Heterologous expression of novel class Ⅰ lanthipeptide in Streptomyces coelicolor A3(2)

Abstract:

[Objective] Class Ⅰ lanthipeptides usually have a wide range of biological activities,with unique antibacterial mechanism and little drug resistance,which leads to the promising clinical applications.This work intended to explore two novel class Ⅰ lanthipeptides in Streptomyces coelicolor A3(2).[Methods] First,antiSMASH was used to analyze the S.coelicolor A3(2) genome sequence and mine lanthipeptide biosynthetic gene clusters.BLAST was employed to annotate gene function and select genes that might be involved in the biosynthesis process.Then,we constructed plasmids via DNA assembly and transferred them into Streptomyces chassis cells for heterologous expression.Finally,high performance liquid chromatography (HPLC),mass spectrometry (MS) and bioactivity assay were performed to detect the fermentation products.[Results] We reconstituted cluster 3(8.9 kb) and cluster 24(9.0 kb) of S.coelicolor A3(2) by adding promoter elements,and pYES-ColE1-SCO-cluster3 and pYES-ColE1-SCO-cluster24 were obtained.pYES-ColE1-SCO-cluster3 was successfully expressed in S.coelicolor M1152 and Streptomyces sp.A14 separately,and the potential target compound coelin 3 was obtained.pYES-ColE1-SCO-cluster24 was heterologously expressed in Streptomyces sp.ZM13,and the potential target compound coelin 24 was obtained.Coelin 3 exerted inhibitory effects on Bacillus subtilis 168 and Escherichia coli ATCC 25922,with the inhibition zone reaching 28 mm.[Conclusion] This study realized the expression of coelin 3 and coelin 24 and conducted the bioactivity assay by promoter activation and heterologous expression,which laid a foundation for subsequent structural and mechanism analysis of the novel class Ⅰ lanthipeptides.