Brain pyridoxal kinase. Purification and characterization.
Pyridoxal kinase has been purified 9000-fold from sheep brain. The purification procedure involves ammonium sulphate fractionation, DEAE-cellulose chromatography, affinity chromatography and Sephadex G-100 gel filtration. The final chromatography step yields a homogeneous preparation of high specific activity with a pI of 5. The molecular mass of the native enzyme was estimated to be approximately 80 kDa by 10-25% gradient polyacrylamide gel electrophoresis and Sephadex G-200 gel filtration. The subunit molecular mass was determined by sodium dodecyl sulphate (SDS)/polyacrylamide gel electrophoresis to be 40 kDa compared with a series of molecular mass standards. This indicates that pyridoxal kinase is a dimeric enzyme. Further results obtained from electron microscopy, using a negative staining technique, provide evidence that pyridoxal kinase exists as a dispherical subunit structure.
D000818:Animals; D001921:Brain / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D008854:Microscopy, Electron; D008970:Molecular Weight; D010766:Phosphorylation; D010770:Phosphotransferases / Q000302:isolation & purification*; D011731:Pyridoxal Kinase / Q000032:analysis / Q000302:isolation & purification*; D012756:Sheep
D010770:Phosphotransferases; D011731:Pyridoxal Kinase