11-BETA-HYDROXYLASE ACTIVITY IN RECOMBINANT YEAST MITOCHONDRIA - IN-VIVO CONVERSION OF 11-DEOXYCORTISOL TO HYDROCORTISONE

In mammals, the final 11 beta-hydroxylation step of the hydrocortisone biosynthesis pathway is performed by a mitochondrial enzyme, namely c ytochrome P-450(11 beta), together with the electron carriers adrenodo xin and NADPH adrenodoxin oxidoreductase. Successful production of a f unctional steroid 11 beta-hydroxylase activity was obtained in recombi nant yeast in vivo. This conversion was achieved by coexpression of a mitochondrially targeted adrenodoxin and a modified bovine P-450(11 be ta) whose natural prese quence was replaced by a yeast presequence, to gether with an unexpected yeast endogenous NADPH-adrenodoxin-reductase -like activity. Adrenodoxin and P-450(11 beta) behave as a mitochondri al matrix and membrane protein, respectively. Saccharomyces cerevisiae apparently produces a mitochondrial protein which is capable of trans ferring electrons to bovine adrenodoxin, which in turn transfers the e lectrons to P-450(11 beta). The endogenous adrenodoxin oxidoreductase gains electrons specifically from NADPH. The notion that a yeast micro somal NADPH P-450 oxidoreductase can transfer electrons to mammalian m icrosomal P-450s can be extended to mitochondria, where an NADPH adren odoxin oxidoreductase protein transfers electrons to adrenodoxin and r enders a mitochondrial mammalian P-450 functional in vivo. The physiol ogical function of this yeast NADPH adrenodoxin oxidoreductase activit y is not known.