title: Discovery and characterization of an isopeptidase that linearizes lasso peptides.
abstract: Lasso peptides are a class of ribosomally derived natural products with diverse bioactivities. The characteristic threaded lasso structure in these peptides derives from an isopeptide bond attaching the N-terminus of the peptide to an acidic side chain. Here we describe the heterologous expression of a lasso peptide gene cluster encoding two lasso peptides, astexin-2 and astexin-3, and solve the solution structure of astexin-3. This cluster also encodes an enzyme annotated as a protease. We show that this enzyme, AtxE2, is a lasso peptide isopeptidase that specifically hydrolyzes astexins-2 and -3, converting them to linear peptides. Astexin-3 is highly thermostable and resists unthreading after extensive heat treatment. In contrast, astexin-2 unthreads upon heat treatment. AtxE2 has no activity toward unthreaded astexin-2, demonstrating that this isopeptidase must recognize a knotted structure in order to function. We also use this isopeptidase as a tool to study evolutionary relationships between lasso peptide gene clusters.
authors.affiliation: Department of Chemical and Biological Engineering, Princeton University, Princeton, New Jersey 08544, USA.
authors.forename: Mikhail O
authors.initials: MO
authors.lastname: Maksimov
authors.forename: A James
authors.initials: AJ
authors.lastname: Link
mesh_terms: D000595:Amino Acid Sequence; D019759:Carbon-Nitrogen Lyases / Q000235:genetics / Q000378:metabolism*; D042063:Caulobacteraceae / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D006868:Hydrolysis; D008958:Models, Molecular; D008969:Molecular Sequence Data; D005810:Multigene Family; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010802:Phylogeny
chemical_list: D010455:Peptides; D019759:Carbon-Nitrogen Lyases; C034300:isopeptidase