Abstract

The initial product upon reacting apomyoglobin with hemin is the holoprotein with the hemin 1:1 rotationally disordered about the a -g -meso axis. Detn. of the rate of equilibration of the disordered state as a function of pH and hemin 2,4-substituents has shown that the rate is highly pH-dependent with a min. near neutral pH and that the relative rates at any pH depend critically on the 2,4-substituent. The slowest rate was obsd. for native hemin (2,4-vinyl), the fastest for deuterohemin (2,4-H), and intermediate for mesohemin (2,4-Et). Competitive reconstitution for 2 different hemins revealed that the forward rate was independent of the 2,4-substituent, and hemin replacement reactions yielded the relative binding consts. for the various hemes. These 2 sets of data yielded the relative dissocn. rates for the various hemes which were shown to be semiquant., the same as the heme reorientation rates, indicating that disruption of the heme cavity was the rate-detg. step in the reorientation. However, competition expts. between heme reorientation and heme replacements revealed that reorientation occurred faster than replacement, indictating that the reorientation occurs by an intramol. mechanism, i.e., without leaving a protein cage.