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Characterization of Lys‐698 to met substitution in human plasminogen catalytic domain
Author(s) -
Terzyan Simon,
Wakeham Nancy,
Zhai Peng,
Rodgers Karla,
Zhang Xuejun C.
Publication year - 2004
Publication title -
proteins: structure, function, and bioinformatics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.699
H-Index - 191
eISSN - 1097-0134
pISSN - 0887-3585
DOI - 10.1002/prot.20070
Subject(s) - chemistry , recombinant dna , protease , biochemistry , activator (genetics) , plasminogen activator , serine protease , plasmin , mutant , enzyme , receptor , biology , gene , endocrinology
Streptokinase (SK) is a human plasminogen (Pg) activator secreted by streptococci. The activation mechanism of SK differs from that of physiological Pg activators in that SK is not a protease and cannot proteolytically activate Pg. Instead, it forms a tight complex with Pg that proteolytically activates other Pg molecules. The residue Lys‐698 of human Pg was hypothesized to participate in triggering activation in the SK–Pg complex. Here, we report a study of the Lys‐698 to Met substitution in the catalytic domain of Pg (μPg) containing the proteolytic activation‐resistant background (R561A). While it remains competent in forming a complex with SK, maintaining a comparable equilibration dissociation constant ( K D ), the recombinant protein shows a nearly 60‐fold reduction in amidolytic activity relative to its R561A background when mixed with native SK. A 2.3 Å crystal structure of this mutant μPg confirmed the correct folding of this recombinant protein. Combined with other biochemical data, these results support the premise that Lys‐698 of human Pg plays a functional role in the so‐called N‐terminal insertion activation mechanism by SK. Proteins 2004. © 2004 Wiley‐Liss, Inc.