Determination of substrate binding energies in individual subsites of a family 18 chitinase.

Thermodynamic parameters for binding of N-acetylglucosamine (GlcNAc) oligomers to a family 18 chitinase, ChiB of Serratia marcescens, have been determined using isothermal titration calorimetry. Binding studies with oligomers of different lengths showed that binding to subsites -2 and +1 is driven by a favorable enthalpy change, while binding to the two other most important subsites, +2 and +3, is driven by entropy with unfavorable enthalpy. These remarkable unfavorable enthalpy changes are most likely due to favorable enzyme-substrate interactions being offset by unfavorable enthalpic effects of the conformational changes that accompany substrate-binding.