Cloning, characterization and expression of the gene encoding cytochrome P-450sca-in2 from Streptomyces carbophilus involved in production of pravastatin, a specific HMG-CoA reductase inhibitor

Abstract

Pravastatin, a drug for treating hypercholesterolemia, is produced by hydroxylation of ML-236B-Na in Streptomyces carbophilus (Sc) catalyzed by the cytochrome P-450sca (CytP-450sca) monooxygenase system. The gene (cytP-450sca-2) encoding CytP-450sca-2 was cloned from Sc. The gene had an open reading frame of 1233 bp, encoding a 410-amino-acid protein. The partial sequencing of the purified CytP-450sca-2 revealed that the N-terminal Met had been removed. CytP-450sca-2 contained the heme-binding HR2 region characteristic of all CytP-450, as well as the putative oxygen-binding site proposed in CytP-450cam from Pseudomonas putida. ML-236B.Na enhanced transcription of cytP-450sca-2, suggesting that substrate induction in Sc is transcriptionally regulated. S. lividans (Sl) transfected with cytP-450sca-2 converted ML-236B.Na to pravastatin, indicating the cloned gene to be functional in Sl.