Two Classes of New Peptaibols Are Synthesized by a Single Non-ribosomal Peptide Synthetase of Trichoderma virens

Abstract

Peptaibols are a group of small peptides having a high α-aminoisobutyric acid (Aib) content and produced by filamentous fungi, especially by the members of the genus Trichoderma (anamorph Hypocrea). These antibiotics are economically important for their anti-microbial and anti-cancer properties as well as ability to induce systemic resistance in plants against microbial invasion. In this study we present sequences of two classes (11-residue and 14-residue) of peptaibols produced by the biocontrol fungus Trichoderma virens. Of the 35 11-residue peptaibols sequenced, 18 are hitherto not described, and all the 53 14-residue sequences described by us here are new. We have also identified a peptaibol synthetase (non-ribosomal peptide synthetase, NRPS) with 14 complete modules in the genome of this fungus and disruption of this single gene (designated as tex2) resulted in the loss of both the classes of peptaibols. We, thus present here an unprecedented case where a single NRPS encodes for two classes of peptaibols. The new peptaibols identified here could have applications as therapeutic agents for the management of human and plant health. Peptaibols are a group of small peptides having a high α-aminoisobutyric acid (Aib) content and produced by filamentous fungi, especially by the members of the genus Trichoderma (anamorph Hypocrea). These antibiotics are economically important for their anti-microbial and anti-cancer properties as well as ability to induce systemic resistance in plants against microbial invasion. In this study we present sequences of two classes (11-residue and 14-residue) of peptaibols produced by the biocontrol fungus Trichoderma virens. Of the 35 11-residue peptaibols sequenced, 18 are hitherto not described, and all the 53 14-residue sequences described by us here are new. We have also identified a peptaibol synthetase (non-ribosomal peptide synthetase, NRPS) with 14 complete modules in the genome of this fungus and disruption of this single gene (designated as tex2) resulted in the loss of both the classes of peptaibols. We, thus present here an unprecedented case where a single NRPS encodes for two classes of peptaibols. The new peptaibols identified here could have applications as therapeutic agents for the management of human and plant health. IntroductionMicrobial resistance to classical antibiotics is a serious medical challenge, and new antimicrobial, membrane-active peptides have been proposed as alternatives to treat infections (1Duclohier H. Chem. Biodivers. 2007; 4: 1023-1026Crossref PubMed Scopus (45) Google Scholar). Peptaibols (a subgroup of peptaibiotics) (2Degenkolb T. Brückner H. Chem. Biodivers. 2008; 5: 1817-1843Crossref PubMed Scopus (72) Google Scholar) are peptides characterized by short linear chain lengths (≤20 residues), C-terminal alcohol residues, and high levels of non-standard amino acids, principally α-aminoisobutyric acid (Aib), 2The abbreviations used are: Aib, α-aminoisobutyric acid; NRPS, non-ribosomal peptide synthetase; Hyp, hydroxyproline; Iva, isovaline. isovaline (Iva), and the imino acid hydroxyproline (Hyp). Naturally occurring peptaibols have been isolated from soil fungi (mostly from the genus Trichoderma (teleomorph Hypocrea) and related genera), and often exhibit antibacterial, antifungal and anti-cancer properties (3Daniel J.F. Filho E.R. Nat. Prod. Rep. 2007; 24: 1128-1141Crossref PubMed Scopus (101) Google Scholar, 4Degenkolb T. von Döhren H. Nielsen K.F. Samuels G.J. Brückner H. Chem. Biodivers. 2008; 5: 671-680Crossref PubMed Scopus (64) Google Scholar, 5Degenkolb T. Gams W. Brückner H. Chem. Biodivers. 2008; 5: 693-706Crossref PubMed Scopus (38) Google Scholar, 6Kubicek C.P. Komoń-Zelazowska M. Sándor E. Druzhinina I.S. Chem. Biodivers. 