Purification, crystallization and preliminary X-ray analysis of native and selenomethionine class I tagatose-1,6-bisphosphate aldolase from Streptococcus pyogenes.
Tagatose-1,6-bisphosphate aldolase (EC 4.1.2.40) is situated at the branching of the tagatose-6-phosphate and Embden-Meyerhof-Parnas (glycolysis) metabolic pathways, where it catalyzes the reversible cleavage of tagatose-1,6-bisphosphate to dihydroxyacetone phosphate and glyceraldehyde 3-phosphate. The recombinant protein from Streptococcus pyogenes was overexpressed in Escherichia coli in its native and selenomethionine-derivative forms and purified using ion-exchange and hydrophobic interaction chromatography. Orthorhombic crystals suitable for structural analysis were obtained by the hanging-drop vapour-diffusion method for both isoforms. The crystals belong to space group P2(1)2(1)2(1), with unit-cell parameters a = 63.7, b = 108.1, c = 238.7 A for the native form and a = 64.1, b = 108.3, c = 239.8 A for the selenomethionine derivative. The asymmetric unit contains four protomers, corresponding to a crystal volume per protein weight (V(M)) of 2.8 A(3) Da(-1) and a solvent content of 56% by volume.
Liotard
Brigitte
B
Département de Biochemie, Université de Montréal, CP 6128, Succursale Centre-Ville, Montréal H3C 3J7, Canada. brigitte.liotard@umontreal.ca
Sygusch
Jurgen
J
D000446:Aldehyde-Lyases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D003460:Crystallization; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000235:genetics; D006601:Hexoses / Q000737:chemistry / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D012645:Selenomethionine / Q000737:chemistry; D013297:Streptococcus pyogenes / Q000201:enzymology* / Q000235:genetics
D001426:Bacterial Proteins; D006601:Hexoses; D011993:Recombinant Fusion Proteins; D012645:Selenomethionine; D000446:Aldehyde-Lyases; C023093:tagatose 1,6-diphosphate aldolase; C030192:tagatose