Skizzle is a novel plasminogen- and plasmin-binding protein from Streptococcus agalactiae that targets proteins of human fibrinolysis to promote plasmin generation.
Skizzle (SkzL), secreted by Streptococcus agalactiae, has moderate sequence identity to streptokinase and staphylokinase, bacterial activators of human plasminogen (Pg). SkzL binds [Glu]Pg with low affinity (K(D) 3-16 mum) and [Lys]Pg and plasmin (Pm) with indistinguishable high affinity (K(D) 80 and 50 nm, respectively). Binding of SkzL to Pg and Pm is completely lysine-binding site-dependent, as shown by the effect of the lysine analog, 6-aminohexanoic acid. Deletion of the COOH-terminal SkzL Lys(415) residue reduces affinity for [Lys]Pg and active site-blocked Pm 30-fold, implicating Lys(415) in a lysine-binding site interaction with a Pg/Pm kringle. SkzL binding to active site fluorescein-labeled Pg/Pm analogs demonstrates distinct high and low affinity interactions. High affinity binding is mediated by Lys(415), whereas the source of low affinity binding is unknown. SkzL enhances the activation of [Glu]Pg by urokinase (uPA) approximately 20-fold, to a maximum rate indistinguishable from that for [Lys]Pg and [Glu]Pg activation in the presence of 6-aminohexanoic acid. SkzL binds preferentially to the partially extended beta-conformation of [Glu]Pg, which is in unfavorable equilibrium with the compact alpha-conformation, thereby converting [Glu]Pg to the fully extended gamma-conformation and accelerating the rate of its activation by uPA. SkzL enhances [Lys]Pg and [Glu]Pg activation by single-chain tissue-type Pg activator, approximately 42- and approximately 650-fold, respectively. SkzL increases the rate of plasma clot lysis by uPA and single-chain tissue-type Pg activator approximately 2-fold, confirming its cofactor activity in a physiological model system. The results suggest a role for SkzL in S. agalactiae pathogenesis through fibrinolytic enhancement.
Wiles
Karen G
KG
Department of Pathology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA.
Panizzi
Peter
P
Kroh
Heather K
HK
Bock
Paul E
PE
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017124:Conserved Sequence; D005341:Fibrinolysin / Q000096:biosynthesis* / Q000378:metabolism*; D005342:Fibrinolysis / Q000502:physiology*; D006801:Humans; D007700:Kinetics; D008666:Metalloendopeptidases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010958:Plasminogen / Q000378:metabolism*; D011485:Protein Binding; D013292:Streptococcus agalactiae / Q000378:metabolism*; D013300:Streptokinase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013379:Substrate Specificity
D001426:Bacterial Proteins; D010958:Plasminogen; D013300:Streptokinase; D005341:Fibrinolysin; D008666:Metalloendopeptidases; C022653:auR protein, Staphylococcus aureus