title: Structural and enzymatic analyses of a glucosyltransferase Alr3699/HepE involved in Anabaena heterocyst envelop polysaccharide biosynthesis.
abstract: Formation of the heterocyst envelope polysaccharide (HEP) is a key process for cyanobacterial heterocyst differentiation. The maturation of HEP in Anabaena sp. strain PCC 7120 is controlled by a gene cluster termed HEP island in addition to an operon alr3698-alr3699, which encodes two putative proteins termed Alr3698/HepD and Alr3699/HepE. Here we report the crystal structures of HepE in the apo-form and three complex forms that bind to UDP-glucose (UDPG), UDP&glucose, and UDP, respectively. The overall structure of HepE displays a typical GT-B fold of glycosyltransferases, comprising two separate β/α/β Rossmann-fold domains that form an inter-domain substrate-binding crevice. Structural analyses combined with enzymatic assays indicate that HepE is a glucosyltransferase using UDPG as a sugar donor. Further site-directed mutageneses enable us to assign the key residues that stabilize the sugar donor and putative acceptor. Based on the comparative structural analyses, we propose a putative catalytic cycle of HepE, which undergoes "open-closed-open" conformational changes upon binding to the substrates and release of products. These findings provide structural and catalytic insights into the first enzyme involved in the HEP biosynthesis pathway.
authors.affiliation: Hefei National Laboratory for Physical Sciences at the Microscale and School of Life Sciences, University of Science and Technology of China, Hefei, Anhui 230027, People's Republic of China.
authors.forename: Xue-Ping
authors.initials: XP
authors.lastname: Wang
authors.affiliation: Hefei National Laboratory for Physical Sciences at the Microscale and School of Life Sciences, University of Science and Technology of China, Hefei, Anhui 230027, People's Republic of China.
authors.forename: Yong-Liang
authors.initials: YL
authors.lastname: Jiang
authors.affiliation: Hefei National Laboratory for Physical Sciences at the Microscale and School of Life Sciences, University of Science and Technology of China, Hefei, Anhui 230027, People's Republic of China.
authors.forename: Ya-Nan
authors.initials: YN
authors.lastname: Dai
authors.affiliation: Hefei National Laboratory for Physical Sciences at the Microscale and School of Life Sciences, University of Science and Technology of China, Hefei, Anhui 230027, People's Republic of China.
authors.forename: Wang
authors.initials: W
authors.lastname: Cheng
authors.affiliation: Hefei National Laboratory for Physical Sciences at the Microscale and School of Life Sciences, University of Science and Technology of China, Hefei, Anhui 230027, People's Republic of China zcz@ustc.edu.cn cyxing@ustc.edu.cn.
authors.forename: Yuxing
authors.initials: Y
authors.lastname: Chen
authors.affiliation: Hefei National Laboratory for Physical Sciences at the Microscale and School of Life Sciences, University of Science and Technology of China, Hefei, Anhui 230027, People's Republic of China zcz@ustc.edu.cn cyxing@ustc.edu.cn.
authors.forename: Cong-Zhao
authors.initials: CZ
authors.lastname: Zhou
mesh_terms: D017033:Anabaena / Q000201:enzymology* / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D005964:Glucosyltransferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D005810:Multigene Family; D011135:Polysaccharides, Bacterial / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics; D000072417:Protein Domains; D014532:Uridine Diphosphate Glucose / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D011135:Polysaccharides, Bacterial; D005964:Glucosyltransferases; D014532:Uridine Diphosphate Glucose
keywords: GT-B fold; crystal structure; cyanobacteria; glycosyltransferase; heterocyst envelope polysaccharide