title: NADP-linked malic enzyme. Purification from maize leaves, Mr and subunit composition.
abstract: 1. The isolation of NADP-linked malic enzyme (EC 1.1.1.40) from maize leaves is described, together with studies of its Mr and subunit composition. 2. The enzyme was purified to apparent homogeneity by affinity chromatography on N6-aminohexyl-2',5'-bisphosphoadenosine-agarose, gel filtration with Sephadex G-100 and ion-exchange chromatography on DEAE-Sephadex A-50. A purification of 140-fold with a 30% yield was obtained. 3. A detailed study of the Mr by several methods revealed the existence of different Mr forms in solution. 4. In the presence of dithiothreitol the enzyme appears to be present in triethanolamine buffer, pH 7.5, as a tetramer with a subunit Mr of 60,000 and an S20,w of 10.75 S. 5. In phosphate buffer, pH 7.0, it seems to be a dimer of Mr 120,000 with an S20,w of 7.95 S. 6. In the absence of dithiothreitol, lower-Mr forms were detected by sedimentation-equilibrium and sedimentation-velocity studies in triethanolamine buffer. 7. Results from gel filtration gave Mr values of about 340,000 in both buffers.
authors.affiliation: Department of Biochemistry, University of Oxford, U.K.
authors.forename: M S
authors.initials: MS
authors.lastname: Thorniley
authors.forename: K
authors.initials: K
authors.lastname: Dalziel
mesh_terms: D002499:Centrifugation, Density Gradient; D002852:Chromatography, Ion Exchange; D004229:Dithiothreitol / Q000494:pharmacology; D046911:Macromolecular Substances; D008291:Malate Dehydrogenase / Q000302:isolation & purification*; D008970:Molecular Weight; D003313:Zea mays / Q000201:enzymology*
chemical_list: D046911:Macromolecular Substances; D008291:Malate Dehydrogenase; C031917:malate dehydrogenase (oxaloacetate-decarboxylating) (NADP+); D004229:Dithiothreitol