Genome shuffling and ribosome engineering of Streptomyces virginiae for improved virginiamycin production

Abstract

The production of virginiamycin (VGM) from Streptomyces virginiae was improved by genome shuffling and ribosome engineering companied with a high-throughput screening method integrating deep-well cultivation and the cylinder-plate detecting. First, a novel high-throughput method was developed to rapidly screen large numbers of VGM-producing mutants. Then, the starting population of genome shuffling was obtained through ultraviolet (UV) and microwave mutagenesis, and four mutants with higher productivity of VGM were selected for genome shuffling. Next, the parent protoplasts were inactivated by UV and heat when a fusant probability was about 98%. Streptomycin resistance was used as an evolutionary pressure to extend positive effects on VGM synthesis. Finally, after five rounds of genome shuffling, a genetically stable strain G5-103 was obtained and characterized to be able to yield 251 mg/L VGM, which was 3.1- and 11.6-fold higher than that of the mutant strain UV 1150 and the wild-type strain, respectively.

Combination of atmospheric and room temperature plasma (ARTP) mutagenesis, genome shuffling and dimethyl sulfoxide (DMSO) feeding to improve FK506 production in Streptomyces tsukubaensis

“…GS has become a promising technique to enhance secondary metabolite production in a variety of microorganisms since it was discovered (Lv et al, 2013;Zhang et al, 2013;Zhao et al, 2012). As an artificial evolution method, it addresses the limitations of classical strain improvement, and can greatly improve the frequency of desired mutation in microbial cells (Tong et al, 2018). Meanwhile, the novel ARTP mutagenesis technology provides a mutational library containing multiple positive mutants for GS.…”

The Application of Ribosome Engineering to Natural Product Discovery and Yield Improvement in Streptomyces

“…Genome shuffling is an approach involving recursive genomic recombination within a population of phenotypically selected bacteria to generate new strain libraries, which may result in pronounced improvement in the screened phenotype. The combination of genome shuffling and ribosome engineering has been widely used in antibiotic development, including avilamycin ( 5 ) [36], daptomycin ( 7 ) [39], nosiheptide ( 1 ) [43], and virginiamycin ( 2 ) [52]. For example, avilamycin ( 5 ), a feed industry antimicrobial agent approved by the European Union, is used to inhibit the growth of multidrug resistant Gram-positive bacteria.…”

“…In addition, ribosome engineering can also activate silent BGCs and discover new active compounds in other bacterial species or fungi. Ochi et al [52] first introduced a rifamycin-induced rpoB mutation into the B. subtilis strain, which led to the production of a new aminosaccharide antibiotic 3,3’-neotrehalosadiamine ( 31 ), for which production is silent in the wild-type strain. In a metabolomic phenotype screening of str or rif mutants in a rare actinomycete Nocardiopsis sp.…”

Improvement of an Industrial Crystallization Process: The Production of Virginiamycin

“…The bactericidal action is related to the specific and irreversible binding in specific positions of the 50S subunit of ribosomes, which inhibits protein synthesis and causes a reduction in growth (bacteriostasis) or the death of bacteria (antibacterial activity). This unique synergy results in a lack of any significant resistance of microorganisms to virginiamycin (VM), even after long-term use. − …”