The meaning of the Michaelis-Menten constant: Kmdescribes a steady-state

Often, in vitro or in vivo enzyme-mediated catalytic events occur far from equilibrium and, therefore, substrate affinity measured as the inverse of ES ⇄ E+S dissociation equilibrium constant ( K d ) has a doubtful physiological meaning; in practice it is almost impossible to determine K d (except using stopped-flow or other sophisticated methodologies). The Michaelis-Menten constant ( K m ), the concentration of substrate ([S]) providing half of enzyme maximal activity, is not the ( K d ). In the simple E+S ⇄ ES → E+P or in more complex models describing S conversion into P, K m must be considered the constant defining the steady state at any substrate concentration. Enzyme kinetics is based on initial rate determination, i.e. in the linear part of the S to P conversion when the concentration of [ES] remains constant while steady state occurs. We also show that Systems Biology issues such as the time required to respond to a system perturbation, is more dependent on k 1 , the kinetic constant defining substrateenzyme association, than on K m . Whereas K m is instrumental for biochemical basic and applied approaches, in any physiological condition, an important parameter to be considered is the substrate association rate ( k 1 ).