title: Isolation of the bifunctional enzyme lysine 2-oxoglutarate reductase-saccharopine dehydrogenase from Phaseolus vulgaris.
abstract: Lysine is catabolyzed by the bifunctional enzyme lysine 2-oxoglutarate reductase-saccharopine dehydrogenase (LOR-SDH) in both animals and plants. LOR condenses lysine and 2-oxoglutarate into saccharopine, using NADPH as cofactor and SDH converts saccharopine into alpha-aminoadipate delta-semialdehyde and glutamic acid, using NAD as cofactor. The distribution pattern of LOR and SDH among different tissues of Phaseolus vulgaris was determined. The hypocotyl contained the highest specific activity, whereas in seeds the activities of LOR and SDH were below the limit of detection. Precipitation of hypocotyl proteins with increasing concentrations of PEG 8000 revealed one broad peak of SDH activity, indicating that two isoforms may be present, a bifunctional LOR-SDH and possibly a monofunctional SDH. During the purification of the hypocotyl enzyme, the LOR activity proved to be very unstable, following ion-exchange chromatography. Depending on the purification procedure, the protein eluted as a monomer of 91-94 kDa containing only SDH activity, or as a dimer of 190 kDa with both, LOR and SDH activities, eluting together.
authors.affiliation: Departamento de Fisiologia Vegetal, Instituto de Biologia, Universidade Estadual de Campinas, Campinas, Brasil. slima@diabetes.ucsf.edu
authors.forename: S T
authors.initials: ST
authors.lastname: Cunha Lima
authors.forename: R A
authors.initials: RA
authors.lastname: Azevedo
authors.forename: L G
authors.initials: LG
authors.lastname: Santoro
authors.forename: S A
authors.initials: SA
authors.lastname: Gaziola
authors.forename: P J
authors.initials: PJ
authors.lastname: Lea
mesh_terms: D008970:Molecular Weight; D027805:Phaseolus / Q000201:enzymology*; D012443:Saccharopine Dehydrogenases / Q000737:chemistry / Q000302:isolation & purification*
chemical_list: D012443:Saccharopine Dehydrogenases; C029496:saccharopine dehydrogenase (NADP+, lysine-forming)