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Proceedings of the 31 st European Peptide SymposiumMichal Lebl, Morten Meldal, Knud J. Jensen, Thomas Hoeg-Jensen (Editors)European Peptide Society, 2010Antimicrobial Oligopeptides and Rapid Alkalinization Factorsin Chilean GrapeAlexander A. Zamyatnin 1,2 and Olga L. Voronina 21 Universidad Técnica Federico Santa Maria, Departamento de Informática, El CentroCientifico Tecnologico de Valparaiso, Valparaiso, 1680, Chile; 2 A.N.Bach Institute ofBiochemistry, Russian Academy of Sciences, Moscow, 119071, Russian FederationIntroductionRegulatory oligopeptides generally do not exceed ~50 amino acid residues [1], and theydiffer substantially from larger polypeptides (proteins) in their physicochemical andbiological properties. The primary structure of more than 10,000 oligopeptides from morethan 1700 different living organisms representing all the biological kingdoms have beenidentified today [2]. Now ~600 new natural oligopeptides emerge annually. Most of theinformation on the structure and function of endogenous oligopeptide molecules iscontained in different databases. Their primary structure is determined either directly or bytranslation from nucleotide sequences. Both ways are experimental and laborious.From the other hand there are a lot of unknown oligopeptide sequences. e.g., only onegrape (Vitis vinifera) oligopeptide primary structure was extracted and characterized beforeamong more than 1000 known plant oligopeptide regulators [2] whereas information onnumerous grape uncharacterized proteins has to be found in different protein databases.Since antimicrobial peptides play important role in the innate defense system of plantsand represent a relatively unexplored source of antimicrobial peptides of biotechnologicalpotential, it is necessary to obtain more information on their structures and functions.Rapid alkalinization factor (RALF) possessing new functional property wasdiscovered in common tobacco [3]. It consisted of 49 amino acid residues and induced arapid alkalinization of the culture medium of tobacco suspension-cultured cells and aconcomitant activation of an intracellular mitogen-activated protein kinase. A family ofoligopeptides inducing rapid pH alkalinization was isolated later from several plant species,but not from grape. Thereupon we have carried out a theoretical structure–function analysisof uncharacterized grape protein amino acid residue sequences in order to identify newprimary structures of oligopeptides possessing antimicrobial and RALF activities.Results and DiscussionThe data of DEGECHIVID database [6] containing primary structures of unrecognizedgrape proteins (more than 300,000 sequences) were used as an object of investigation.Their sequences were compared with EROP-Moscow database containing information onstructure and functions of plant regulatory oligopeptides [2] using specially createdcomputer programs [4,5].This method permitted to reveal 10 grape protein structure sites homologues to knownregulatory oligopeptides elucidated from other plant species. Six potentially activeantimicrobial oligopeptide sequences (Figure 1) have been identified among them. Theyconsisted of from 49 amino acid residues and their similarity with known grape primarystructure Vv-AMP1 [7] was from 66.0 to 100.0%. All these primary structures containedeight Cys residues forming potentially four disulfide bridges. One structure was the same asVv-AMP1 but had the difference in pre-pro-peptide region of oligopeptide precursor.Primary structures of several AMPs were found by us earlier [4]. However AMP3 andAMP4 had unique amino acid residue sequences not described before.This method revealed also four structures of potentially active rapid alkalinizationfactors (Figure 2). They consisted of from 49 to 57 amino acid residues and their similaritywith common tobacco RALF primary structure [3] was from 57.1 to 89.8%. All of themwere also characterized by a conserved sequences but containing four cysteine residuesforming potentially two disulfide bridges. Primary structures of these RALF1-RALF4found in other databases were described by us before [8].412