Putative F-G loop is involved in association with the membrane in P450scc (P450 11A1).

Cytochrome P450scc (P450 11A1) catalyzes the conversion of cholesterol to pregnenolone, the first step in overall steroid hormone biosynthesis in steroidogenic tissues. On the basis of alignment with structurally characterized cytochromes P450 (P450s), we have identified the putative F-G loop of P450 11A1 and mutated amino acid residues in this region. Wild type and mutant P450s 11A1 were expressed in E. coli and compared with respect to subcellular distribution and turnover number. About 30% of the wild type P450 was found in the bacterial cytosol indicating loose association of the enzyme with the membrane. The N210S/V211M and L218R/F219Y replacements increased the fraction of P450 in the bacterial cytosol 1.5-1.7-fold and the latter mutant also showed an almost two-fold increase of the turnover number. These data indicate that the putative F-G loop is the site of attachment to the membrane in P450 11A1 and changes in the enzyme-membrane interactions may affect the rate of catalysis.