Mechanism of salinomycin overproduction in Streptomyces albus as revealed by comparative functional genomics

Abstract

The anticoccidial salinomycin is a polyketide produced by Streptomyces albus, and the high-yield strain BK 3-25 produces 18.0 g/L salinomycin under lab condition. In order to elucidate the overproduction mechanism, the genome of BK 3-25 was fully sequenced and compared with the wild-type DSM 41398. Strain BK 3-25 has a 75-kb large deletion, containing type-I polyketide gene cluster PKS-9, and 60 additional InDels and SNVs affecting 55 CDSs, including a 1-bp deletion in type-I PKS gene cluster PKS-6. Subsequently, individual or combined deletions of the 75-kb region and PKS-6 in the wild-type resulted in improved salinomycin yields from 2.60 to 5.20, 6.90, and 9.50 g/L (53% of BK 3-25), respectively, suggesting a redirected flux of polyketide precursors to salinomycin biosynthesis. Moreover, due to the much higher transcription of salinomycin biosynthetic genes (sln) in the high-yield BK 3-25 than in the wild-type, 13 putative regulatory genes among the 55 CDSs were individually inactivated and 7 were proved to be negatively involved in the transcription of sln genes. Combined deletions of two major negative regulatory genes SLNWT_3357 and SLNWT_7015 caused further improved transcription of sln genes as well as the yield, from 2.60 to 7.30 g/L (40% of BK 3-25). Therefore, the comparative genomics approach combined with functional experiments identified that the multiple deletions and mutations of competing gene clusters and negative regulatory genes are crucial for salinomycin overproduction, setting an example for rational titer improvement of other polyketide natural products.

Subtilisin-Involved Morphology Engineering for Improved Antibiotic Production in Actinomycetes

“…Whereas A. pretiosum ATCC 31280 undergoes mycelial fragmentation at early stage of cultivation and has less biomass accumulation, most Streptomyces strains have robust growth and form large dense pellets consisting of interconnected hyphae, which usually limit the transfer of nutrients and oxygen and consequent antibiotic production [ 12 ]. In an attempt to reduce to the size of pellets, the subtilisin-like gene APASM_4178 under the control of promoter kasOp* was introduced into salinomycin-producing Streptomyces albus BK 3-25 [ 50 ] and validamycin-producing Streptomyces hygroscopicus TL01 [ 51 ]. Interestingly, the overexpression of APASM_4178 in both strains indeed led to dispersed mycelia ( Figure 5 A,B).…”

Reconstitution of a mini‐gene cluster combined with ribosome engineering led to effective enhancement of salinomycin production in Streptomyces albus

“…So strategies towards regulatory system are of importance for those complex biosynthetic pathways. In terms of metabolic modulation‐related approaches, diligent efforts have been made to improve the production of antibiotics via various techniques (Lu et al ., 2016 ; Tanaka et al ., 2017 ; Zhang et al ., 2017a , 2017b , 2019 , 2017a , 2017b , 2019 ). Also, the whole sal gene cluster was cloned and heterologously expressed in S. coelicolor A3(2) (Yin et al ., 2015 ).…”

Positive regulation of the MarR-type regulator slnO and improvement of salinomycin production by Streptomyces albus by multiple transcriptional regulation

“…Salinomycin, a polyether ionophore antibiotic produced by S. albus, is widely used as anticoccidial drugs and growth promoters in animal husbandry. It has been used for 30 years as an anticoccidial drug and to improve nutrient absorption and feed efficiency in ruminants and swine D r a f t (Zhang et al 2017). It has the ability to selectively chelate metal ions in a hydrophobic matrix, and then transport the ions to the cell membrane, which destroys the physiological ion gradients of the cells and eventually results in cell death (Zhu et al 2017).…”