Albomycin (ABM), also known as grisein, is a sulfur-containing metabolite produced by Streptomyces griseus ATCC 700974. Genes predicted to be involved in the biosynthesis of ABM and ABM-like molecules are found in the genomes of other actinomycetes. ABM has potent antibacterial activity, and as a result, many attempts have been made to develop ABM into a drug since the last century. Although the productivity of S. griseus can be increased with random mutagenesis methods, understanding of Streptomyces sulfur amino acid (SAA) metabolism, which supplies a precursor for ABM biosynthesis, could lead to improved and stable production. We previously characterized the gene cluster (abm) in the genome-sequenced S. griseus strain and proposed that the sulfur atom of ABM is derived from either cysteine (Cys) or homocysteine (Hcy). The gene product, AbmD, appears to be an important link between primary and secondary sulfur metabolic pathways. Here, we show that propargylglycine or iron supplementation in growth media increased ABM production by significantly changing the relative concentrations of intracellular Cys and Hcy. An SAA metabolic network of S. griseus was constructed. Pathways toward increasing Hcy were shown to positively impact ABM production. The abmD gene and five genes that increased the Hcy/Cys ratio were assembled downstream of hrdBp promoter sequences and integrated into the chromosome for overexpression. The ABM titer of one engineered strain, SCAK3, in a chemically defined medium was consistently improved to levels ∼400% of the wild type. Finally, we analyzed the production and growth of SCAK3 in shake flasks for further process development.
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This work was supported by grants from U.S. National Institutes of Health (AI087849 to S.V.L.) and the National Basic Research Program of China (2011CBA00803 and 2012CB721101 to W.Z.).
Kulkarni, Aditya; Zeng, Yu; Zhou, Wei; Van Lanen, Steven; Zhang, Weiwen; and Chen, Shawn, "A Branch Point of Streptomyces Sulfur Amino Acid Metabolism Controls the Production of Albomycin" (2016). Pharmaceutical Sciences Faculty Publications. 67.
Supplemental File 1 - Comparison of key sulfur amino acid metabolism genes (Table S1), semi-log plot of zone of inhibition diameter versus amount of pure ABM (Fig. S1), HPLC quantification curve of pure ABM (Fig. S2), UV-Vis absorption spectra (Fig. S3), bioassay plates (Fig. S4), comparison of NBRC 13350 whole genome and ATCC 700974 draft genome (Fig. S5), PCR analysis of the construction of mutants (Fig. S6), HPLC-fluorescence detection (Fig. S7), and the proposed AbmD reaction that forms the -C-S-C- group of ABM (Fig. S8).