[NAD+-kinase from Neurospora crassa: isolation and properties].
The NAD+ kinase (EC 2.7.1.23) of the filamentous fungus N. crassa is localized in cytosol. The activity in the dialyzed cell free extract has a pH optimum 8.3; it utilizes only ATP but not inorganic polyphosphates as a phosphoryl donor. A method for 200-fold purification of NAD+ kinase with a 20% yield has been developed. The procedure includes 105000 g centrifugation, fractionation with (NH4)2SO4, isoelectrofocusing in a Ultrodex layer and preparative electrophoresis in polyacrylamide gel. The molecular heterogeneity of NAD+ kinase was demonstrated by polyacrylamide gradient electrophoresis and by gel filtration through Sephadex G-200. The molecular weights of four individual forms of the enzyme are: 330000-338000, 305000-306000, 215000-229000 and 203000 Da. The Km values for the reaction catalyzed by purified NAD+ kinase for NAD+ and ATP are 3.0 X 10(-4) M and 0.9 X 10(-3) M, respectively.
D003035:Cobalt / Q000494:pharmacology; D003956:Dialysis; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008970:Molecular Weight; D009491:Neurospora / Q000201:enzymology*; D009492:Neurospora crassa / Q000201:enzymology*; D010766:Phosphorylation; D010770:Phosphotransferases / Q000032:analysis / Q000302:isolation & purification*; D017853:Phosphotransferases (Alcohol Group Acceptor); D013347:Subcellular Fractions / Q000201:enzymology; D013379:Substrate Specificity; D013438:Sulfhydryl Compounds / Q000032:analysis
D013438:Sulfhydryl Compounds; D003035:Cobalt; D010770:Phosphotransferases; D017853:Phosphotransferases (Alcohol Group Acceptor); C021844:NAD kinase; D008274:Magnesium
NAD+-kinaza Neurospora crassa: vydelenie i svoĭstva.