QUANTITATION OF CHEMICAL-EXCHANGE RATES USING PULSED-FIELD-GRADIENT DIFFUSION MEASUREMENTS

A new approach to the quantitation of chemical exchange rates is prese nted, and its utility is illustrated with application to the exchange of protein amide protons with bulk water. The approach consists of a s elective-inversion exchange HMQC experiment in which a short spin echo diffusion filter has been inserted into the exchange period. In this way, the kinetics of exchange are encoded directly in an apparent diff usion coefficient which is a function of the position of the diffusion filter in the pulse sequence. A detailed theoretical analysis of this experiment indicates that, in addition to the measurement of simple e xchange rates, the experiment is capable of measuring the effect of me diated exchange, e.g. the transfer of magnetization from bulk water to an amide site mediated by an internal bound water molecule or a labil e protein side-chain proton in fast exchange with bulk water. Experime ntal results for rapid water/amide exchange in acyl carrier protein ar e shown to be quantitatively consistent with the exchange rates measur ed using a selective-inversion exchange experiment.