1. Two unprecedented natural Aib-peptides with the (Xaa-Yaa-Aib-Pro) motif and an unusual C-terminus: structures, membrane-modifying and antibacterial properties of pseudokonins KL III and KL VI from the fungus Trichoderma pseudokoningii
S Rebuffat, C Goulard, S Hlimi, B Bodo J Pept Sci. 2000 Oct;6(10):519-33. doi: 10.1002/1099-1387(200010)6:103.0.CO;2-6.
Pseudokonins KL III and KL VI are two natural ten-residue peptides, which both contain the (Xaa-Yaa-Aib-Pro) motif and exhibit an unusual C-terminus. They have been isolated from the fungus Trichoderma pseudokoningii by intensive reversed-phase HPLC, beside peptaibols classically C-ended by a beta-amino alcohol. The amino acid sequences and the chemical structures of the C-ends have been determined by the combined use of positive ion LSI-MS and two-dimensional homo- and heteronuclear NMR, including COSY, TOCSY, ROESY, 13C heteronuclear single quantum correlation (HSQC) and heteronuclear multiple bond correlation (HMBC). Instead of one of the amino alcohols usually found as C-terminal residue in peptaibols, pseudokonins KL III and KL VI are characterized by -Pro-NH2 and cyclo-(Aib-L-Proal) (Proal, prolinal), respectively. Such backbone modifications are described here for the first time for peptaibol antibiotics. The unusual cyclo-(Aib-L-Proal) C-terminus is probably the result of an intramolecular cyclization of the two last Aib and Pro residues of a ten-amino acid precursor, via a Proal intermediate. A secondary structure stabilized by -C=O...H-N-hydrogen bonds of the 1<--4 type has been deduced for both peptides from ROESY data, 3JNHCalphaH couplings and amide proton temperature coefficient values. The (Xaa-Yaa-Aib-Pro) beta-bend ribbon spiral, which has been described for the first time in the case of a 14-residue peptaibol containing three repetitive (Xaa-Yaa-Aib-Pro) motifs (Segalas G et al. Biopolymers 1999; 50: 71-85) appears to be maintained in the two shortened modified peptides. The beta-bend ribbon structure thus appears to be initiated by a single (Xaa-Yaa-Aib-Pro) motif and unaffected by the C-terminal modifications. However, the membrane and antibiotic properties of pseudokonins KL III and KL VI, point to the unfavourable effect of both shortening and cyclization of the peptide chain.
2. Sequence and solution conformation of the 20-residue peptaibols, saturnisporins SA II and SA IV
S Rebuffat, L Conraux, M Massias, C Auvin-Guette, B Bodo Int J Pept Protein Res. 1993 Jan;41(1):74-84. doi: 10.1111/j.1399-3011.1993.tb00117.x.
Saturnisporins SA II and SA IV are the major components of the 20-residue peptaibol mixture isolated from a culture of the fungus Trichoderma saturnisporum. These peptides exhibit antibiotic activity against Staphylococcus aureus. Their sequences were derived from unequivocal methodology implying the combined use of positive ion FAB mass spectrometry and NMR: the majority of the sequences result from mass spectrometry fragmentations and the location of isomeric residues arises either from analysis of ROESY cross-peaks between contiguous amide protons or from heteronuclear 2J or 3J 1H-13C couplings detected in long-range 1H-13C COSY experiments. The sequence specific 1H and 13C NMR assignments are described. Saturnisporins SA II and SA IV exhibit similar secondary structures, as deduced from their ROESY patterns and 3JNHC alpha H coupling constant values, together with amide hydrogen-deuterium exchange rates and temperature coefficients of amide and carbonyl groups. An overall alpha-helical structure is proposed, maintaining two regions of distortion to this regular structure; i) the N-terminal part, which contains 3(10) and mixed alpha-3(10) turns, and ii) the Aib10-Val15 region, including a Pro residue which accommodates a bend stabilized by two 3(10) H-bonds.
3. Sequences and antimycoplasmic properties of longibrachins LGB II and LGB III, two novel 20-residue peptaibols from Trichoderma longibrachiatum
G Leclerc, C Goulard, Y Prigent, B Bodo, H Wróblewski, S Rebuffat J Nat Prod. 2001 Feb;64(2):164-70. doi: 10.1021/np000240s.
Longibrachins are members of the class of natural Aib-containing peptides designated as peptaibols. Six longibrachins, LGA I-IV and LGB II and III, were purified from a Trichoderma longibrachiatum strain by a procedure employing several chromatography steps including reversed-phase HPLC. The amino acid sequence determination was based on a combination of liquid secondary ion mass spectrometry (LSIMS) and two-dimensional 1H and 13C NMR spectroscopy. Longibrachins are 20-residue peptaibols with a C-terminal phenylalaninol and either neutral (LGA; Gln18) or acidic (LGB; Glu18) character. Longibrachins LGB II and III have novel sequences. Both longibrachins LGA and LGB show significant bactericidal activity against mycoplasmas (Acholeplasma, Mycoplasma, and Spiroplasma), with minimal inhibitory concentrations in the range 1.56-12.5 microM (3-25 micrograms/mL), and also perturb the permeability of membrane bilayers. Longibrachin LGA IV is the most potent of the presently known 18-20-residue peptaibols. The antimicrobial and membrane-perturbing properties of longibrachins, which are described here for the first time, were shown to be correlated.
