Akere Aishat, Liu Qian, Wu Shibo, Hou Bingkai, Yang Min
The School of Pharmacy, University College London 29-39 Brunswick Square London WC1N 1AX UK
Key Laboratory of Plant Cell Engineering and Germplasm Innovation, Ministry of Education of China, School of Life Sciences, Shandong University Jinan 250100 China.
RSC Adv. 2018 Aug 24;8(53):30080-30086. doi: 10.1039/c8ra03947j.
In this report, we cloned and characterised four members of group H glycosyltransferases (GTs) by studying their substrate specificities and kinetics. The formation of products and possible glycosylation position was confirmed using MS/MS. The results revealed that 76E1 and 76E5 have broader donor specificity, including UDP-glucose (UDPGlc), UDP-galactose (UDPGal) and UDP--acetylglucosamine (UDPGlcNAc) with various flavonoids as acceptor substrates. Pseudo-single substrate kinetics data showed a relatively low , indicating a high affinity for substrate UDPGlc and also supported that 76E5 is more of a galactosyl and -acetylglucosamine transferase. Sequence alignment and site-directed mutagenesis studies indeed suggested that serine is a crucial residue in the UDPGlcNAc and UDPGal activity.
在本报告中，我们通过研究H组糖基转移酶（GTs）的底物特异性和动力学，克隆并鉴定了其中的四个成员。使用串联质谱（MS/MS）确认了产物的形成和可能的糖基化位置。结果表明，76E1和76E5具有更广泛的供体特异性，包括以各种黄酮类化合物作为受体底物的UDP-葡萄糖（UDPGlc）、UDP-半乳糖（UDPGal）和UDP-N-乙酰葡糖胺（UDPGlcNAc）。伪单底物动力学数据显示相对较低的Km值，表明对底物UDPGlc具有高亲和力，这也支持76E5更像是一种半乳糖基和N-乙酰葡糖胺转移酶。序列比对和定点诱变研究确实表明，丝氨酸是UDPGlcNAc和UDPGal活性中的关键残基。
