Production, characterization and identification using proteomic tools of a polygalacturonase from Fusarium graminearum

ORTEGA, L.; KIKOT. G.E.; LOPEZ, N.L; N.L. ROJAS,; ASTORECA, A.; ALCONADA, T.

JOURNAL OF BASIC MICROBIOLOGY

WILEY-V C H VERLAG GMBH

Lugar: Weinheim; Año: 2014 vol. 54 p. 170 - 177

0233-111X

A Fusarium graminearum isolate obtained from infected wheat spikes of Argentina Pampa region, with high polygalacturonase activity was selected for this study. A polygalacturonase has been purified 375 times from 2-day-old culture filtrates by gel filtration and ion-exchange chromatography successively. The purified sample showed two protein bands in SDS- polyacrylamide gels, with a molecular mass of 40 and 55 kDa. The protein bands were identified as an endopolygalacturonase and as a serine carboxypeptidase of F. graminearum respectively, by peptide mass fingerprinting and MALDI TOF/TOF fragment ion analysis. The pattern of substrate degradation analyzed by thin layer chromatography confirmed the mode of action of the enzyme as an endopolygalacturonase. High activity of the polygalacturonase against polygalacturonic acid was observed between 4 and 6 of pH, and between 30 and 50ºC, being 5 and 50ºC the optimum pH and temperature, respectively. The enzyme was fully stable at pH 5 for 120 min and 30ºC and sensible to the presence of some metal ions. The significance of the study of endopolygalacturonase activity is based on that such activity is produced in the early stages of infection on the host, suggesting a crucial role in the establishment of disease.