Screening Biosynthetic Peptide Libraries for Antimicrobial Activity

摘要

We have recently demonstrated the feasibility of utilizing the ligase activity of inteins for the in vivo backbone cyclization of peptidic chains. This procedure-called SICLOPPS for Split Intein Circular Ligation Of Peptides and ProteinS -provides a biosynthetic pathway for peptides that are metabolically stable and can be produced with spatial and temporal control. To screen for bacteriotoxic peptides, a SICLOPPS library was introduced into an Escherichia coli population, such that each bacterium encodes a different peptide sequence. SICLOPPS library over-expression afforded six distinct bacteriostatic peptides that reduce cell growth. One of these peptides (LN05) also caused cell aggregation. An E. coli genomic library was introduced into cells encoding LN05. Co-expression of the genomic library and LN05 peptide rescues growth only in cells expressing genomic fragments able to counteract peptide toxicity. Genomic library and LN05 co-expression resulted in enrichment of a single genomic construct, a fragment of the NarZ gene. NarZ is part j of a nitrate reductase complex and has a role in tuberculosis persistence.