Increased vancomycin production by overexpression of MbtH-like protein in Amycolatopsis orientalis KFCC10990P.

UNLABELLED: Recently, many in vitro studies have reported that MbtH-like proteins are very necessary for the adenylation of amino acid by adenylating enzymes present in the biosynthetic machineries of nonribosomal peptides (NRPs). However, in vivo studies on mbtH-like genes are somewhat controversial since their mutants still produce the target compounds. Here, we report unambiguous evidence of the crucial role of MbtH-like protein in the biosynthesis of NRP based on in vivo study of vancomycin producer, Amycolatopsis orientalis. Deletion of mbtH-like gene (vcm11) in the vancomycin biosynthetic gene cluster completely abolished production of vancomycin and its complementation strain showed almost full recovery of vancomycin production. As a result, we propose that the mbtH-like gene is a good genetic engineering target to increase the yield of NRP, as verified by increased vancomycin production (by 60 and 80%) upon overexpression of cognate (Vcm11) as well as noncognate (CloY) MbtH-like proteins. SIGNIFICANCE AND IMPACT OF THE STUDY: Elucidation and application of biosynthetic machineries of bioactive compounds containing amino acids such as antibiotics, immunosuppressants and siderphores etc. are significant for the production and development of drugs. Here, we observed an apparent increase in the yield of vancomycin, a type of NRP, upon overexpression of MbtH-like protein in Amycolatopsis orientalis. Our result is the first example of increased NRP(s) yield following overexpression of mbtH-like genes to develop the strain for economic production and elucidate the role of MbtH-like protein in vivo for combinatorial biosynthesis.