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Lasso Peptide Biosynthetic Protein LarB1 Binds Both Leader and Core Peptide Regions of the Precursor Protein LarA

Author(s): Cheung, Wai Ling; Chen, Maria Y.; Maksimov, Mikhail O.; Link, A. James

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Abstract: Lasso peptides are a member of the superclass of ribosomally synthesized and posttranslationally modified peptides (RiPPs). Like all RiPPs, lasso peptides are derived from a gene-encoded precursor protein. The biosynthesis of lasso peptides requires two enzymatic activities: proteolytic cleavage between the leader peptide and the core peptide in the precursor protein, accomplished by the B enzymes, and ATP-dependent isopeptide bond formation, accomplished by the C enzymes. In a subset of lasso peptide biosynthetic gene clusters from Gram-positive organisms, the B enzyme is split between two proteins. One such gene cluster is found in the organism Rhodococcus jostii, which produces the antimicrobial lasso peptide lariatin. The B enzyme in R. jostii is split between two open reading frames, larB1 and larB2, both of which are required for lariatin biosynthesis. While the cysteine catalytic triad is found within the LarB2 protein, LarB1 is a PqqD homologue expected to bind to the lariatin precursor LarA based on its structural homology to other RiPP leader peptide binding domains. We show that LarB1 binds to the leader peptide of the lariatin precursor protein LarA with a sub-micromolar affinity. We used photocrosslinking with the noncanonical amino acid p-azidophenylalanine and mass spectrometry to map the interaction of LarA and LarB1. This analysis shows that the LarA leader peptide interacts with a conserved motif within LarB1 and, unexpectedly, the core peptide of LarA also binds to LarB1 in several positions. A Rosetta model built from distance restraints from the photocrosslinking experiments shows that the scissile bond between the leader peptide and core peptide in LarA is in a solvent-exposed loop.
Publication Date: 26-Oct-2016
Electronic Publication Date: 29-Sep-2016
Citation: Cheung, Wai Ling, Chen, Maria Y, Maksimov, Mikhail O, Link, A James. (2016). Lasso Peptide Biosynthetic Protein LarB1 Binds Both Leader and Core Peptide Regions of the Precursor Protein LarA. ACS Central Science, 2 (10), 702 - 709. doi:10.1021/acscentsci.6b00184
DOI: doi:10.1021/acscentsci.6b00184
ISSN: 2374-7943
EISSN: 2374-7951
Pages: 702 - 709
Type of Material: Journal Article
Journal/Proceeding Title: ACS Central Science
Version: Final published version. This is an open access article.

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