Environment of the Heme in Myoglobins. NMRD and EPR Spectroscopy of Val68X (X = Asn, Asp, and Glu) Mutants of Human Myoglobin
Analyses of the temperature dependences and shapes of nuclear magnetic relaxation dispersion (NMRD) signals of site-directed mutants of human myoglobin indicate that a water molecule is bound to the sixth coordination site of the ferric heme in proteins in which the valine at position 68 is changed to either aspartate (Val68Asp) or asparagine (Val68Asn). Both NMRD data and electron paramagetic resonance (EPR) spectra show that carboxylate is axially ligated to Fe(III) in the Va168Glu mutant. The EPR spectra of the Va168Asp and Va168Asn derivatives show much smaller rhombic splittings than the spectrum of the Va168Glu protein.
© 1994 American Chemical Society. Received August 23, 1993. We thank Seymour Koenig and Rodney Brown of the IBM Thomas J. Watson Research Center for allowing us access to their facilities and for assistance with many of the experiments. During the course of the work, we enjoyed several discussions with Claudio Luchinat, Lucia Banci, and Paul Smith. Research at Caltech was supported by the National Science Foundation, the National Institutes of Health, and the Arnold and Mabel Beckman Foundation.