P57
Activation of the silent actinorhodin gene cluster of Streptomyces lividans by DNA fragments from Saccharopolyspora erythraea
Sunday, January 11, 2015
California Ballroom C and Santa Fe Room
The discovery of silent antibiotic biosynthetic gene clusters in actinomycetes and the development of new methods for their activation has renewed interest in natural product drug discovery. In this study we transferred genes from the erythromycin-producing organism Saccharopolyspora erythraea into Streptomyces lividans and looked for genes that would activate the silent actinorhodin pathway. A library of 2 - 5 kb DNA fragments were transformed into S. lividans on a high copy plasmid and the transformants were visually screened for blue pigment production. Of 50,000 transformants that were screened, four plasmids were found to strongly activate blue pigment production. The strongest activation was obtained by over-expression of a single gene, SACE_1669, a putative membrane peptidase. SACE_1669 encodes a predicted protein of 28.9 kDa. The N-terminal sequence has a signal peptide followed by an 81 amino acid domain similar to lysostaphin and other bacteriocin-like peptidases. A 37 amino acid C-terminal domain was 46% identical to a domain from sapA, a developmentally regulated spore coat protein from Streptomyces coelicolor. Activation of actinorhodin production by SACE_1669 was starch dependent. Strong activation of actinorhodin was also observed by over expression of SACE_0905, 4-hydroxyphenylpyruvate dioxygenase, an enzyme of tyrosine catabolism widely found in nature. The other two plasmids appeared to activate actinorhodin due to an increased copy number of intergenic DNA sequences. The mechanism of activation for these genes and DNA fragments has not been elucidated but the results indicate that there is a high degree of diversity in the types of activation mechanisms that are possible.