Analysis of blm: a gene of unclear function involved in the biosynthesis of clavulanic acid in Streptomyces clavuligerus

Abstract

Streptomyces clavuligerus produces the medicinally important specialized metabolite clavulanic acid (CA), but detailed understanding of the roles played by many genes involved in CA biosynthesis is still lacking. These include the overlapping blm (orf12), orf13 and orf14 genes, disruptions of which cause defective CA production in S. clavuligerus. I investigated the potential polarity associated with disrupting blm in S. clavuligerus, along with the cellular localization of Blm. This was accomplished using complementation of prepared blm deletion mutants and western analysis of strains expressing epitope (FLAG) tagged copies of the protein. blm-like genes from other microorganisms containing sequenced CA gene clusters were examined for their ability to complement a S. clavuligerus blm in-frame mutant. Results demonstrate that the blm mutation is not associated with polar effects and that Blm is a cytoplasmic protein, suggesting that it could either play a regulatory or enzymatic role during CA production in S. clavuligerus.