Overexpression and puriWcation of scytovirin, a potent, novel anti-HIV protein from the cultured cyanobacterium Scytonema varium

Scytovirin (SVN) is a novel anti-human immunodeWciency virus (HIV) protein isolated from aqueous extracts of the cultured cyanobacterium Scytonema varium. The protein consists of a single 95-amino acid chain with signiWcant internal sequence duplication and 10 cysteines forming Wve intrachain disulWde bonds. A synthetic gene that encodes scytovirin was constructed, and expressed in Escherichia coli, with thioredoxin (TRX) fused to its N-terminius (TRX-SVN). Most of the expressed protein was in soluble form, which was puriWed by a polyhistidine tag aYnity puriWcation step. SVN was then cleaved from TRX with enterokinase and separated from the TRX partner by C18 reversed-phase HPLC. This production method has proven superior to earlier synthetic attempts and recombinant procedures using a standard expression system. The current system resulted in yields of 5–10 mg/L of puriWed SVN for structural studies and for preclinical development of SVN as a topical microbicide for HIV prophylaxis. Published by Elsevier Inc.