Heterologous Expression Of Type Ⅲ Lanthipeptide From Lentzea Jiangxiensis And Streptomyces Davaonensis

As a large family of ribosomally synthesized and post-translationally modified peptides(Ri PPs),lanthipeptides have shown a variety of biological activities such as antibacterial,antiviral,antiallodynic and other biological activities,and their genome mining and biosynthetic pathways have been continuously explored and studied.Bacteria are currently the only source of lanthipeptides,their genomes contain a large number of lanthipeptide biosynthetic gene clusters,and show a variety of evolutionary sources,the research on lanthipeptides is constantly advancing,and the discovery of heterozygous gene clusters provides further research space for new lanthipeptides.As the mainstream means to study biosynthetic gene clusters and obtain their coding natural products,heterologous expression allows us to quickly explore related gene clusters with a clear metabolic background and convenient gene editing.We targeted type III lanthipeptide,looked for relevant biosynthetic gene clusters that may have novel modifications through genome mining,and carried out research through heterologous expression.First of all,we targeted lipolanthines,which had been found in recent years,which had been heterozygosity for fatty acid chains and lanthipeptides,and found a gene cluster encoding polyketone synthase and type III lanthipeptide heterozygosity in Lentzea jiangxiensis through bioinformatics analysis.Through continuous optimization of experimental methods,heterologous expression in Streptomyces successfully activated the silencing gene cluster,a new type III lanthipeptide product was discovered,and four products of the same series were found after fermentation and separation,adding new members to the lanthipeptide family,and also discovering a new S8 family serine protease,which may participate in the lead peptide resection of type III lanthipeptide,enriching the few type III lanthipeptide protease family.However,at present,no type III lanthipeptide products with fatty acylation have been found,and work will continue around this part.In order to explore the biosynthesis of glycosylated lanthipeptide,through bioinformatics analysis of glycosyltransferases,we found a type III lanthipeptide biosynthetic gene cluster encoding glycosyltransferase in Streptomyces davaonensis,which was expressed heterologous in Streptomyces and Escherichia coli,by adding promoters,co-expressing with Chaperone Plasmid,etc.The precursor peptide was successfully expressed in E.coli,and the dehydrated precursor peptide product was successfully discovered by co-expression with dehydrated cyclase Str KC and glycosyltransferase Str G,which laid a preliminary foundation for the further development of glycosylated type III lanthipeptide.In this thesis,two type III lanthipeptide biosynthetic gene clusters were studied by heterologous expression,and a new type III lanthipeptide product and corresponding protease were found,adding members of type III lanthipeptide,and the new type of protease may participated in the biosynthesis of type III lanthpeptide,which enriched the biosynthetic diversity of this compound.The reaction sequence of dehydration cyclization and glycosylation needs to be further studied,which puts forward new possibilities for the biosynthesis of type III lanthipeptide,and the biosynthetic mechanism of more types of lanthipeptide remains to be explored.At the same time,a variety of ways to optimize the activation of gene clusters in the experiment also provide a reference for heterologous expression to generate lanthipeptide.