Kinetic measurements of T----R structural transitions in insulin

Abstract

The T6----T3R3 and T3R3----R6-structural transitions of cobalt insulin hexamers as induced by SCN ions or m-cresol were studied in stopped-flow experiments using the absorption in the visible for monitoring their time course. The T6----T3R3 transition induced by either SCN or limited concentrations of m-cresol is mono-exponential with a rate constant of 0.1 s-1 and 0.4 s-1, respectively. A mono-exponential time course is also encountered for the m-cresol-induced T3R3----R6 transition when starting from the T3R3 state preestablished by either SCN or m-cresol. The corresponding rate constants are 1.3 s-1 and 0.49 s-1, respectively. If m-cresol is used beyond the concentration range where transformation is limited to one trimer, two exponentials are required for fitting the time course. The second exponential corresponds to the T3R3----R6 step with a concentration-independent rate constant of 0.4 s-1. The rate constant for the faster T6----T3R3 transition, however, increases with increasing excess of m-cresol.