HIGHLY PURIFIED BIOTIN SYNTHASE CAN TRANSFORM DETHIOBIOTIN INTO BIOTIN IN THE ABSENCE OF ANY OTHER PROTEIN, IN THE PRESENCE OF PHOTOREDUCEDDEAZAFLAVIN

Biotin synthase from Bacillus Sphaericus has been purified to homogene ity from a recombinant strain. The UV-visible spectrum of the pure pro tein reveals the presence of a [2Fe-2S] cluster. The enzyme is active in the conversion of dethiobiotin to biotin in vitro, in the presence of NADPH, AdoMet and additional unidentified components from the I cru de extract of B. sphaericus wild type. We have also found that photore duced deazaflavin can substitute for the crude extract and NADPH. In t his system, biotin synthase is capable of transforming dethiobiotin in to biotin in the absence of any other protein but at a substoichiometr ic level. When this assay was conducted in the presence of [S-35]cyste ine, no S-35 was incorporated into biotin, contrary to what happens in the presence of the crude extract. (C) 1995 Academic Press. Inc.