Structural and functional basis of plasminogen activation by staphylokinase

Thromb Haemost. 1999 Apr;81(4):479-85.

Abstract

Staphylokinase (Sak), a 15.5-kDa bacterial protein, forms a complex with human plasmin, which in turn activates other plasminogen molecules to plasmin. Three recombinant DNA-based approaches, (i) site directed substitution with alanine, (ii) search for proximity relationships at the complex interface, and (iii) active-site accessibility to protease inhibitors have been used to deduce a coherent docking model of the crystal structure of Sak on the homology-based model of microplasmin (microPli), the serine protease domain of plasmin. Sak binding on microPli is primarily mediated by two surface-exposed loops, loops 174 and 215, at the rim of the active-site cleft, while the binding epitope of Sak on microPli involves several residues located in the flexible NH2-terminal arm and in the five-stranded mixed beta-sheet. Several Sak residues located within the unique alpha-helix and the beta2 strand do not contribute to the binding epitope but are essential to induce plasminogen activating potential in the Sak:microPli complex. These residues form a topologically distinct activation epitope, which, upon binding of Sak to the catalytic domain of microPli, protrudes into a broad groove near the catalytic triad of microPli, thereby generating a competent binding pocket for micro-plasminogen (microPlg), which buries approximately 2500 A of the Sak:microPli complex upon binding. This structural and functional model may serve as a template for the design of improved Sak-derived thrombolytic agents. Following the completion and presentation of the present study, the deduced Sak:microPli:microPlg complex was fully confirmed by X-ray crystallography, which further illustrates the power and potential of the present approach.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacteriophages
  • Binding Sites
  • Drug Interactions
  • Enzyme Activation / drug effects
  • Enzyme Activation / physiology
  • Fibrinolysin / genetics
  • Genetic Variation
  • Humans
  • Metalloendopeptidases / genetics
  • Metalloendopeptidases / pharmacology*
  • Mutation
  • Plasminogen / metabolism*
  • Plasminogen Activators / chemistry*
  • Plasminogen Activators / pharmacology
  • Plasminogen Activators / physiology*
  • Protease Inhibitors / chemistry
  • Protein Structure, Tertiary
  • Recombinant Proteins / genetics
  • Staphylococcus / virology

Substances

  • Protease Inhibitors
  • Recombinant Proteins
  • Plasminogen
  • Plasminogen Activators
  • Fibrinolysin
  • Metalloendopeptidases
  • auR protein, Staphylococcus aureus