2007; 4: 1068-1082Crossref PubMed Scopus (34) Google Scholar, 7Pruksakorn P. Arai M. Kotoku N. Vilchèze C. Baughn A.D. Moodley P. Jacobs Jr., W.R. Kobayashi M. Bioorg. Med. Chem. Lett. 2010; 20: 3658-3663Crossref PubMed Scopus (122) Google Scholar, 8Sang Y. Blecha F. Animal Health Res. Rev. 2008; 9: 227-235Crossref PubMed Scopus (181) Google Scholar, 9Chutrakul C. Alcocer M. Bailey K. Peberdy J.F. Chem. Biodivers. 2008; 5: 1694-1706Crossref PubMed Scopus (34) Google Scholar). The amphipathic nature of peptaibols allows them to self-associate into oligomeric ion-channel assemblies which span the width of lipid bi-layer membranes. Their antibiotic functions arise from their membrane insertion and pore-forming abilities (10Sansom M.S. Quart. Rev. Biophys. 1993; 26: 365-421Crossref PubMed Scopus (169) Google Scholar). Some of these peptides have also been shown to inhibit the replication of enveloped viruses such as influenza A virus, vesicular stomatitis, and human immunodeficiency virus (11Degenkolb T. Berg A. Gams W. Schlegel B. Gräfe U. J Pept. Sci. 2003; 9: 666-678Crossref PubMed Scopus (110) Google Scholar). In addition, some possess anticancer activities or promote wound healing (12Marahiel M.A. J Pept. Sci. 2009; 15: 799-807Crossref PubMed Scopus (101) Google Scholar). All these properties make these peptides good candidates as new therapeutic agents (13Shi M. Wang H.N. Xie S.T. Luo Y. Sun C.Y. Chen X.L. Zhang Y.Z. Mol. Cancer. 2010; 9: 26Crossref PubMed Scopus (63) Google Scholar). Recently it has been shown for the first time that peptaibols are regulators involved in both apoptosis and autophagy, suggesting that the class of peptaibols might also serve as potential suppressors of tumor cells (13Shi M. Wang H.N. Xie S.T. Luo Y. Sun C.Y. Chen X.L. Zhang Y.Z. Mol. Cancer. 2010; 9: 26Crossref PubMed Scopus (63) Google Scholar).Trichoderma spp., the major producers of peptaibols, are a unique group of filamentous fungi that are omnipresent in soil as highly competitive saprophytes, as well as rhizosphere and root colonizers. They are capable of protecting crops from root-invading fungi, parasitizing mushrooms and other fungi, and in some instances being opportunistic human pathogens (14Mukherjee P.K. Nautiyal C.S. Mukhopadhyay A.N. Nautiyal C.S. Dion P. Molecular Mechanisms of Plant and Microbe Coexistence (Soil Biology Series). Springer, Netherlands2008Google Scholar, 15Verma M. Brar S.K. Tyagi R.D. Surampalli R.Y. Surampalli J. Valéro J.R. Biochem. Eng. J. 2007; 37: 1-20Crossref Scopus (419) Google Scholar). Thus, these fungi occupy important roles in agriculture, medicine as well as industry (16Brotman Y. Kapuganti J.G. Viterbo A. Curr. Biol. 2010; 20: R390-R391Abstract Full Text Full Text PDF PubMed Scopus (69) Google Scholar, 17Schuster A. Schmoll M. Appl. Microbiol. Biotechnol. 2010; 87: 787-799Crossref PubMed Scopus (405) Google Scholar).Within the genus Trichoderma/Hypocrea, so far peptaibol subfamilies 1, 4, 5 and 9 have been described (18Szekeres A. Leitgeb B. Kredics L. Antal Z. Hatvani L. Manczinger L. Vágvölgyi C. Acta. Microbiol. Immunol. Hung. 2005; 52: 137-168Crossref PubMed Scopus (105) Google Scholar). Subfamily 1 (SF1) comprises about half of the known structures and includes peptides ranging from 18 to 20 residues in length. All of these peptides have partial sequence identities or similarities. Subfamily 4 (SF4) is comprised of peptides of 11–14 residues, also sharing sequence similarities, but having no sequence relationship to SF1. Subfamilies 5 and 9 (SF5 and -9) have only a few members and comprise peptides with 11 or 6 and 7 residues, respectively, again with no sequence similarities to the other subfamilies (19Neuhof T. Dieckmann R. Druzhinina I.S. Kubicek C.P. von Döhren H. Microbiology. 