The Biosynthsis Of Giant Lasso Peptide Derived From Enterobacterium

Among the many microbial natural drugs,Ribosomally Synthesized and Posttranslationally Modified Peptides(RiPPs)are important ones.RiPPs is a kind of natural product which is synthesized by ribosomes and obtained through translation and modification.It has a very wide variety of structure and biodiversity.The good antibacterial activity and unique mechanism of these peptides have attracted wide attention,making these compounds a hot trend in the study of anti-infective drugs.In many ribosomal peptides,lasso polypeptide have relatively simple modifications compared to other chemicals.However,its unique "lasso" structure has attracted a lot of attention.This unique tertiary structure provides unique stability and antibacterial activity to the lasso peptides.In this study,enterobacterium was used as the object of study to explore novel lasso peptide antibiotics.First of all,through the method of genomic excavations,we found a lasso polypeptide derived from enterobacteria.Through the structure prediction,this kind of polypeptide may be the largest lasso polypeptide in the world,consisting of up to 24 amino acids.The gene cluster of this giant lasso polypeptide was cloned into pET41 and expressed in the host of escherichia coli.After mass spectrometry analysis,the recombinant bacteria were successfully obtained,and the optimal fermentation conditions were determined.After 5 hours of culture at 37? and three days of induction,the yield of polypeptides was highest.By modifying the gene cluster of giant lasso polypeptide,we inserted the transporter fragment into the gene cluster and inserted the ribosome binding site.The modified gene cluster was cloned into pET41 and expressed in the host of escherichia coli,and the results showed that the yield of the polypeptide compound was further improved about 6 times by liquid phase detection.A large number of recombinant strains were fermented and purified by liquid phase purification,and it has successfully purified the 3mg lasso polypeptides.