Lysine 156 promotes the anomalous proenzyme activity of tPA: X-ray crystal structure of single-chain human tPA.
Martin Renatus,Richard A. Engh,Richard A. Engh,Milton T. Stubbs,Robert Huber,Stephan Fischer,Ulrich Kohnert,Wolfram Bode +7 more
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TLDR
Comparisons with the structures of other serine proteinases that also possess Lys156, such as trypsin, factor Xa and human urokinase plasminogen activator (uPA), identify a set of secondary interactions which are required for Lys156 to fulfil this activating role.Abstract:
Tissue type plasminogen activator (tPA) is the physiological initiator of fibrinolysis, activating plasminogen via highly specific proteolysis; plasmin then degrades fibrin with relatively broad specificity. Unlike other chymotrypsin family serine proteinases, tPA is proteolytically active in a single-chain form. This form is also preferred for therapeutic administration of tPA in cases of acute myocardial infarction. The proteolytic cleavage which activates most other chymotrypsin family serine proteinases increases the catalytic efficiency of tPA only 5- to 10-fold. The X-ray crystal structure of the catalytic domain of recombinant human single-chain tPA shows that Lys156 forms a salt bridge with Asp194, promoting an active conformation in the single-chain form. Comparisons with the structures of other serine proteinases that also possess Lys156, such as trypsin, factor Xa and human urokinase plasminogen activator (uPA), identify a set of secondary interactions which are required for Lys156 to fulfil this activating role. These findings help explain the anomalous single-chain activity of tPA and may suggest strategies for design of new therapeutic plasminogen activators.read more
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References
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A slow clearing, fibrin-specific, PAI-1 resistant variant of t-PA (T103N, KHRR 296-299 AAAA)
Nicholas F. Paoni,Bruce Keyt,Canio J. Refino,Alice M. Chow,Hung V. Nguyen,L. Berleau,J M Badillo,Luis C. Pena,K. Brady,Florian M. Wurm +9 more
TL;DR: These results show that it is possible to combine mutations in different domains of t-PA to construct a variant which is simultaneously slower clearing, less reactive towards plasminogen in the absence of a fibrin clot, and resistant to inactivation by PAI-1.
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Converting tissue-type plasminogen activator into a zymogen.
Kathy Tachias,Edwin L. Madison +1 more
TL;DR: It is proposed that the presence or absence of an acidic residue at position 144 (chymotrypsin numbering system) is the primary determinant of the distinct zymogenicities of the two enzymes, t-PA and u-PA.
Journal ArticleDOI
Converting Tissue Type Plasminogen Activator into a Zymogen IMPORTANT ROLE OF Lys156
Kathy Tachias,Edwin L. Madison +1 more
TL;DR: One of the variants characterized in this study, t-PA/R15E,K156Y, possessed substantially enhanced response to and selectivity among fibrin co-factors, resistance to inhibition by plasminogen activator inhibitor type 1, and significantly increased zymogenicity.
High resolution analysis
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Enzymatic activities of activated sludge
M. Teuber,K. E. U. Brodisch +1 more
TL;DR: It is established that the corresponding phosphatase, cyclic phosphodiesterase, glycosidase and aminopeptidase activities can be conveniently tested with diluted activated sludge within 10–20 min incubation at 30°C.