revealed five major fragments resulting from cleavage of four methionyl residues. The fragments were isolated by gel filtration in guanidine-HCl on Sepharose 6B and Bio-Gel P10 and P60. Compositional analyses of the five cyanogen bromide fragments accounted for 248-253 amino acid residues in agreement with the number of residues determined for the intact beta chain. Most of the carbohydrate was attached to CNBr II. Automated amino-terminal sequence analysis and carboxyl-terminal hydrolysis with carboxypeptidase of the haptoglobin beta chain and cyanogen bromide fragments identified 139 residues, or about 55% of the beta-chain molecule. The placement of the fragments within the beta-chain molecule was established by sequence analysis of whole beta chain and a plasmin cleavage fragment. The position of CNBr V was confirmed by the absence of homoserine or homoserine lactone. Cyanogen bromide reaction of intact haptoglobin 1-1 resulted in the isolation of a beta-chain fragment, CNBr III, covalently attached to the intact alpha1 chain by a single disulfide bond. The beta chain was shown to have primary structural similarities to the chymotrypsin family of serin eproteases. Partial sequence analysis of CNBr V established the region which is comparable to the serine-195 active-site region: /Asp-Thr-Cys-Tyr-Gly-Asp-Ala-Gly-Ser-Ala-Phe/ (residues 189-199, chymotrypsinogen A numbering). The active-site serine-195 is replaced by alanine; however, the specificity residue of the trypsin-like enzymes, Asp-189, is preserved. Several minor cyanogen bromide cleavage products were also identified in yields of up to 15%. These minor cleavage products give evidence that tryptophanyl residues in proteins, or glycoproteins, are also susceptible to cyanogen bromide cleavage.
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PMID:Characterization of the cyanogen bromide fragments of the beta chain of human haptoglobin. 99 9

Plasmin(ogen) receptors are expressed by many gram-positive and gram-negative bacteria. We previously isolated a plasmin receptor from a pathogenic group A streptococcal strain (C. C. Broder, R. Lottenberg, G. O. von Mering, K. H. Johnston, and M. D. P. Boyle, J. Biol. Chem. 266:4922-4928, 1991). The gene encoding this plasmin receptor, plr, was isolated from a lambda gt11 library of chromosomal DNA from group A streptococcal strain 64/14 by screening plaques with antibodies raised against the purified streptococcal plasmin receptor protein. The gene was subcloned by using a low-copy-number plasmid and stably expressed in Escherichia coli, resulting in the production of an immunoreactive and functional receptor protein. The DNA sequence of the gene contained an open reading frame encoding 335 amino acids with a predicted molecular weight of 35,787. Upstream of the open reading frame, putative promoter and ribosomal binding site sequences were identified. The experimentally derived amino acid sequences of the N terminus and three cyanogen bromide fragments of the purified streptococcal plasmin receptor protein corresponded to the predicted sequence encoded by plr. The deduced amino acid sequence for the plasmin receptor protein revealed significant similarity (39 to 54% identical amino acid residues) to glyceraldehyde 3-phosphate dehydrogenases.
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PMID:Cloning, sequence analysis, and expression in Escherichia coli of a streptococcal plasmin receptor. 132 83

Streptokinase is an extracellular protein produced by several strains of streptococci. It functions in the species-specific conversion of plasminogen to plasmin. In this paper we describe the purification of streptokinase by affinity chromatography on human plasminogen acylated with p'-nitrophenyl p-guanidinobenzoate. The acylated and non-acylated plasminogen and plasmin were coupled to cyanogen bromide-activated Sepharose 4B and evaluated for streptokinase purification. These results show that a homogeneous preparation of streptokinase with high specific activity and high yield can be obtained using acylated plasminogen. This method permits the binding of one milligram of streptokinase per milliliter of swollen gel.
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PMID:Purification of streptokinase by affinity chromatography on immobilized acylated human plasminogen. 153 7

The rate of activation of plasminogen by tissue-type plasminogen activator (t-PA) is greatly increased by fibrin, but much less by fibrinogen. Fibrin(ogen) fragments such as the fibrin(ogen) cyanogen bromide fragment FCB-2 and FCB-5, and a synthetic peptide with the sequence of fibrinogen A alpha-(148-160), a constituent of FCB-2, also have rate-enhancing properties. In order to find a possibly smaller, still stimulating site within A alpha-(148-160) we synthesized successive linear amino-terminally acylated hexapeptides [i.e. A alpha-(148-153), A alpha-(149-154)'d, .... A alpha-(155-160)] from the sequence A alpha-(148-160). The only hexapeptide within the sequence A alpha-(148-160) capable of enhancing the rate of plasminogen-to-plasmin conversion by t-PA appears to be the amino-terminally acylated peptide comprising the sequence A alpha-(154-159). This peptide enhances the plasminogen activation rate six-fold; half-maximal activation rate is reached at a peptide concentration of 56 microM.
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PMID:The sequence A alpha-(154-159) of fibrinogen is capable of accelerating the t-PA catalysed activation of plasminogen. 193 32

