title: Expression, purification, crystallization and preliminary X-ray crystallographic analysis of fructose-1,6-bisphosphate aldolase from Escherichia coli.
abstract: Fructose-1,6-bisphosphate aldolase is one of the most important enzymes in the glycolytic pathway and catalyzes the reversible cleavage of fructose-1,6-bisphosphate to dihydroxyacetone phosphate and glyceraldehyde 3-phosphate. The full-length fbaB gene encoding fructose-1,6-bisphosphate aldolase class I (FBPA I) was cloned from Escherichia coli strain BL21. FBPA I was overexpressed in E. coli and purified. Biochemical analysis found that the optimum reaction temperature of FBPA I is 330.5 K and that the enzyme has a high temperature tolerance. Crystals of recombinant FBPA I were obtained by the sitting-drop vapour-diffusion technique in a condition consisting of 19 mg ml(-1) FBPA I in 0.1 M Tris pH 9.0, 10%(w/v) polyethylene glycol 8000 and diffracted to 2.0 Å resolution. The crystals belonged to the monoclinic space group C2, with unit-cell parameters a = 217.7, b = 114.9, c = 183.9 Å, β = 124.6°. The asymmetric unit of these crystals may contain ten molecules, giving a Matthews coefficient of 2.48 Å(3) Da(-1) and a solvent content of 50.5%.
authors.affiliation: State Key Laboratory of Genetic Engineering, School of Life Sciences, Fudan University, Shanghai 200433, People's Republic of China.
authors.forename: Li
authors.initials: L
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Genetic Engineering, School of Life Sciences, Fudan University, Shanghai 200433, People's Republic of China.
authors.forename: Zheng
authors.initials: Z
authors.lastname: Guo
authors.affiliation: State Key Laboratory of Genetic Engineering, School of Life Sciences, Fudan University, Shanghai 200433, People's Republic of China.
authors.forename: Jing
authors.initials: J
authors.lastname: Huang
authors.affiliation: State Key Laboratory of Genetic Engineering, School of Life Sciences, Fudan University, Shanghai 200433, People's Republic of China.
authors.forename: Meiruo
authors.initials: M
authors.lastname: Liu
authors.affiliation: State Key Laboratory of Genetic Engineering, School of Life Sciences, Fudan University, Shanghai 200433, People's Republic of China.
authors.forename: Yuandong
authors.initials: Y
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Genetic Engineering, School of Life Sciences, Fudan University, Shanghai 200433, People's Republic of China.
authors.forename: Chaoneng
authors.initials: C
authors.lastname: Ji
mesh_terms: D003460:Crystallization; D018360:Crystallography, X-Ray; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification; D005634:Fructose-Bisphosphate Aldolase / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification; D013696:Temperature
chemical_list: D029968:Escherichia coli Proteins; D005634:Fructose-Bisphosphate Aldolase
keywords: Ec-FBPA I; fructose-1,6-bisphosphate aldolase; thermostability