Biochemical Properties of a Chitin-Binding Class III Chitinase in Pumpkin Leaves

  • Lee, Kyun-Oh (Department of Biochemistry, College of Natural Sciences, Gyeongsang National University) ;
  • Kim, Min-Gab (Department of Biochemistry, College of Natural Sciences, Gyeongsang National University) ;
  • Jang, Ho-Hee (Department of Biochemistry, College of Natural Sciences, Gyeongsang National University) ;
  • Lee, Ji-Yeun (Department of Biochemistry, College of Natural Sciences, Gyeongsang National University) ;
  • Kim, Sun-Chang (Department of Biological Science, Korea Advanced Institute of Science and Technology) ;
  • Lee, Sang-Yeol (Department of Biochemistry, College of Natural Sciences, Gyeongsang National University)
  • Received : 0
  • Accepted : 0
  • Published : 0

Abstract

When we compared the chitinase activity of various plant sources using colorimetric or active gel-staining assay methods, the specific activity of pumpkin leaves was the highest among the samples we analyzed. The highly active chitinase from pumpkin leaves (designated PL-ChtIII) was purified to homogeneity using affinity chitin gel and HPLC Mono-Q anion-exchange cloumn chromatographies. In contrast to other members of the class III chitinase family, PL-ChtIII showed a strong binding affinity to the regenerated chitin gel column. The apparent molecular weight of PL-ChtIII was estimated to be 29 kDa on SDS-PAGE gel, while its optimum pH and temperature were shown to be pH 6.0 and $60^{\circ}C$, respectively. Analyzing the reaction products of PL-ChtIII with swollen chitin as substrate, the dimer and tetramer of N-acetylglucosamine were produced as major products in the first hour of the enzymatic reaction along with a small amount of monomers and trimers. As the reaction time increased, dimeric N-acetylglucosamine became the predominant form of reaction product.

Keywords

Chitin binding activity;Class III chitinase;Enzymatic properties;Pumpkin leaves