1H-n.m.r. investigation of the interaction between cytochrome c and cytochrome b5

Abstract

The interaction between eukaryotic cytochrome c and the tryptic fragment of bovine liver microsomal cytochrome b5 was studied by 1H-n.m.r. spectroscopy, and a procedure was developed that may be generally applicable to the study of macromolecular interactions by n.m.r. At pH6.3 (27 degrees C, I approx. 0.04) the two ferricytochromes were found to form a 1:1 complex with an association constant of approx. 10(3) M -1. The protein-protein-interaction region was found to encompass the region of the surface of horse cytochrome c that includes Ile-81, Phe-82, Ala-83 and Ile-85, and Lys-13 and Lys-72 of horse cytochrome c were suggested to be involved in two important intermolecular interactions. Me3Lys-72 of Candida krusei cytochrome c was shown to be involved in the interaction.