The rates of electron transfer from the tryptic fragment of bovine liver cytochrome b5 to FeIIINTA, FeIIIATP, CuIINTA, CuIIATP, and CuIIHis have been measured by anaerobic stopped-flow techniques. The rates of reduction of the Fe(III) complexes are independent of ionic strength, enhanced at low pH, and slightly inhibited by ZnIINTA. Saturation kinetics are observed with CuIINTA (kappa et = 0.05 s-1, K = 8.6 M-1), CuIIHis (kappa et = 0.2 s-1, K = 2.6 X 10(3) M-1), and CuIIATP (kappa et = 0.6 s-1, K = 4.5 X 10(3) M-1), thereby indicating that binding of Cu(II) to the protein occurs prior to electron transfer. 1H NMR resonances of the three surface histidines and some neighboring residues have been assigned by two-dimensional NMR techniques. NMR titration experiments show unequivocally that CuIINTA binds preferentially at a site near His-26 and Tyr-27.
