title: Crystallization and preliminary X-ray analysis of (1R,6R)-2-succinyl-6-hydroxy-2,4-cyclohexadiene-1-carboxylate (SHCHC) synthase (MenD) from Escherichia coli.
abstract: (1R,6R)-2-Succinyl-6-hydroxy-2,4-cyclohexadiene-1-carboxylate (SHCHC) synthase, also called MenD, participates in the menaquinone (vitamin K2) biosynthetic pathway. The enzyme is a part of the superfamily of ThDP-dependent enzymes; however, it is the only enzyme known to catalyze a Stetter-like 1,4-addition of a ThDP adduct to the beta-carbon of an unsaturated carboxylate. This is the first reported crystallization of the apoenzyme and holoenzyme forms of MenD. The apoenzyme crystals were obtained by sitting-drop vapour diffusion with 70% MPD. However, the crystals were too small to collect diffraction data and a search for better conditions was not successful. Single crystals of the holoenzyme with ThDP and Mn2+ as cofactors were obtained by the hanging-drop vapour-diffusion method with 35% ethylene glycol as precipitant. Diffraction data were collected on a cryocooled crystal to a resolution of 2.0 A at BioCARS, Advanced Photon Source (APS), Chicago, IL, USA. The crystal was found to belong to space group P2(1)2(1)2(1), with unit-cell parameters a = 106.86, b = 143.06, c = 156.85 A, alpha = beta = gamma = 90 degrees.
authors.affiliation: Department of Chemistry, University of Saskatchewan, Saskatoon, Saskatchewan S7N 5C9, Canada.
authors.forename: Edyta A L
authors.initials: EA
authors.lastname: Sieminska
authors.forename: Andrea
authors.initials: A
authors.lastname: Macova
authors.forename: David R J
authors.initials: DR
authors.lastname: Palmer
authors.forename: David A R
authors.initials: DA
authors.lastname: Sanders
mesh_terms: D001051:Apoenzymes / Q000737:chemistry; D003460:Crystallization; D003510:Cyclohexanes; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D020035:Holoenzymes / Q000737:chemistry; D012459:Salicylates / Q000737:chemistry* / Q000378:metabolism; D013386:Succinates / Q000737:chemistry* / Q000378:metabolism
chemical_list: C087010:2-succinyl-6-hydroxycyclohexa-2,4-diene-1-carboxylic acid; D001051:Apoenzymes; D003510:Cyclohexanes; D020035:Holoenzymes; D012459:Salicylates; D013386:Succinates