Sasaki Chiye, Vårum Kjell M, Itoh Yoshifumi, Tamoi Masahiro, Fukamizo Tamo
Department of Advanced Bioscience, Kinki University, Nakamachi, Nara, Japan.
Glycobiology. 2006 Dec;16(12):1242-50. doi: 10.1093/glycob/cwl043. Epub 2006 Sep 6.
Sugar recognition specificities of class III (OsChib1a) and class I (OsChia1cDeltaChBD) chitinases from rice, Oryza sativa L., were investigated by analyzing (1)H- and (13)C-nuclear magnetic resonance spectra of the enzymatic products from partially N-acetylated chitosans. The reducing end residue of the enzymatic products obtained by the class III enzyme was found to be exclusively acetylated, whereas both acetylated and deacetylated units were found at the nearest neighbor to the reducing end residue. Both acetylated and deacetylated units were also found at the nonreducing end residue and its nearest neighbor of the class III enzyme products. Thus, only subsite (-1) among the contiguous subsites (-2) to (+2) of the class III enzyme was found to be specific to an acetylated residue. For the class I enzyme, the reducing end residue was preferentially acetylated, although the specificity was not absolute. The nearest neighbor to the acetylated reducing end residue was specifically acetylated. Moreover, the nonreducing end residue produced by the class I enzyme was exclusively acetylated, although there was a low but significant preference for deacetylated units at the nearest neighbor to the nonreducing end. These results suggest that the three contiguous subsites (-2), (-1), and (+1) of the class I enzyme are specific to three consecutive GlcNAc residues of the substrate. In rice plants, the target of the class I enzyme might be a consecutive GlcNAc sequence probably in the cell wall of fungal pathogen, whereas the class III enzyme might act toward an endogenous complex carbohydrate containing GlcNAc residue.
通过分析部分N-乙酰化壳聚糖酶解产物的（1）H-和（13）C-核磁共振谱，研究了来自水稻（Oryza sativa L.）的III类（OsChib1a）和I类（OsChia1cDeltaChBD）几丁质酶的糖识别特异性。发现III类酶产生的酶解产物的还原端残基完全被乙酰化，而在还原端残基的最近邻位发现了乙酰化和脱乙酰化单元。在III类酶产物的非还原端残基及其最近邻位也发现了乙酰化和脱乙酰化单元。因此，发现III类酶相邻亚位点（-2）至（+2）中只有亚位点（-1）对乙酰化残基具有特异性。对于I类酶，还原端残基优先被乙酰化，尽管这种特异性不是绝对的。乙酰化还原端残基的最近邻位被特异性乙酰化。此外，I类酶产生的非还原端残基完全被乙酰化，尽管在非还原端最近邻位对脱乙酰化单元有较低但显著的偏好。这些结果表明，I类酶的三个相邻亚位点（-2）、（-1）和（+1）对底物的三个连续GlcNAc残基具有特异性。在水稻植株中，I类酶的作用靶点可能是真菌病原体细胞壁中可能存在的连续GlcNAc序列，而III类酶可能作用于含有GlcNAc残基 的内源性复合碳水化合物。
