Localization of the binding site of tissue-type plasminogen activator to fibrin.
01 Jul 1986-Journal of Clinical Investigation (American Society for Clinical Investigation)-Vol. 78, Iss: 1, pp 163-169
TL;DR: Results indicated that the binding site of tissue-type plasminogen activator to fibrin was located in the kringle-2 segment, which was found to be responsible for the binding to lysine-Sepharose or fibrIn.
Abstract: Functionally active A and B chains were separated from a two-chain form of recombinant tissue-type plasminogen activator after mild reduction and alkylation. The A chain was found to be responsible for the binding to lysine-Sepharose or fibrin and the B chain contained the catalytic activity of tissue-type plasminogen activator. An extensive reduction of two-chain tissue-type plasminogen activator, however, destroyed both the binding and catalytic activities. A thermolytic fragment, Fr. 1, of tissue-type plasminogen activator that contained a growth factor and two kringle segments retained its lysine binding activity. Additional thermolytic cleavages in the kringle-2 segment of Fr. 1 caused a total loss of the binding activity. These results indicated that the binding site of tissue-type plasminogen activator to fibrin was located in the kringle-2 segment.
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TL;DR: This work has shown that the protease Nexin I antagonist acts as a “spatially aggregating force” to drive down the activity of the plasminogen during the courtship process.
Abstract: PLASMINOGEN ACTIVATOR INHIBITORS 104 Plasminogen Activator Inhibitor 1 104 Plasminogen Activator Inhibitor 2 106 Protease Nexin I .... .... 108 PHYSIOLOGICAL FUNCTIONS OF PLASMINOGEN ACTIVATION ........ 109 Matrix Breakdown 109 Cell Migration 111 Ovulation 113 CONCLUDING REMARKS 115
877 citations
TL;DR: The authors reviewed the epidemiology, pathogenesis and various prevention strategies of adhesion formation, using Medline and PubMed search, and found that several preventive agents against postoperative peritoneal adhesions have been investigated but most are contradictory and achieved mostly in animal model.
Abstract: Peritoneal adhesions represent an important clinical challenge in gastrointestinal surgery. Peritoneal adhesions are a consequence of peritoneal irritation by infection or surgical trauma, and may be considered as the pathological part of healing following any peritoneal injury, particularly due to abdominal surgery. The balance between fibrin deposition and degradation is critical in determining normal peritoneal healing or adhesion formation. Postoperative peritoneal adhesions are a major cause of morbidity resulting in multiple complications, many of which may manifest several years after the initial surgical procedure. In addition to acute small bowel obstruction, peritoneal adhesions may cause pelvic or abdominal pain, and infertility. In this paper, the authors reviewed the epidemiology, pathogenesis and various prevention strategies of adhesion formation, using Medline and PubMed search. Several preventive agents against postoperative peritoneal adhesions have been investigated. Their role aims in activating fibrinolysis, hampering coagulation, diminishing the inflammatory response, inhibiting collagen synthesis or creating a barrier between adjacent wound surfaces. Their results are encouraging but most of them are contradictory and achieved mostly in animal model. Until additional findings from future clinical researches, only a meticulous surgery can be recommended to reduce unnecessary morbidity and mortality rates from these untoward effects of surgery. In the current state of knowledge, pre-clinical or clinical studies are still necessary to evaluate the effectiveness of the several proposed prevention strategies of postoperative peritoneal adhesions.
363 citations
TL;DR: TISSUE plasminogen activator (t-PA) is a naturally occurring protein that catalyzes the conversion of the inactive proenzyme plAsminogen into the active serine protease plasmine.
Abstract: TISSUE plasminogen activator (t-PA) is a naturally occurring protein that catalyzes the conversion of the inactive proenzyme plasminogen into the active serine protease plasmin. It is one of two ma...
277 citations
TL;DR: The organization and structure of the gene coding for plasminogen has been determined by a combination of in vitro amplification of leukocyte DNA from normal individuals and isolation of unique clones from three different human genomic libraries.
