Comparative study of the reaction mechanism of family 18 chitinases from plants and microbes

Chiye Sasaki, Ai Yokoyama, Yoshifumi Itoh, Masayuki Hashimoto, Takeshi Watanabe, Tamo Fukamizo

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57 Citations (Scopus)


Hydrolytic mechanisms of family 18 chitinases from rice (Oryza sativa L.) and Bacillus circulans WL-12 were comparatively studied by a combination of HPLC analysis of the reaction products and theoretical calculation of reaction time-courses. All of the enzymes tested produced β-anomers from chitin hexasaccharide [(GlcNAc)6], indicating that they catalyze the hydrolysis through a retaining mechanism. The rice chitinases hydrolyzed predominantly the fourth and fifth glycosidic linkages from the nonreducing end of (GlcNAc)6, whereas B. circulans chitinase A1 hydrolyzed the second linkage from the nonreducing end. In addition, the Bacillus enzyme efficiently catalyzed transglycosylation, producing significant amounts of chitin oligomers larger than the initial substrate, but the rice chitinases did not. The time-courses of (GlcNAc)6 degradation obtained by HPLC were analyzed by theoretical calculation, and the subsite structures of the rice chitinases were identified to be (-4)(-3)(-2)(-1)(+1) (+2). From the HPLC profile of the reaction products previously reported [Terwisscha van Scheltinga et al. (1995) Biochemistry 34, 15619-15623], family 18 chitinase from rubber tree (Hevea brasiliensis) was estimated to have the same type of subsite structure. Theoretical analysis of the reaction time-course for the Bacillus enzyme revealed that the enzyme has (-2)(-1) (+1)(+2)(+3)(+4)-type subsite structure, which is identical to that of fungal chitinase from Coccidioides immitis [Fukamizo et al. (2001) Biochemistry 40, 2448-2454]. The Bacillus enzyme also resembled the fungal chitinase in its transglycosylation activity. Minor structural differences between plant and microbial enzymes appear to result in such functional variations, even though all of these chitinases are classified into the identical family of glycosyl hydrolases.

Original languageEnglish
Pages (from-to)557-564
Number of pages8
JournalJournal of biochemistry
Issue number4
Publication statusPublished - 2002 Apr

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology


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