title: A trypsin-like proteinase in the midgut of Ectomyelois ceratoniae Zeller (Lepidoptera: Pyralidae): purification, characterization, and host plant inhibitors.
abstract: A trypsin-like proteinase was purified and characterized in the midgut of Ectomyelois ceratoniae. A purification process that used Sepharyl G-100 and DEAE-cellulose fast flow chromatographies revealed a proteinase with specific activity of 66.7 μmol/min/mg protein, recovery of 27.04 and purification fold of 23.35. Molecular weight of the purified protein was found to be 35.8 kDa. Optimal pH and temperature were obtained 9 and 20°C for the purified trypsin proteinase, respectively. The purified enzyme was significantly inhibited by PMSF, TLCK, and SBTI as specific inhibitors of trypsins in which TLCK showed the highest inhibitory effect. Trypsin proteinase inhibitors were extracted from four varieties of pomegranate including Brait, Torsh-Sabz, May-Khosh, and Shirin by ion exchange chromatography. It was found that fractions 17-20 of Brait; fractions 18 and 21-26 of Torsh-Sabz; fractions 1-7, 11-17, and 19-21 of May-Khosh and fraction 8 for Shirin showed presence of trypsin inhibitor in these host. Comparison of their inhibitory effects on the purified trypsin proteinase of E. ceratoniae demonstrated that fractions from May-khosh variety had the highest effect on the enzyme among other extracted fractions. Characterization of serine proteinases of insects mainly trypsins is one of the promising methods to decrease population and damages via extracting their inhibitors and providing resistant varieties.
authors: Department of Plant Protection, Faculty of Agricultural Sciences, University of Guilan, Rasht, Iran.
authors: Mina
authors: M
authors: Ranjbar
authors: Arash
authors: A
authors: Zibaee
authors: Jalal Jalali
authors: JJ
authors: Sendi
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002852:Chromatography, Ion Exchange; D004064:Digestive System / Q000201:enzymology; D004789:Enzyme Activation / Q000187:drug effects; D006863:Hydrogen-Ion Concentration; D007814:Larva; D007915:Lepidoptera / Q000201:enzymology*; D029561:Lythraceae / Q000737:chemistry; D010936:Plant Extracts / Q000494:pharmacology; D013696:Temperature; D014357:Trypsin / Q000302:isolation & purification* / Q000378:metabolism*; D014361:Trypsin Inhibitors / Q000302:isolation & purification / Q000494:pharmacology
chemical_list: D010936:Plant Extracts; D014361:Trypsin Inhibitors; D014357:Trypsin
keywords: Ectomyelois ceratoniae; proteinase inhibitor; purification; trypsin proteinase