title: Structural and functional analysis of validoxylamine A 7'-phosphate synthase ValL involved in validamycin A biosynthesis.
abstract: Validamycin A (Val-A) is an effective antifungal agent widely used in Asian countries as crop protectant. Validoxylamine A, the core structure and intermediate of Val-A, consists of two C(7)-cyclitol units connected by a rare C-N bond. In the Val-A biosynthetic gene cluster in Streptomyces hygroscopicus 5008, the ORF valL was initially annotated as a validoxylamine A 7'-phosphate(V7P) synthase, whose encoded 497-aa protein shows high similarity with trehalose 6-phosphate(T6P) synthase. Gene inactivation of valL abolished both validoxylamine A and validamycin A productivity, and complementation with a cloned valL recovered 10% production of the wild-type in the mutant, indicating the involvement of ValL in validoxylamine A biosynthesis. Also we determined the structures of ValL and ValL/trehalose complex. The structural data indicates that ValL adopts the typical fold of GT-B protein family, featuring two Rossmann-fold domains and an active site at domain junction. The residues in the active site are arranged in a manner homologous to that of Escherichia coli (E.coli) T6P synthase OtsA. However, a significant discrepancy is found in the active-site loop region. Also noticeable structural variance is found around the active site entrance in the apo ValL structure while the region takes an ordered configuration upon binding of product analog trehalose. Furthermore, the modeling of V7P in the active site of ValL suggests that ValL might have a similar SNi-like mechanism as OtsA.
authors.affiliation: Department of Biotechnology, College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan, Hubei, China.
authors.forename: Lina
authors.initials: L
authors.lastname: Zheng
authors.forename: Xiang
authors.initials: X
authors.lastname: Zhou
authors.forename: Huaidong
authors.initials: H
authors.lastname: Zhang
authors.forename: Xiaofeng
authors.initials: X
authors.lastname: Ji
authors.forename: Lei
authors.initials: L
authors.lastname: Li
authors.forename: Lin
authors.initials: L
authors.lastname: Huang
authors.forename: Linquan
authors.initials: L
authors.lastname: Bai
authors.forename: Houjin
authors.initials: H
authors.lastname: Zhang
mesh_terms: D019883:Alkyl and Aryl Transferases / Q000737:chemistry / Q000235:genetics*; D000595:Amino Acid Sequence; D000935:Antifungal Agents / Q000737:chemistry; D002384:Catalysis; D020134:Catalytic Domain; D004926:Escherichia coli / Q000201:enzymology; D005816:Genetic Complementation Test; D005964:Glucosyltransferases / Q000737:chemistry; D007294:Inositol / Q000031:analogs & derivatives* / Q000096:biosynthesis / Q000737:chemistry; D008956:Models, Chemical; D008958:Models, Molecular; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D005810:Multigene Family; D009154:Mutation; D016366:Open Reading Frames; D017386:Sequence Homology, Amino Acid; D013302:Streptomyces / Q000235:genetics*; D014199:Trehalose / Q000737:chemistry
chemical_list: D000935:Antifungal Agents; C030790:validamycin A; C052534:validoxylamine A; D007294:Inositol; D014199:Trehalose; D005964:Glucosyltransferases; C023180:trehalose-6-phosphate synthase; D019883:Alkyl and Aryl Transferases; C574538:validoxylamine A 7'-phosphate synthase, Streptomyces hygroscopicus