title: Improved methods for the purification of enzymes of the folate pathway in Escherichia coli. I. Chromatographic methods.
abstract: A sequence of chromatographic procedures is described for the isolation of three consecutive enzymes of the folate pathway in Escherichia coli: hydroxymethyldihydropteridine pyrophosphokinase (E.C. 2.7.6.3) (I), 7,8-dihydropteroate synthase (E.C. 2.5.1.15) (II) and 7,8-dihydrofolate reductase, (E.C. 1.5.1.3) (III). Starting with the crude extract, ion-exchange chromatography on a DEAE-Sepharose CL-6B column with a salt gradient completely separated I, II and III. I and II were further purified by hydrophobic-interaction chromatography on Phenyl-Sepharose CL-4B, followed by size-exclusion chromatography on Ultrogel AcA 54. For III only size-exclusion chromatography was used. The overall enrichment factors, on the basis of protein, were 13,700-fold for I, 280-fold for II and 500-fold for III. Bacterial batches of more than 500 g were handled.
authors.affiliation: Forschungsinstitut Borstel, Medizinisch-Pharmazeutische Chemie, F.R.G.
authors.forename: R
authors.initials: R
authors.lastname: Bartels
authors.forename: L
authors.initials: L
authors.lastname: Bock
mesh_terms: D002852:Chromatography, Ion Exchange / Q000379:methods*; D004094:Dihydropteroate Synthase / Q000302:isolation & purification*; D017854:Diphosphotransferases; D004926:Escherichia coli / Q000201:enzymology*; D005492:Folic Acid / Q000378:metabolism*; D010770:Phosphotransferases / Q000302:isolation & purification*; D013762:Tetrahydrofolate Dehydrogenase / Q000302:isolation & purification*
chemical_list: D005492:Folic Acid; D013762:Tetrahydrofolate Dehydrogenase; D004094:Dihydropteroate Synthase; D010770:Phosphotransferases; D017854:Diphosphotransferases; C019684:2-amino-4-hydroxy-6-hydroxymethyldihydropteridine pyrophosphokinase