Balázs Leitgeb

Recent Results in Alamethicin Research

by László Kredics*a), András Szekeresa), Dorina Czifraa), Csaba Vágvçlgyia), and Balázs Leitgebb) a) Department of Microbiology, Faculty of Science and Informatics, University of Szeged, Kçzép fasor 52, H-6726 Szeged (phone: þ36-62-544-516; fax: þ36-62-544-823; e-mail: kredics@bio.u-szeged.hu) b) Institute of Biophysics, Biological Research Centre of the Hungarian Academy of Sciences, Temesvári krt. 62, H-6726 Szeged

Structural characterization of the short peptaibols trichobrachins by molecular-dynamics methods.

A structural characterization was carried out by molecular‐dynamics methods for eight trichobrachin peptides, to identify the conformational features of these short peptaibols. For all peptides, the backbone and side‐chain conformations were investigated, different secondary structures, such as type‐I and ‐III β‐turns as well as β‐bend ribbon spirals, were determined in certain tetrapeptide units of the molecules, and the preferred rotamers of the side chains of amino acids were identified. Furthermore, the end‐to‐end and residueresidue distances were examined, as well as the fluctuations of backbone atoms were studied. Based on these results, the peptides were compared to one another. Our theoretical study indicated that trichobrachins could be characterized by typical structural properties, and both conformational similarities and dissimilarities were observed between these peptaibols. In summary, this structural investigation supplied a characterization of the various conformational features of eight trichobrachin peptides.

Exploring and characterizing the folding processes of Lys- and Arg-containing Ala-based peptides: A molecular dynamics study

In this study, molecular dynamics simulations were carried out on Lys- and Arg-containing Ala-based peptides (i.e. Ace-(AAAAK)(n)A-NH(2) and Ace-(AAAAR)(n)A-NH(2), where n=1-4), in order to explore and characterize their folding processes. For the oligopeptides, the evolution of α-helical structure with regard to the whole conformation, as well as to each residue was investigated, and the helix-forming propensities were characterized. On the basis of the helicity curves, representing the alteration of average helicity as a function of time, the typical time values describing the folding processes and subprocesses were identified. In the case of each peptide, the evolution and role of helix-stabilizing, non-local and side-chain-to-backbone H-bonds were examined. The appearing i←i+4 H-bonds pointed out the role of these interactions in the stabilization of α-helical conformations, while the occurring i←i+3 H-bonds indicated the presence of β-turn or 3(10)-helical structures. Studying the formation and role of non-local and side-chain-to-backbone H-bonds led to the observation that these types of interactions produced an effect on the evolution of helical conformations, as well as on the folding processes.

Diversity Profile and Dynamics of Peptaibols Produced by Green Mould Trichoderma Species in Interactions with Their Hosts Agaricus bisporus and Pleurotus ostreatus

Certain Trichoderma species are causing serious losses in mushroom production worldwide. Trichoderma aggressivum and Trichoderma pleuroti are among the major causal agents of the green mould diseases affecting Agaricus bisporus and Pleurotus ostreatus, respectively. The genus Trichoderma is well‐known for the production of bioactive secondary metabolites, including peptaibols, which are short, linear peptides containing unusual amino acid residues and being synthesised via non‐ribosomal peptide synthetases (NRPSs). The aim of this study was to get more insight into the peptaibol production of T. aggressivum and T. pleuroti. HPLC/MS‐based methods revealed the production of peptaibols closely related to hypomurocins B by T. aggressivum, while tripleurins representing a new group of 18‐residue peptaibols were identified in T. pleuroti. Putative NRPS genes enabling the biosynthesis of the detected peptaibols could be found in the genomes of both Trichoderma species. In vitro experiments revealed that peptaibols are potential growth inhibitors of mushroom mycelia, and that the host mushrooms may have an influence on the peptaibol profiles of green mould agents.

Characterizing the structural and folding properties of long-sequence hypomurocin B peptides and their analogs

We studied the folding processes of long‐sequence hypomurocin (HM) peptides and their analogs by means of molecular dynamics methods, focusing on the formation of various helical structures and intramolecular H‐bonds. The evolution of different helical conformations, such as the 310‐, α‐, and left‐handed α‐helices, was examined, taking into account the entire sequence and each amino acid of peptides. The results indicated that the HM peptides and their analogs possessed a propensity to adopt helical conformations, and they showed a preference for the 310‐helical structure over the α‐helical one. The evolution of a variety of the intramolecular H‐bonds, including local and non‐local interactions, was also investigated. The results pointed out that on the one hand, the appearance of local, helix‐stabilizing H‐bonds correlated with the presence of helical conformations, and on the other hand, the non‐local H‐bonds did not affect significantly the formation of helical structures. Additionally, comparing the structural and folding features of HM peptides and their analogs, our study led to the observation that the L‐D isomerism of isovaline amino acid induced effects on the folding processes of these long‐sequence peptaibol molecules. Accordingly, the HM peptides and their analogs could be characterized by typical structural and folding properties.

