Temperature and pH dependence of the proton magnetic hyperfine resonances of cytochrome c peroxidase-cyanide. Evidence for hindered vinyl group rotation as a mediator of the enzymes activity.

Abstract

Proton nmr studies of the hyperfine resonances of cytochrome c peroxidase reveal that two pH-dependent processes can be monitored. One of these is the simple pH titration of a resonance which has been previously assigned to the alpha-vinyl proton at heme position 4. Combined with this proton's temperature dependence, the pH data indicate that the rotational position of vinyl 4 is changing with a pK which is similar to that which regulates the enzyme's activity. The second process, slow on the nmr time scale, occurs above pH 8. This is beyond what is normally considered to be the optimum pH range for cytochrome c peroxidase's activity and we interpret this to indicate a protein conformational change.