2007; 153: 3417-3437Crossref PubMed Scopus (54) Google Scholar). An extensive survey of 28 Trichoderma/Hypocrea species for peptaibol production by intact-cell MALDI-TOF mass spectrometry (ICMS) revealed that peptaibols are produced by all strains with some producing up to five peptide families of different sizes (19Neuhof T. Dieckmann R. Druzhinina I.S. Kubicek C.P. von Döhren H. Microbiology. 2007; 153: 3417-3437Crossref PubMed Scopus (54) Google Scholar). In addition to terrestrial strains, peptaibols have also been isolated from strains of Trichoderma P. Arai M. Kotoku N. Vilchèze C. Baughn A.D. Moodley P. Jacobs Jr., W.R. Kobayashi M. Bioorg. Med. Chem. Lett. 2010; 20: 3658-3663Crossref PubMed Scopus (122) Google Scholar, N. L. M. J. P. 2007; PubMed Scopus Google Scholar, J. C. L. M. Chen J. Z. P. W. J Nat. Prod. 2009; PubMed Scopus Google Scholar, M. M. J.F. F. P. M. J. 20: PubMed Scopus Google of peptaibols in Trichoderma is as of the has been of new peptaibols from a The of peptaibols in and has been M. T. Dieckmann R. von Döhren H. A. Kubicek C.P. Druzhinina I.S. 2007; PubMed Scopus Google and their in resistance in plants has been (18Szekeres A. Leitgeb B. Kredics L. Antal Z. Hatvani L. Manczinger L. Vágvölgyi C. Acta. Microbiol. Immunol. Hung. 2005; 52: 137-168Crossref PubMed Scopus (105) Google Scholar, J. T. F. W. Chem. PubMed Scopus Google Scholar, A. A. Y. Mol. Plant 2007; PubMed Scopus Google Scholar, B. Y. Y. K. W. Lett. Scopus (45) Google Scholar). a of peptaibols isolated from an biocontrol of Trichoderma has been to have antifungal activities against important thus their in biocontrol L. A. A. T. Microbiology. 2009; PubMed Scopus Google Scholar). The peptaibols from T. have been shown to exhibit J. A. R. M.A. M.S. Appl. Microbiol. PubMed Scopus Google are by known as non-ribosomal peptide that from a of amino and or which or M.A. T. Chem. Rev. PubMed Scopus Google Scholar, R. U. PubMed Google Scholar). have a in which is a that and a single of the peptide M.A. T. Chem. Rev. PubMed Scopus Google Scholar). into and which a of a NRPS T. J. A. B. P. M. R. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The first of the of an NRPS in peptaibol from of where we that an NRPS the peptaibol produced by T. A. A. Y. Mol. Plant 2007; PubMed Scopus Google Scholar, A. C. B. C. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus (169) Google Scholar, F. J Microbiol. 2005; PubMed Scopus Google Scholar). so have been no the of the peptaibols, of these peptaibols are in Trichoderma/Hypocrea (19Neuhof T. Dieckmann R. Druzhinina I.S. Kubicek C.P. von Döhren H. Microbiology. 2007; 153: 3417-3437Crossref PubMed Scopus (54) Google Scholar). In this we present new sequences of the and 14-residue peptaibols of T. and for the first that a single NRPS is for the of these two classes of of the and a the of the which a of and peptaibols in we have the of which and from of T. these could to the peptaibols not In peptaibols as ranging from to to and to for and to a the of of mass and of peptaibols from T. in in as shown in for of the of to the described by N. L. M. J. P. 2007; PubMed Scopus Google Scholar, M. M. J.F. F. P. M. J. 20: PubMed Scopus Google in by and and a for as in for the 14-residue peptaibol of and 14-residue peptaibols identified by from are in 1 and sequence the is sequences for are in The 11-residue peptaibols to a of sequences and with of and five sequences of 11-residue peptaibols identified from They to the peptide the sequence where is is α-aminoisobutyric to the and to amino sequences new and the to known 11-residue peptaibols such as N. L. M. J. P. 2007; PubMed Scopus Google Scholar, C. J. Brückner H. Chem. Biodivers. 