PURIFICATION AND PRIMARY STRUCTURE OF TWO ANTIMICROBIAL PEPTIDES FROM SKIN SECRETION OF Leptodacytlus pentadacylus (AMPHIBIAN: ANURA)

CORDEIRO-SOUSA J. (1), FONTENELE-BERTO R. F. (1), OLIVEIRA R. G. (1), REBOUÇAS JR J. P. (1), HONÓRIO JR J.E.R. (2), OLINDA A. C. C (1), CARVALHO I. F. (1), ADED DA SILVA P. (1), FONTENELE-CARDI N. C. F. (4), PIMENTA D. C. (3), CARDI B. A. (1,2), CARVALHO K. M.(1)

(1) Lab. de Toxinologia e Farmacologia Molecular, UECE; (2) Lab. de Animais Peçonhentos, UECE; (3) Centro de Toxinologia Aplicada (CAT/CEPID – FAPESP), Instituto Butantan ; (4) Hospital São José de Doenças Infecciosas, Ceará.

The potency of a particular peptide against microorganisms is determined by a complex interaction between factors relating to conformation cationicity, relatively hydrophobic, and have propensity to form an amphipathic a-helice in membrane-mimetic environment. The present study describes the purification and the structural and biological characterization of two peptides with antimicrobial activity from skin secretions of L. pentactylus. Adult specimens of L. pentadctylus were collected in Paraipaba (Ceará, Brazil) and maintained in captivity. The secretion from their cutaneous glands was obtained by mild electrical stimulation (2-7V), collected in a becker freezing, lyophilized and kept at -25ºC. The antimicrobial peptides were purified by HPLC with a C18 column (25X250mm), eluted with a flow of 4,5ml/min and using a 0-80% linear gradient of acetonitrile. The molecular mass was analyzed by ESI-Q-TOF Ultima API (Micromass). The purification assay resulted in the isolation of two fractions with molecular mass of 2275,34 and 2208,34 Da. The primary structure of new pentdacytins 1 and 2 were established with ESI-Q-TOF as GVLDLLKGAAKDLAGHLASKALD-NH2 and GVLDLLKGAAKNVVGLASKALD-NH2. A search using non-redundant database by CLUSTALW revealed 70% of homology to amphibian antimicrobial peptides pentadactylins. Prediction of the secondary structure indicates that these peptides have the propensity to adopt an extended a-helice conformation from residue (3-22).

KEY WORDS: Leptodactylus, antimicrobial peptides

FINANCIAL SUPPORT: FUNCAP, CAPES, CNPq

CORRESPONDENCE TO: carvalhokris@gmail.com