title: Characterisation of the peptaibiome of the biocontrol fungus Trichoderma atroviride by liquid chromatography/tandem mass spectrometry.
abstract: The present study describes the liquid chromatography/tandem mass spectrometry (LC/MS/MS)-based screening and characterisation of linear antibiotic alpha-aminoisobutyric acid (Aib)-containing non-ribosomal peptides (NRP) in culture samples of the filamentous fungus Trichoderma atroviride ATCC 74058. Fungal culture filtrates were enriched by solid-phase extraction (SPE) and separated by reversed-phase high-performance liquid chromatography (HPLC), prior to mass spectrometric (MS) and tandem mass spectrometric (MS/MS) analysis on a triple quadrupole-linear ion trap tandem mass spectrometer. A workflow consisting of two alternative screening strategies was applied to search for NRP. Various MS full scan and MS/MS measurement modes led to the identification of 16 trichorzianines and diagnostic in-source fragment ions of another four trichorzianines. Furthermore, we detected 15 novel Aib-containing peptides with putative molecular weights ranging from 951.7 to 1043.7 g/mol (monoisotopic masses), composed of up to 9 amino acids. While the amino acid sequences of the novel peptaibiotics showed typical microheterogeneity and consisted of the amino acids Leu/Ile, Aib, Ser, Val/Iva, Gly, Ac-Aib, Tyr and Phe, the mass increments at the C-termini of the peptides were not assignable to any residues described in the literature. The amino acid sequences were confirmed and structure proposals made for both molecule termini by high-resolution MS and MS/MS analysis. We propose the group name 'trichoatrokontins' for the newly identified peptaibiotics. As no other peptaibiotics were found in the culture samples, the peptaibiome of the investigated strain of T. atroviride consists of at least 20 trichorzianines and 15 trichoatrokontins.
authors.affiliation: Department for Agrobiotechnology , University of Natural Resources and Applied Life Sciences, Vienna, Konrad Lorenz Str. 20, A-3430 Tulln, Austria.
authors.forename: Norbert
authors.initials: N
authors.lastname: Stoppacher
authors.forename: Susanne
authors.initials: S
authors.lastname: Zeilinger
authors.forename: Markus
authors.initials: M
authors.lastname: Omann
authors.forename: Paul-Gerhard
authors.initials: PG
authors.lastname: Lassahn
authors.forename: Andreas
authors.initials: A
authors.lastname: Roitinger
authors.forename: Rudolf
authors.initials: R
authors.lastname: Krska
authors.forename: Rainer
authors.initials: R
authors.lastname: Schuhmacher
mesh_terms: D000595:Amino Acid Sequence; D000621:Aminoisobutyric Acids / Q000737:chemistry; D002853:Chromatography, Liquid / Q000295:instrumentation / Q000379:methods*; D054848:Peptaibols / Q000032:analysis* / Q000737:chemistry; D010455:Peptides / Q000737:chemistry; D010572:Pest Control, Biological; D012015:Reference Standards; D020539:Sequence Analysis, Protein; D052616:Solid Phase Extraction; D053719:Tandem Mass Spectrometry / Q000379:methods*; D013997:Time Factors; D014242:Trichoderma / Q000737:chemistry*
chemical_list: D000621:Aminoisobutyric Acids; D054848:Peptaibols; D010455:Peptides; C100049:2-aminoisobutyric acid