2007; 4: PubMed Scopus (34) Google A. T. T. A. Schlegel B. Gräfe U. J. Pept. Sci. 2003; 9: PubMed Scopus (34) Google A. N. M. N. T. Chem. PubMed Scopus Google A. M. T. H. A. M. K. Chem. PubMed Scopus Google and from M. Brückner H. Scopus Google which to H. T. T. Gams W. Brückner H. J Chem. PubMed Scopus Google of 5 from to of the of the by in of the are by to major and by are in Aib, and the C-terminal of 11-residue peptaibols produced by T. in a new of 14-residue peptaibols produced by T. in a new of 14-residue peptaibols identified in characterized by a in 6 or The major from 1 to and with of and to the peptide as isolated from of T. which are characterized by a in 6 C. B. J Chem. Scopus Google Scholar). new sequences identified the sequence The other 14-residue peptaibol the They from and with of and different sequence The to the known of an in 6 C. B. J Chem. Scopus Google Scholar). new sequences have been identified this The a new group of 14-residue peptaibols. In this unprecedented is with a and the is by a to the in sequences A of the 11-residue and 14-residue peptaibols revealed an the amino 4, and 6 are in the 11-residue peptaibols 4 and of and 14-residue peptaibols produced by T. are in Aib, and the C-terminal to the proposed survey of the genome the of NRPS with high to the described a gene for an NRPS of 14 linear modules and with 7 modules is no unique NRPS with in the The NRPS is by a single of and by of and The has a the and 14 complete modules of in a linear A of all the of both the and from species of Trichoderma T. and T. with other from Trichoderma and other fungi that the to the peptaibol class of not An of the sequences the and from modules of from all is all but the first and of with the the T. M.A. Chem. Biol. Full Text PDF PubMed Scopus Google Scholar) The C. T. W. Res. 2005; PubMed Scopus Google the for the first as and is also good the of the of the other the of the the for is the of the is to A in the that is highly to the sequence of T. and a of an fungus and respectively, and for in the peptaibol synthetase used for The of for by is The of an the and to gene disruption is the of in case of the with the and the is present in the to the single is shown The with and with the that used for the disruption of the of of and residues are and The amino by as well as the are The residues and T. M.A. Chem. Biol. Full Text PDF PubMed Scopus Google Scholar) are by the single used to T. P.K. J. R. 2003; PubMed Scopus Google we (designated as and for of the the of the in of the and the single by no major the and the in of that the a All the both the and 14-residue peptaibols as from the MALDI-TOF disruption of a single gene resulted in loss of two classes of peptaibols, we the is no the of 6 is as a We also the of and not of and a that both the 11-residue and 14-residue peptaibols are in the of and 5 of are the major of by known as non-ribosomal peptide These peptides have potential applications as and anticancer in addition to their as of plant In this we isolated and characterized a peptaibol of 11 and 14 amino acid residues from the The of both of peptaibols is in peptaibols also produced by and to the group and have been in a study A. C. B. C. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus (169) Google Scholar). to their sequence 11-residue peptaibols isolated from to the major and of 11-residue peptaibols produced by Trichoderma genus for which sequences have been in the A high in the 11-residue peptaibol produced by T. of 35 18 and from other by in 1, 4, and These are in with which related high of 11-residue peptaibol isolated from T. and T. N. L. M. J. P. 2007; PubMed Scopus Google Scholar, C. J. Brückner H. Chem. Biodivers. 2007; 4: PubMed Scopus (34) Google 14-residue peptaibol sequences have been described in to 11-residue sequences have been to the group isolated from the T. strains and C. B. J Chem. Scopus Google Scholar). In the present 14-residue sequences to group have been from with residues in 1, and The known of by the which or have been in all of these sequences identified to new sequences of the different a in and in 6 is for the first is the of sequences so far for 14-residue peptaibols, and the first of sequences identified from a T. to are peptaibol sequences (2Degenkolb T. Brückner H. Chem. Biodivers. 