In:
Thrombosis and Haemostasis, Georg Thieme Verlag KG, Vol. 76, No. 05 ( 1996), p. 755-760
Abstract:
Structure/function relationships in the activation of plasminogen with staphylokinase were studied using mutants of recombinant staphylokinase (Sak42D). Deletion of up to 10 NH2-terminal amino acids (Sak42DΔN10) did not affect plasminogen activation, but removal of 11 amino acids completely abolished the ability to activate plasminogen. Elimination of potential plasmin cleavage sites in the NH2-terminal region yielding mutants Sak42D(K8H,K10H,KllH) and Sak42D(K6H,K8H,KllH) did not alter the rate of the exposure of a proteolytically active site (amidolytic activity) in equimolar mixtures with plasminogen, but destroyed the plasminogen activator properties of these muteins. Deleting two residues following the preferred processing site at position 10 (Sak42Δ(K11,G12)) resulted in a mutein also inactive in plasminogen activation. Removal of the COOH-terminal Lys136, yielding Sak42DΔCl, or of Lysl35 and Lysl36 in Sak42DΔC2 resulted in proteins with strongly reduced plasminogen activation capacity. In contrast, substitution of Lys135 and Lys136 with Ala in Sak42D(K135A,K136A) did not affect activation. Cyanogen bromide cleavage of Sak42D(M26L,E61M,D82E) produced a 61 amino acid NH2-terminal and a 65 amino acid COOH-terminal fragment which did not activate plasminogen, but bound to plasminogen with affinity constants Ka of 4.0 × 105 M−1 and 1.4 × 107 M−1, respectively (as compared to a Ka of 1.1 × 108 M−1 for Sak42D). These results indicate that Lysl 1 and the COOH-terminal region of staphylokinase play a key role in the activation of plasminogen.
Type of Medium:
Online Resource
ISSN:
0340-6245
,
2567-689X
DOI:
10.1055/s-0038-1650656
Language:
English
Publisher:
Georg Thieme Verlag KG
Publication Date:
1996
