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Expression, characterization, and site-directed mutagenesis of UDP-glycosyltransferase UGT88A1 from Arabidopsis thaliana.
Bioengineered ( IF 4.2 ) Pub Date : 2019-05-09 , DOI: 10.1080/21655979.2019.1607710
Jingyuan Weng 1 , Liangliang Chen 1 , Yinchu Cheng 1 , Yan Li 1 , Honghua Jia 1 , Hua Zhou 1 , Ping Wei 1
Affiliation  

Quercetin-4′-O-glucoside is one of the major quercetin derivatives in the mature red onion bulb. It has an adjuvant effect on allergies, asthma, arthritis, and cancer. The present study aimed to use uridine diphosphate glycosyltransferase 88A1 (UGT88A1) from Arabidopsis thaliana to achieve the enzymatic synthesis of quercetin-4′-O-glucoside from quercetin. The results showed that UGT88A1 was most active at pH 9.0. The optimum temperature of UGT88A1 for synthesizing quercetin-4′-O-glucoside was 45°C, which was a little lower than that for synthesizing quercetin-3-O-glucoside (50°C). One mutant, V18R, of UGT88A1 was obtained by site-directed mutation and showed a greater affinity (Km 0.20 mM) and twice the enzyme activity (552.3 mU/mg) towards quercetin compared with the wild-type enzyme (0.36 mM and 227.6 mU/mg, respectively). The possible reason could be attributed to the distance change between the 18th amino-acid residue of UGT88A1 and the substrate quercetin, as deduced by molecular simulation.



中文翻译:


拟南芥 UDP-糖基转移酶 UGT88A1 的表达、表征和定点诱变。



Quercetin-4′- O- glucoside 是成熟红洋葱球茎中的主要槲皮素衍生物之一。对过敏、哮喘、关节炎、癌症等有辅助作用。本研究旨在利用拟南芥尿苷二磷酸糖基转移酶88A1(UGT88A1)实现槲皮素酶法合成槲皮素-4'- O-葡萄糖苷。结果显示UGT88A1在pH 9.0时最活跃。 UGT88A1合成槲皮素-4'- O-葡萄糖苷的最适温度为45℃,略低于合成槲皮素-3- O-葡萄糖苷的最适温度(50℃)。 UGT88A1 的一种突变体 V18R 是通过定点突变获得的,与野生型酶(0.36 mM 和 227.6)相比,它对槲皮素表现出更大的亲和力( K m 0.20 mM)和两倍的酶活性(552.3 mU/mg)。分别为 mU/mg)。通过分子模拟推断,可能的原因是UGT88A1的第18个氨基酸残基与底物槲皮素之间的距离变化。

更新日期:2019-05-09
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