2008; 5: 1817-1843Crossref PubMed Scopus (72) Google the of their is a few such as A. A. Y. Mol. Plant 2007; PubMed Scopus Google Scholar, A. C. B. C. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus (169) Google Scholar, F. J Microbiol. 2005; PubMed Scopus Google and a partial peptaibol synthetase gene in T. L. B. E. Microbiol. PubMed Scopus Google Scholar). genome revealed that in addition to an peptaibol synthetase, the T. genome two with high with a and a These are related to other the peptaibol MALDI-TOF revealed the of an a 14-residue and an 11-residue peptaibol the Trichoderma a 11-residue peptaibol or a 14-residue or both (19Neuhof T. Dieckmann R. Druzhinina I.S. Kubicek C.P. von Döhren H. Microbiology. 2007; 153: 3417-3437Crossref PubMed Scopus (54) Google Scholar). we that the of resulted in the loss of all and peptaibols, it by us and group that for only the peptaibol A. A. Y. Mol. Plant 2007; PubMed Scopus Google Scholar, A. C. B. C. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus (169) Google Scholar, F. J Microbiol. 2005; PubMed Scopus Google Scholar). The of the 11-residue peptaibol has thus been a as is no NRPS gene in the Trichoderma In the present for the first present that disruption of a single NRPS gene in the loss of two peptaibols, both 14-residue and the of the and 14-residue allows the of sequence similarities the two of peptaibols 4 and both of peptaibols could from a single peptide the sequence of 14-residue peptaibols, (19Neuhof T. Dieckmann R. Druzhinina I.S. Kubicek C.P. von Döhren H. Microbiology. 2007; 153: 3417-3437Crossref PubMed Scopus (54) Google Scholar) that and 14-residue and could produced by a single peptaibol synthetase by of modules into the of the it that in 11-residue peptaibols could from 14-residue peptaibols by of the residues in 4, and 6 or could by an that is the modules for of residues 4, and the 11-residue peptaibol is the 14-residue peptaibol the amino 4, and it that of these in the of 11-residue from the 14-residue both are the and the and the of the Thus, is no that the NRPS arise by an of the of the NRPS not the production of or 11-residue peptaibols not are two by which the gene could for of both 11-residue and 14-residue peptaibols, an of the gene or of these modules by the NRPS of an as are no to these modules The only is of the modules of the 11-residue peptaibols. peptaibol has not been but this of of non-ribosomal peptides to M. Wang Curr. Chem. 2010; Scopus Google Scholar, R. J Microbiol. Biotechnol. PubMed Scopus Google Scholar, W. K. K. M. M. T. H. R. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, R. Curr. Chem. Biol. 2005; 9: PubMed Scopus Google Scholar, B. B. M. H. R. Chem. Chem. 2005; PubMed Scopus Google Scholar, P. T. R. Chem. PubMed Scopus Google Scholar, Bailey Chem. 2005; PubMed Scopus Google Scholar). of two of 11-residue and 14-residue by a single could have for fungi, T. as these are characterized by microbial and the peptaibols are known to addition to the we here the sequences of 35 which 18 are 11-residue and 53 14-residue peptaibols in T. These new peptaibol sequences described here could a of and important therapeutic agents for management of human and plant health. IntroductionMicrobial resistance to classical antibiotics is a serious medical challenge, and new antimicrobial, membrane-active peptides have been proposed as alternatives to treat infections (1Duclohier H. Chem. Biodivers. 2007; 4: 1023-1026Crossref PubMed Scopus (45) Google Scholar). Peptaibols (a subgroup of peptaibiotics) (2Degenkolb T. Brückner H. Chem. Biodivers. 