A small but consistent proportion of the von Willebrand factor (vWF) in normal plasma is composed of 189, 176, and 140 kD fragments cleaved from the 225 kD subunit. A monoclonal antibody map of vWF, based on the reactivity of individual antibodies with cyanogen bromide and tryptic fragments of known carboxy and/or amino termini, showed that in normal and IIA von Willebrand disease (vWD) plasmas the 140 kD fragment was derived from the amino-terminal region, whereas the 176 kD fragment was derived from the carboxy-terminal region of the subunit. In type IIA vWD, however, the fragments comprised a greater proportion of circulating vWF. In contrast, plasmin cleaved a 176 kD fragment from the amino terminus and a 145 kD fragment from the carboxy terminus of the subunit. Species similar to these plasmin-cleaved fragments were demonstrated in plasmas from four patients treated with fibrinolytic agents, but not in IIA vWD.
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PMID:Epitope mapping of the von Willebrand factor subunit distinguishes fragments present in normal and type IIA von Willebrand disease from those generated by plasmin. 243 8

Sulfation of human alpha 2-antiplasmin, the major plasma inhibitor of fibrinolysis, was examined using both protein isolated from human plasma and protein synthesized and biosynthetically labeled with [35S]sulfate by a human hepatoma-derived cell line. Linkage of sulfate to tyrosine was demonstrated by recovery of labeled tyrosine sulfate after base hydrolysis of sulfate-labeled alpha 2-antiplasmin. Analysis by reverse-phase high performance liquid chromatography of peptides released from alpha 2-antiplasmin by cleavage with trypsin or cyanogen bromide indicated that sulfate is linked to a single segment of the protein. A cyanogen bromide peptide corresponding to the sulfate-labeled peptide was prepared from alpha 2-antiplasmin isolated from human plasma. Consistent with the presence of tyrosine sulfate in this peptide, its chromatographic elution was altered by treatment with acid under conditions which release sulfate from a tyrosine residue. No peptide in the total digest of alpha 2-antiplasmin by cyanogen bromide eluted at the position of the peptide following desulfation, suggesting that all of the protein is in a sulfated form. The sequence of the sulfate-containing cyanogen bromide peptide as determined by sequential Edman degradation, amino acid composition, and fast atom-bombardment-mass spectrometry was: Glu-Glu-Asp-Tyr(SO4)-Pro-Gln-Phe-Gly-Ser-Pro-Lys-COOH. This peptide is a segment of the previously identified plasmin-binding domain of alpha 2-antiplasmin.
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PMID:Sulfation of a tyrosine residue in the plasmin-binding domain of alpha 2-antiplasmin. 243 96

The complete amino acid sequence of canine miniplasminogen (Mr 36,678, 333 residues) was determined with the aid of fragments obtained by cleavage with BNPS-skatole, cyanogen bromide or clostripain. The fragments were aligned with overlapping sequences. Sequence comparison with miniplasminogens of other species gave identities in the range of 80% (bovine) and 88% (human), indicating the presence of the same structural and functional domains as in the other species. Sequence comparison of different miniplasminogens showed that plasminogens of species activated by streptokinase have identical residues in positions 49, 83 and 161 of the plasmin light chain. The triad of these amino acids may represent at least one of eventually several prerequisites for the interaction and activation of plasminogen with streptokinase.
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PMID:Complete amino acid sequence of canine miniplasminogen. 262 24

Arterial thrombi and atherosclerotic lesions were analyzed immunochemically and examined histologically. The extent of in vivo proteolytic cleavage of the amino-terminal end of fibrinogen by thrombin and plasmin was determined and quantitated by specific radioimmunoassays. The samples were treated with cyanogen bromide (CNBr), and the total amount of fibrinogen and fibrin-derived protein was determined as NDSK, the NH2-terminal disulfide knot of fibrinogen. Thrombin-releasable fibrinopeptides A and B were used to quantitate fibrinogen and fibrin I. Previous plasmin cleavage of the B beta chain was inferred from the amount of B beta 1-42 and B beta 15-42 in undigested NDSK. The results obtained in both acute and organized thrombi indicate that approximately 60% of the total protein (as determined by amino acid analysis) was fibrinogen-derived and that 70% to 80% of the fibrinogen-derived material was fibrin II. These findings support the hypothesis that fibrin II as distinct from fibrin I is the predominant component in a thrombus. In samples from normal and atherosclerotic aortas, fibrinogen-derived protein comprised less than 10% of the total protein. Samples from grossly normal aortas contained only fibrinogen and fibrin I. Fibrinogen concentration decreased and fibrin II concentration increased with increasing severity of the lesions, suggesting that increased fibrin II formation is associated with progression of atheromas.
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PMID:Immunochemical characterization of fibrinogen, fibrin I, and fibrin II in human thrombi and atherosclerotic lesions. 295 Sep 41


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