STEREOCHEMISTRY OF THE HYDROLYSIS REACTION CATALYZED BY ENDOGLUCANASE-Z FROM ERWINIA-CHRYSANTHEMI
STEREOCHEMISTRY OF THE HYDROLYSIS REACTION CATALYZED BY ENDOGLUCANASE-Z FROM ERWINIA-CHRYSANTHEMI
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DOI：
10.1016/0014-5793(92)80183-h
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发表时间：
1992-03-30
期刊：
影响因子：
3.5
通讯作者：
HENRISSAT, B
中科院分区：
文献类型：
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作者：
BARRAS, F;BORTOLIGERMAN, I;HENRISSAT, B
通过微晶纤维素 Avicel PH101 上的亲和层析纯化来自植物病原菌菊欧文氏菌（菌株 3937）的内切葡聚糖酶 Z。使用对硝基苯基 β-D-纤维二糖苷和对硝基苯基 β-D-乳糖苷作为底物进行动力学表征：内切葡聚糖酶 Z 对对硝基苯基 β-D-纤维二糖苷的 K(m) 值分别为 3 mM 和 7.5 mM，V(m) 值分别为 129 和 40 nmol.min-1.mg-1 (k(cat) = 0.1 s-1）和 对硝基苯基 β-D-乳糖苷（k(cat) = 0.03 s-1），分别）。通过内切葡聚糖酶 Z 水解纤维四醇，然后进行 HPLC 和 H-1 NMR。结果表明，纤维二糖醇和 β-纤维二糖最初形成，表明该酶通过保留异头构型的分子机制发挥作用。这表明糖基酶中间体的参与。
Endoglucanase Z from the phytopathogenic bacterium Erwinia chrysanthemi (strain 3937) was purified by affinity chromatography on microcrystalline cellulose Avicel PH101. A kinetic characterization using p-nitrophenyl beta-D-cellobioside and p-nitrophenyl beta-D-lactoside as substrates was conducted: endoglucanase Z exhibited K(m) values of 3 mM and 7.5 mM and V(m) values of 129 and 40 nmol.min-1.mg-1 towards p-nitrophenyl beta-D-cellobioside (k(cat) = 0.1 s-1) and p-nitrophenyl beta-D-lactoside (k(cat) = 0.03 s-1), respectively). The hydrolysis of cellotetraitol by endoglucanase Z was followed by HPLC and H-1 NMR. Results show that cellobiitol and beta-cellobiose are initially formed, demonstrating that the enzyme is acting by a molecular mechanism retaining the anomeric configuration. This suggests the involvement of a glycosyl-enzyme intermediate.