2008; 5: 1817-1843Crossref PubMed Scopus (72) Google Scholar) are peptides characterized by short linear chain lengths (≤20 residues), C-terminal alcohol residues, and high levels of non-standard amino acids, principally α-aminoisobutyric acid (Aib), 2The abbreviations used are: Aib, α-aminoisobutyric acid; NRPS, non-ribosomal peptide synthetase; Hyp, hydroxyproline; Iva, isovaline. isovaline (Iva), and the imino acid hydroxyproline (Hyp). Naturally occurring peptaibols have been isolated from soil fungi (mostly from the genus Trichoderma (teleomorph Hypocrea) and related genera), and often exhibit antibacterial, antifungal and anti-cancer properties (3Daniel J.F. Filho E.R. Nat. Prod. Rep. 2007; 24: 1128-1141Crossref PubMed Scopus (101) Google Scholar, 4Degenkolb T. von Döhren H. Nielsen K.F. Samuels G.J. Brückner H. Chem. Biodivers. 2008; 5: 671-680Crossref PubMed Scopus (64) Google Scholar, 5Degenkolb T. Gams W. Brückner H. Chem. Biodivers. 2008; 5: 693-706Crossref PubMed Scopus (38) Google Scholar, 6Kubicek C.P. Komoń-Zelazowska M. Sándor E. Druzhinina I.S. Chem. Biodivers. 2007; 4: 1068-1082Crossref PubMed Scopus (34) Google Scholar, 7Pruksakorn P. Arai M. Kotoku N. Vilchèze C. Baughn A.D. Moodley P. Jacobs Jr., W.R. Kobayashi M. Bioorg. Med. Chem. Lett. 2010; 20: 3658-3663Crossref PubMed Scopus (122) Google Scholar, 8Sang Y. Blecha F. Animal Health Res. Rev. 2008; 9: 227-235Crossref PubMed Scopus (181) Google Scholar, 9Chutrakul C. Alcocer M. Bailey K. Peberdy J.F. Chem. Biodivers. 2008; 5: 1694-1706Crossref PubMed Scopus (34) Google Scholar). The amphipathic nature of peptaibols allows them to self-associate into oligomeric ion-channel assemblies which span the width of lipid bi-layer membranes. Their antibiotic functions arise from their membrane insertion and pore-forming abilities (10Sansom M.S. Quart. Rev. Biophys. 1993; 26: 365-421Crossref PubMed Scopus (169) Google Scholar). Some of these peptides have also been shown to inhibit the replication of enveloped viruses such as influenza A virus, vesicular stomatitis, and human immunodeficiency virus (11Degenkolb T. Berg A. Gams W. Schlegel B. Gräfe U. J Pept. Sci. 2003; 9: 666-678Crossref PubMed Scopus (110) Google Scholar). In addition, some possess anticancer activities or promote wound healing (12Marahiel M.A. J Pept. Sci. 2009; 15: 799-807Crossref PubMed Scopus (101) Google Scholar). All these properties make these peptides good candidates as new therapeutic agents (13Shi M. Wang H.N. Xie S.T. Luo Y. Sun C.Y. Chen X.L. Zhang Y.Z. Mol. Cancer. 2010; 9: 26Crossref PubMed Scopus (63) Google Scholar). Recently it has been shown for the first time that peptaibols are regulators involved in both apoptosis and autophagy, suggesting that the class of peptaibols might also serve as potential suppressors of tumor cells (13Shi M. Wang H.N. Xie S.T. Luo Y. Sun C.Y. Chen X.L. Zhang Y.Z. Mol. Cancer. 2010; 9: 26Crossref PubMed Scopus (63) Google Scholar).Trichoderma spp., the major producers of peptaibols, are a unique group of filamentous fungi that are omnipresent in soil as highly competitive saprophytes, as well as rhizosphere and root colonizers. They are capable of protecting crops from root-invading fungi, parasitizing mushrooms and other fungi, and in some instances being opportunistic human pathogens (14Mukherjee P.K. Nautiyal C.S. Mukhopadhyay A.N. Nautiyal C.S. Dion P. Molecular Mechanisms of Plant and Microbe Coexistence (Soil Biology Series). Springer, Netherlands2008Google Scholar, 15Verma M. Brar S.K. Tyagi R.D. Surampalli R.Y. Surampalli J. Valéro J.R. Biochem. Eng. J. 2007; 37: 1-20Crossref Scopus (419) Google Scholar). Thus, these fungi occupy important roles in agriculture, medicine as well as industry (16Brotman Y. Kapuganti J.G. Viterbo A. Curr. Biol. 2010; 20: R390-R391Abstract Full Text Full Text PDF PubMed Scopus (69) Google Scholar, 17Schuster A. Schmoll M. Appl. Microbiol. Biotechnol. 