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利用“酶晶体低温抑活底物固定”方法,在-20℃条件下,将6-磷酸-β-葡萄糖苷酶(BglA)晶体与底物对硝基苯-β-D-吡喃葡萄糖苷-6-磷酸(pNPβG6P)进行浸泡试验,通过X射线进行衍射数据收集并处理,获得了完整的底物电子密度图。研究结果表明,在不突变酶中关键基团使酶失活,以及不选用底物类似物替代原有底物的情况下,通过上述方法,可以获得酶与底物复合物的实际结构。该研究结果可为今后低温酶学及复合物中间态的进一步研究提供帮助。
Using the method of immobilization of low temperature substrate by enzyme crystal, the crystal of 6-phospho-β-glucosidase (BglA) and the substrate p-nitrophenyl-β-D-pyran Glucopyranosine-6-phosphate (pNPβG6P) was immersed in the test, the diffraction data were collected by X-ray and processed to obtain the complete electron density map of the substrate. The results show that the actual structure of the complex of enzyme and substrate can be obtained by the above method without deactivating the key group in the mutant enzyme and without replacing the original substrate with the substrate analogue. The results of this study may be helpful for further research on the low temperature enzymology and the complex intermediate state.