The mechanism of a bacterial plasminogen activator intermediate between streptokinase and staphylokinase.
The therapeutic properties of plasminogen activators are dictated by their mechanism of action. Unlike staphylokinase, a single domain protein, streptokinase, a 3-domain (alpha, beta, and gamma) molecule, nonproteolytically activates human (h)-plasminogen and protects plasmin from inactivation by alpha(2)-antiplasmin. Because a streptokinase-like mechanism was hypothesized to require the streptokinase gamma-domain, we examined the mechanism of action of a novel two-domain (alpha,beta) Streptococcus uberis plasminogen activator (SUPA). Under conditions that quench trace plasmin, SUPA nonproteolytically generated an active site in bovine (b)-plasminogen. SUPA also competitively inhibited the inactivation of plasmin by alpha(2)-antiplasmin. Still, the lag phase in active site generation and plasminogen activation by SUPA was at least 5-fold longer than that of streptokinase. Recombinant streptokinase gamma-domain bound to the b-plasminogen.SUPA complex and significantly reduced these lag phases. The SUPA-b.plasmin complex activated b-plasminogen with kinetic parameters comparable to those of streptokinase for h-plasminogen. The SUPA-b.plasmin complex also activated h-plasminogen but with a lower k(cat) (25-fold) and k(cat)/K(m) (7.9-fold) than SK. We conclude that a gamma-domain is not required for a streptokinase-like activation of b-plasminogen. However, the streptokinase gamma-domain enhances the rates of active site formation in b-plasminogen and this enhancing effect may be required for efficient activation of plasminogen from other species.
Cardiovascular Biology Laboratory, Harvard School of Public Health and the Massachusetts General Hospital, Boston, Massachusetts 02114, USA.
I Y
IY
Sazonova
A K
AK
Houng
S A
SA
Chowdhry
B R
BR
Robinson
L
L
Hedstrom
G L
GL
Reed
D000595:Amino Acid Sequence; D000818:Animals; D000933:Antifibrinolytic Agents / Q000494:pharmacology; D001665:Binding Sites; D002417:Cattle; D003001:Cloning, Molecular; D005341:Fibrinolysin / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008666:Metalloendopeptidases / Q000737:chemistry* / Q000378:metabolism*; D008969:Molecular Sequence Data; D010958:Plasminogen / Q000378:metabolism; D010960:Plasminogen Activators / Q000737:chemistry* / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013291:Streptococcus / Q000201:enzymology*; D013300:Streptokinase / Q000737:chemistry* / Q000378:metabolism*
D000933:Antifibrinolytic Agents; D011994:Recombinant Proteins; D010958:Plasminogen; D013300:Streptokinase; D010960:Plasminogen Activators; D005341:Fibrinolysin; D008666:Metalloendopeptidases; C022653:auR protein, Staphylococcus aureus