2010; 87: 787-799Crossref PubMed Scopus (405) Google Scholar).Within the genus Trichoderma/Hypocrea, so far peptaibol subfamilies 1, 4, 5 and 9 have been described (18Szekeres A. Leitgeb B. Kredics L. Antal Z. Hatvani L. Manczinger L. Vágvölgyi C. Acta. Microbiol. Immunol. Hung. 2005; 52: 137-168Crossref PubMed Scopus (105) Google Scholar). Subfamily 1 (SF1) comprises about half of the known structures and includes peptides ranging from 18 to 20 residues in length. All of these peptides have partial sequence identities or similarities. Subfamily 4 (SF4) is comprised of peptides of 11–14 residues, also sharing sequence similarities, but having no sequence relationship to SF1. Subfamilies 5 and 9 (SF5 and -9) have only a few members and comprise peptides with 11 or 6 and 7 residues, respectively, again with no sequence similarities to the other subfamilies (19Neuhof T. Dieckmann R. Druzhinina I.S. Kubicek C.P. von Döhren H. Microbiology. 2007; 153: 3417-3437Crossref PubMed Scopus (54) Google Scholar). An extensive survey of 28 Trichoderma/Hypocrea species for peptaibol production by intact-cell MALDI-TOF mass spectrometry (ICMS) revealed that peptaibols are produced by all strains with some producing up to five peptide families of different sizes (19Neuhof T. Dieckmann R. Druzhinina I.S. Kubicek C.P. von Döhren H. Microbiology. 2007; 153: 3417-3437Crossref PubMed Scopus (54) Google Scholar). In addition to terrestrial strains, peptaibols have also been isolated from strains of Trichoderma P. Arai M. Kotoku N. Vilchèze C. Baughn A.D. Moodley P. Jacobs Jr., W.R. Kobayashi M. Bioorg. Med. Chem. Lett. 2010; 20: 3658-3663Crossref PubMed Scopus (122) Google Scholar, N. L. M. J. P. 2007; PubMed Scopus Google Scholar, J. C. L. M. Chen J. Z. P. W. J Nat. Prod. 2009; PubMed Scopus Google Scholar, M. M. J.F. F. P. M. J. 20: PubMed Scopus Google of peptaibols in Trichoderma is as of the has been of new peptaibols from a The of peptaibols in and has been M. T. Dieckmann R. von Döhren H. A. Kubicek C.P. Druzhinina I.S. 2007; PubMed Scopus Google and their in resistance in plants has been (18Szekeres A. Leitgeb B. Kredics L. Antal Z. Hatvani L. Manczinger L. Vágvölgyi C. Acta. Microbiol. Immunol. Hung. 2005; 52: 137-168Crossref PubMed Scopus (105) Google Scholar, J. T. F. W. Chem. PubMed Scopus Google Scholar, A. A. Y. Mol. Plant 2007; PubMed Scopus Google Scholar, B. Y. Y. K. W. Lett. Scopus (45) Google Scholar). a of peptaibols isolated from an biocontrol of Trichoderma has been to have antifungal activities against important thus their in biocontrol L. A. A. T. Microbiology. 2009; PubMed Scopus Google Scholar). The peptaibols from T. have been shown to exhibit J. A. R. M.A. M.S. Appl. Microbiol. PubMed Scopus Google are by known as non-ribosomal peptide that from a of amino and or which or M.A. T. Chem. Rev. PubMed Scopus Google Scholar, R. U. PubMed Google Scholar). have a in which is a that and a single of the peptide M.A. T. Chem. Rev. PubMed Scopus Google Scholar). into and which a of a NRPS T. J. A. B. P. M. R. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The first of the of an NRPS in peptaibol from of where we that an NRPS the peptaibol produced by T. A. A. Y. Mol. Plant 2007; PubMed Scopus Google Scholar, A. C. B. C. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus (169) Google Scholar, F. J Microbiol. 2005; PubMed Scopus Google Scholar). so have been no the of the peptaibols, of these peptaibols are in Trichoderma/Hypocrea (19Neuhof T. Dieckmann R. Druzhinina I.S. Kubicek C.P. von Döhren H. Microbiology. 2007; 153: 3417-3437Crossref PubMed Scopus (54) Google Scholar). In this we present new sequences of the and 14-residue peptaibols of T. and for the first that a single NRPS is for the of these two classes of peptaibols.