THE ASSOCIATION RATE-CONSTANT FOR HEME-BINDING TO GLOBIN IS INDEPENDENT OF PROTEIN-STRUCTURE

Rate constants for CO-heme binding to 35 different recombinant apomyog lobins and several other apoproteins were measured in an effort to und erstand the factors governing heme affinity and the velocity of the as sociation reaction. Surprisingly, the rate constant for the binding of monomeric heme is approximate to 1 x 10(8) M(-1) s(-1) regardless of the structure or overall affinity of the apoprotein for iron-porphyrin . Major differences between the proteins are reflected primarily in th e rates of dissociation of the prosthetic group. Slow phases observed in the reaction of CO heme with excess apomyoglobin result from format ion of nonspecific heme-protein complexes which must dissociate before heme can bind specifically in the heme pocket. Once the specific heme -globin complex is formed, the heme pocket rapidly collapses around th e porphyrin, simultaneously forming the bond between the proximal His( 93) and the heme iron atom. The overall affinity of sperm whale apomyo globin for hemin is similar to 1 x 10(14) M(-1). Nonspecific hydrophob ic interactions between the porphyrin and the apolar heme cavity accou nt for a factor of 10(5)-10(4). Covalent bond formation between Fe3+ a nd His(93)(F8) provides an additional factor of 10(3)-10(4). Specific interactions with conserved amino acids in the heme pocket contribute the final factor of 10(3)-10(4).