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TL;DR: Plasminogen was prepared from human plasma by affinity chromatography on L-lysine-substituted Sepharose with a specific activity of 100 caseinolytic units per milligram of nitrogen.
Abstract: Plasminogen was prepared from human plasma by affinity chromatography on L-lysine-substituted Sepharose. Thirty milligrams of plasminogen, with a specific activity of 100 caseinolytic units (Committee on Thrombolytic Agents) per milligram of nitrogen, were obtained from 340 milliliters of plasma. This corresponds to over 200-fold purification from plasma. Disc-gel electrophoresis at pH 8.3 indicated seven distinct bands, all of which contained activity.
2,051 citations
TL;DR: Bacterial clones containing human tissue-type plasminogen activator cDNA sequences were identified in a cDNA library prepared using gel-fractionated mRNA from human melanoma cells and a polypeptide was produced having the fibrinolytic properties characteristic of authentic human t-PA.
Abstract: Bacterial clones containing human tissue-type plasminogen activator (t-PA) cDNA sequences were identified in a cDNA library prepared using gel-fractionated mRNA from human melanoma cells. A plasmid containing the Escherichia coli trp promoter and the cDNA sequence coding f or the 527-amino acid mature t-PA protein was constructed for expression in E. coli. A polypeptide was produced having the fibrinolytic properties characteristic of authentic human t-PA.
1,219 citations
TL;DR: In immunodiffusion, as well as in quenching experiments of the fibrinolytic activities, the melanoma plasminogen activator appeared to be immunologically identical with the uterine tissue plasmineg activator, but unrelated to urokinase.
784 citations
TL;DR: A genomic clone carrying the human tissue-type plasminogen activator (t-PA) gene was isolated from a cosmid library, and the gene structure was elucidated by restriction mapping, Southern blotting, and DNA sequencing, revealing a gene organization similar to other serine proteases.
Abstract: A genomic clone carrying the human tissue-type plasminogen activator (t-PA) gene was isolated from a cosmid library, and the gene structure was elucidated by restriction mapping, Southern blotting, and DNA sequencing. The cosmid contained all the coding parts of the mRNA, except for the first 58 bases in the 5' end of the mRNA, and had a total length of greater than 20 kilobases. It was separated into at least 14 exons by at least 13 introns, and the exons seemed to code for structural or functional domains. Thus, the signal peptide, the propeptide, and the domains of the heavy chain, including the regions homologous to growth factors, and to the "finger" structure of fibronectin, are all encoded by separate exons. In addition, the two kringle regions of t-PA were both coded for by two exons and were cleaved by introns at identical positions. The region coding for the light chain, comprising the serine protease part of the molecule was split by four introns, revealing a gene organization similar to other serine proteases.
307 citations
TL;DR: The sequence data presented complete the primary structure of high molecular mass urokinase from human urine, and revealed a significant homology to peptide chains of other serine proteinases.
Abstract: The complete sequence of 157 amino acids of the light (A) chain of high molecular mass urokinase from human urine was determined. The fragmentation strategy included cyanogen bromide cleavage of the S-carboxymethylated A chain at the methionine and/or tryptophan residues and use of the specific endoproteinase Lys-C. For sequence determination automated solid- or liquid-phase techniques of Edman degradation were used. C-terminal amino acids of the A chain were determined by consecutive treatment with carboxypeptidase A and B. The amino acid sequence obtained revealed a significant homology to peptide chains of other serine proteinases. Accordingly, the sequence of the A chain can be divided into three domains: 1) The growth factor domain with homologies to murine epidermal growth factor and a particular sequence of bovine clotting factor X, 2) The "kringle" domain with homologies to "kringle" structures, e.g. in plasminogen, and 3) the connecting peptide domain containing the A1 chain of low molecular mass urokinase. Together with the amino acid sequence of the B chain, which was presented by us in an earlier communication, the sequence data presented complete the primary structure of high molecular mass urokinase from human urine.
292 citations