PROPERTIES OF HUMAN HEMOGLOBINS WITH INCREASED POLARITY IN THE ALPHA-HEME OR BETA-HEME POCKET - CARBONMONOXY DERIVATIVES

The spectroscopic, conformational, and functional properties of mutant carbonmonoxy hemoglobins in which either the beta-globin Val(67)(Ell) or the alpha-globin Val(62)(Ell) is replaced by threonine have been i nvestigated. The thermal evolution of the Soret absorption band and th e stretching frequency of the bound CO were used to probe the stereody namic propel-ties of the heme pocket. The functional properties mere i nvestigated by kinetic measurements. The spectroscopic and functional data were related to the conformational properties through molecular a nalysis. The effects of this nonpolar-to-polar isosteric mutation are: (i) increase of heme pocket anharmonic motions, (ii) stabilization of the A(o) conformer in the IR, spectrum, (iii) increased CO dissociati on rates. The spectroscopic data indicate that for the carbonmonoxy de rivatives, the Val --> Thr mutation has a larger conformational effect on the beta-subunits than on the alpha-subunits. This is at variance with the deoxy derivatives where the conformational modification was l arger in the heme pocket of the alpha-subunit (Cupane, A., Leone, Ri., Militello, V,, Friedman, R. K., Koley, A. P,, Vasquez, G. P,, Briniga r, W, S., Kara; vitis, M., and Fronticelli, C. (1997) J. Biol. Chern. 272, 26271-26278), These effects are attributed to a different electro static interaction between O-gamma of Thr(E11) and the bound CO molecu le. Molecular analysis indicates a more favorable interaction of the b ound CO with Thr OY in the beta-subunit heme pocket.