Phosphoprotein affinity purification identifies proteins involved in S-adenosyl-L-methionine-induced enhancement of antibiotic production in Streptomyces coelicolor

Abstract

Streptomycetes are the major natural source of clinical antibiotics. The enhanced secondary metabolite production of many streptomycetes by S-adenosylmethionine (SAM) in previous studies suggested the existence of a common SAM regulatory effect. We screened nine proteins using the phosphoprotein purification column from Streptomyces coelicolor. Among them, genes (SCO5477, SCO5113, SCO4647, SCO4885 and SCO1793) for five proteins were disrupted by insertion mutation. The undecylprodigiosin and actinorhodin productions were changed in all mutations. The SAM-induced enhancement of actinorhodin production was abolished by all mutations except SCO4885 mutation, which reduced the production of actinorhodin and undecylprodigiosin with SAM treatment. This study demonstrates that phosphoprotein affinity purification can be used as a screening method to identify the proteins involved SAM signaling. Keywords: actinorhodin; protein phosphorylation; S-adenosylmethionine; Streptomyces coelicolor; undecylprodigiosin INTRODUCTIONStreptomyces is the major genus that produces useful antibiotics of natural origin. Secondary metabolite production in Streptomyces is usually strain specific; thus, it is difficult to develop a universal method to promote the production of secondary metabolites in most Streptomyces strains. 1,2 We observed that increased concentration of S-adenosylmethionine (SAM) by overexpression of SAM synthetase gene stimulated actinorhodin production in Streptomyces lividans, 3,4 and exogenous SAM treatment in culture media enhanced streptomycin production in S. griseus, 5 as well as other secondary metabolite production in many Streptomyces strains. 6 On the basis of these results, we sought to identify SAM-response proteins, which might be targets, to improve secondary metabolite production in many Streptomyces spp.SAM-enhanced secondary metabolite production in all tested actinomycetes suggested the existence of a common signal transduction pathway. To identify cellular proteins that regulate the SAM response, we hypothesized that changes in protein phosphorylation constitute part of the signal transduction of SAM responses in S. coelicolor, a model strain for actinomycetes. As a new approach to

Quantitative Proteome and Phosphoproteome Analyses of Streptomyces coelicolor Reveal Proteins and Phosphoproteins Modulating Differentiation and Secondary Metabolism

“…3): OhkA, a regulator of secondary metabolism and morphological differentiation (59); RarA-C “restoration of aerial mycelium formation” proteins (60); SarA, a protein involved in the regulation of sporulation and secondary metabolism (61); AtrA, an activator of the actinorhodin biosynthetic gene expressions (62); DevA, a GntR-Like transcriptional regulator controlling aerial mycelium development (63); NsdA, a BldD target regulating morphological differentiation and secondary metabolism (64); and ScbR, a γ-butyrolactone binding protein regulating morphological differentiation and secondary metabolism (65). By contrast, some regulatory proteins were down-regulated at the MII stages: CspB, a repressor of secondary metabolism (66) and NusG, a transcription antiterminator repressing undecylprodigiosin production (67).…”

Diverse control of metabolism and other cellular processes in Streptomyces coelicolor by the PhoP transcription factor: genome-wide identification of in vivo targets

“…Both are implicated in antibiotic production and development and both are induced by supplementation with S-adenosylmethionine (SAM) (e.g. (46)). It is also notable that metK , encoding SAM synthetase, is substantially up-regulated in a phoP null mutant (Supplementary Table S4).…”

Comparative Proteomic Analysis of Streptomyces lividans Wild-Type andppkMutant Strains Reveals the Importance of Storage Lipids for Antibiotic Biosynthesis

“…SCO5113 is thought to be involved in morphological differentiation by allowing the transport of the morphogenic oligopeptide Bld261 (40,41), whereas the ABC transporters, including SCO5477, were reported to play a role in the upregulation of actinorhodin production by S-adenosylmethionine (42,43). These transport systems are thought to be involved in triggering antibiotic production.…”