Bibliographic Citation
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| Title | Functional role of proteolytic cleavage at arginine-275 of human tissue plasminogen activator as assessed by site-directed mutagenesis |
| Creator/Author | Tate, K.M. ; Higgins, D.L. ; Holmes, W.E. ; Winkler, M.E. ; Heyneker, H.L. ; Vehar, G.A. |
| Publication Date | 1987 Jan 27 |
| OSTI Identifier | OSTI ID: 6790677 |
| Other Number(s) | Journal ID: CODEN: BICHA |
| Resource Type | Journal Article |
| Resource Relation | Journal Name: Biochemistry; (United States); Journal Volume: 26:2 |
| Research Org | Genentech, Inc., South San Francisco, CA |
| Subject | 62 RADIOLOGY AND NUCLEAR MEDICINE; PEPTIDE HYDROLASES; AUTORADIOGRAPHY; MUTAGENESIS; ARGININE; ELECTROPHORESIS; ENZYMATIC HYDROLYSIS; FIBRINOLYSIN; GLUTAMIC ACID; IODINE 125; ION EXCHANGE CHROMATOGRAPHY; PLASMINOGEN; SERINE; SODIUM IODIDES; STAPHYLOCOCCUS; ALKALI METAL COMPOUNDS; AMINO ACIDS; BACTERIA; BETA DECAY RADIOISOTOPES; CARBOXYLIC ACIDS; CHEMICAL REACTIONS; CHROMATOGRAPHY; DAYS LIVING RADIOISOTOPES; DECOMPOSITION; DRUGS; ELECTRON CAPTURE RADIOISOTOPES; ENZYMES; FIBRINOLYTIC AGENTS; HALIDES; HALOGEN COMPOUNDS; HEMATOLOGIC AGENTS; HYDROLASES; HYDROLYSIS; HYDROXY ACIDS; INORGANIC PHOSPHORS; INTERMEDIATE MASS NUCLEI; IODIDES; IODINE COMPOUNDS; IODINE ISOTOPES; ISOTOPES; LYSIS; MICROORGANISMS; NUCLEI; ODD-EVEN NUCLEI; ORGANIC ACIDS; ORGANIC COMPOUNDS; PHOSPHORS; RADIOISOTOPES; SEPARATION PROCESSES; SERINE PROTEINASES; SODIUM COMPOUNDS; SOLVOLYSIS |
| Description/Abstract | Activation of the zymogen form of a serine protease is associated with a conformational change that follows proteolysis at a specific site. Tissue-type plasminogen activator (t-PA) is homologous to mammalian serine proteases and contains an apparent activation cleavage site at arginine-275. To clarify the functional consequences of cleavage at arginine-275 of t-PA, site-specific mutagenesis was performed to convert arginine-275 to a glutamic acid. The mutant enzyme (designated Arg-275 ..-->.. Glu t-PA) could be converted to the two-chain form by Staphylococcus aureus V8 protease but not by plasmin. The one-chain form was 8 times less active against the tripeptide substrate H-D-isoleucyl-L-prolyl-L-arginine-rho-nitroanilide (S-2288), and the ability of the enzyme to activate plasminogen in the absence of fibrinogen was reduced 20-50 times compared to the two-chain form. In contrast, one-chain Arg-275 ..-->.. Glu t-PA has equal activity to the two-chain form when assayed in the presence of physiological levels of fibrinogen and plasminogen. Fibrin bound significantly more of the one-chain form of t-PA than the two-chain form for both the wild-type and mutated enzymes. One- and two-chain forms of the wild-type and mutated plasminogen activators slowly formed complexes with plasma protease inhibitors, although the one-chain forms showed decreased complex formation with ..-->../sub 2/-macroglobulin. The one-chain form of t-PA therefore is fully functional under physiologic conditions and has a increased fibrin binding compared to the two-chain form. |
| Country of Publication | United States |
| Language | English |
| Format | Medium: X; Size: Pages: 338-343 |
| System Entry Date | 2009 Dec 17 |
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