Effects of missense mutation on structure and function of photoreceptor

Phytochromes (PHYs) are photoreceptors of the red (R ~660 nm) and far-red (FR ~730 nm) light, and they control a wide range of responses affecting crucial aspects of plant life. There are five genes PHYA-PHYE encoding for phytochromes of different but overlapping function. One of these, PHYA has the unique function controlling specific responses in high irradiance far-red, as well as in very weak light. Appropriate PHYA functioning requires not only the photoreversibility of molecule but also the proper nuclear localization and degradation of receptor. Recently, we identified and described a mutant PHYA allele (phyA-5) in Arabidopsis thaliana, which showed reduced binding affinity to FHY1/FHL, the proteins regulating its nuclear transport, resulting in impaired nuclear localization and altered signaling under certain conditions. We present here a hypothesis to explain how the identified amino acid substitution may lead to structural changes manifested as altered signaling and phenotype displayed by the phyA-5 mutant.

Bioactive Peptaibols of Forest-Derived Trichoderma Isolates from Section Longibrachiatum

Filamentous fungi are producers of a large number of secondary metabolites with wide spectra of biological effects. Among them, peptaibols represent a group of compounds produced mainly by members of the mycotrophic filamentous fungal genus Trichoderma. A simple peptaibol characterization strategy including purification and structural elucidation steps was applied to examine the peptaibol production of three strains from the Longibrachiatum section of genus Trichoderma, T. aethiopicum TUCIM 1817, T. novae-zelandiae TUCIM 4158 and T. pseudokoningii TUCIM 1277, all deriving from natural forest habitats (disturbed semiforest, native Notophagus forest and the bark of Beilschmiedia tawa, respectively). After the solid phase clean-up of culture extracts, mass spectrometric analysis of peptaibols produced by the examined strains was performed by on-line reversed-phase high performance liquid chromatography coupled to electrospray ionization ion trap mass spectrometry. All three examined species produced 20-residue trichobrachin-like compounds, some of which are known from the literature, while others proved to be different from any peptaibols reported so far. The spectra of the peptaibols produced by these isolates were entirely different from each other. The largest amount of peptaibols consisting of four yet unknown compounds was produced by T. pseudokoningii TUCIM 1277, while ten and eight new, trichobrachin-like compounds were detected from T. aethiopicum TUCIM 1817 and T. novae-zelandiae TUCIM 4158, respectively. Feline fetal lung cell proliferation inhibition tests and membrane damage bio-assay with boar sperm cells revealed that although T. novae-zelandiae TUCIM 4158 produced the least amount of peptaibols, its compounds were the most inhibitory to mammalian cells.

In silico conformational analysis of the short-sequence hypomurocin a peptides.

In this theoretical study, a conformational analysis was performed on short-sequence hypomurocin A peptides, in order to identify their characteristic structural properties. For each hypomurocin A molecule, not only the backbone conformations, but also the side-chain conformations were examined. The results indicated that certain tetrapeptide units could be characterized by types I and III β-turn structures, and considering the helical conformations, it could be concluded that the hypomurocin A peptides showed a preference for the 310-helical structure over the α-helical structure. Beside the backbone conformations, the side-chain conformations were investigated, and the preferred rotamer states of the side-chains of amino acids were determined. Furthermore, the occurrence of i ← i + 3 and i ← i + 4 intramolecular H-bonds was studied, which could play a role in the structural stabilization of β-turns and helical conformations. On the whole, our theoretical study supplied a comprehensive characterization of the three-dimensional structure of short-sequence hypomurocin A peptides.

Studying the Structural and Folding Features of Long-Sequence Trichobrachin Peptides

In this theoretical study, the folding processes of long‐sequence trichobrachin peptides (i.e., TB IIb peptides) were investigated by molecular dynamics methods. The formation of various helical structures (i.e., 310‐, α‐, and left‐handed α‐helices) was studied with regard to the entire sequence of peptides, as well as to each amino acid. The results pointed out that TB IIb molecules showed a propensity to form helical conformations, and they could be characterized by 310‐helical structure rather than by α‐helical structure. The formation of local (i.e., i←i+3 and i←i+4) as well as of non‐local (i.e., i←i+n, where n>4; and all i→i+n) H‐bonds was also examined. The results revealed that the occurrence of local, helix‐stabilizing H‐bonds was in agreement with the appearance of helical conformations, and the non‐local H‐bonds did not produce relevant effects on the evolution of helical structures. Based on the data obtained by our structural investigation, differences were observed between the TB IIb peptides, according to the type of amino acid located in the 17th position of their sequences. In summary, the folding processes were explored for TB IIb molecules, and our theoretical study led to the conclusion that these long‐sequence peptaibols showed characteristic structural and folding features.

Spatial relationships between the pharmacophores of endomorphin-2: a comparative study of stereoisomers

AbstractThe spatial relationships between the pharmacophore elements were investigated in the case of four different stereoisomeric forms of opioid tetrapeptide, endomorphin-2, taking into account the L-D and cis-trans isomerisms. On the basis of distances and angles measured between the pharmacophoric points, a comparative analysis of conformational distributions was performed, applying a variety of distance-angle maps. The results obtained by this theoretical study indicated that the stereoisomers of endomorphin-2 could be distinguished from one another, based on the comparative analysis of distance-angle maps. Nevertheless, it could be concluded that this method proved to be suitable to examine the effects of L-D and cis-trans isomerisms on the spatial relationships of the pharmacophores of tetrapeptide.