Occurrence and characterization of peptaibols from Trichoderma citrinoviride, an endophytic fungus of cork oak, using electrospray ionization quadrupole time-of-flight mass spectrometry.
A cork oak endophytic strain of Trichoderma citrinoviride, previously selected for its antagonistic potential against various fungal pathogens involved in oak decline, was screened for the production of bioactive secondary metabolites. From liquid culture a mixture of polypeptide antibiotics (peptaibols) belonging to the paracelsin family was isolated and characterized. This peptide mixture was purified by column chromatography and preparative TLC on silica gel, and separated by analytical HPLC. It was analysed by MALDI-TOF MS and nano-ESI-QTOF MS. Tandem mass experiments were performed to determine the amino acid sequences based on the fragmentation pattern of selected parent ions. The mixture comprised 20-residue peptides with C-terminal phenylalaninol and N-terminal acetylation. Twenty-eight amino acid sequences were identified, and amino acid exchanges were located in positions 6, 9, 12 and 17. Among them, seven sequences are new as compared to those reported in the database specifically for peptaibols and in the literature. In addition, we obtained experimental evidence suggesting the existence of non-covalent dimeric forms (homo- and hetero-) of the various peptaibol species. The peptide mixture showed strong antifungal activity toward seven important forest tree pathogens, and it was highly toxic in an Artemia salina (brine shrimp) bioassay. These results emphasize the cryptic role of endophytic fungi as a source of novel bioactive natural products and biocontrol agents.
Maddau
Lucia
L
Dipartimento di Protezione delle Piante, Sezione di Patologia Vegetale, Università di Sassari, Via E. De Nicola 9, 07100 Sassari, Italy.
Cabras
Annalisa
A
Dipartimento di Protezione delle Piante, Sezione di Patologia Vegetale, Università di Sassari, Via E. De Nicola 9, 07100 Sassari, Italy.
Franceschini
Antonio
A
Dipartimento di Protezione delle Piante, Sezione di Patologia Vegetale, Università di Sassari, Via E. De Nicola 9, 07100 Sassari, Italy.
Linaldeddu
Benedetto T
BT
Dipartimento di Protezione delle Piante, Sezione di Patologia Vegetale, Università di Sassari, Via E. De Nicola 9, 07100 Sassari, Italy.
Crobu
Salvatore
S
Porto Conte Ricerche Srl, Tramariglio, Alghero, Italy.
Roggio
Tonina
T
Porto Conte Ricerche Srl, Tramariglio, Alghero, Italy.
Pagnozzi
Daniela
D
Porto Conte Ricerche Srl, Tramariglio, Alghero, Italy.
D000818:Animals; D000935:Antifungal Agents / Q000032:analysis* / Q000302:isolation & purification* / Q000494:pharmacology / Q000633:toxicity; D001156:Artemia / Q000187:drug effects; D002851:Chromatography, High Pressure Liquid; D002853:Chromatography, Liquid; D002855:Chromatography, Thin Layer; D013058:Mass Spectrometry / Q000379:methods; D054848:Peptaibols / Q000032:analysis* / Q000302:isolation & purification* / Q000494:pharmacology / Q000633:toxicity; D029963:Quercus / Q000382:microbiology; D016019:Survival Analysis; D014242:Trichoderma / Q000737:chemistry* / Q000302:isolation & purification*
D000935:Antifungal Agents; D054848:Peptaibols