Kinetic applications using high substrate and competitive inhibitor concentrations to determine Ki or Km

Anal Biochem. 1994 Aug 15;221(1):53-6. doi: 10.1006/abio.1994.1377.


Conventional procedures for determining Km or Ki values generally employ subsaturating concentrations of substrate and competitive inhibitor; however, this is impractical in some cases if a significant percentage would be enzyme-bound or metabolized. Applications employing high and competing concentrations of substrates and inhibitors are described. Ki or Km values can be calculated from the equations. Ki = [I]/((V0)/V - 1)(1 + [S]/Km)) Km = [S](V0/V - 1)/(1 - V0/V + [I]/Ki), where V0 and V are the rates in the absence and presence of inhibitor. This is illustrated by measurements of inhibition of human urinary kallikrein by synthetic peptides.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Kallikreins / antagonists & inhibitors
  • Kallikreins / metabolism*
  • Kinetics
  • Mathematics


  • Kallikreins