6533b85efe1ef96bd12c08cc
RESEARCH PRODUCT
Alkylation at the active site of the D-3-hydroxybutyrate dehydrogenase (BDH), a membrane phospholipid-dependent enzyme, by 3-chloroacetyl pyridine adenine dinucleotide (3-CAPAD)
Norbert LatruffeM.s. El Kebbajsubject
AlkylationStereochemistryAffinity labelMitochondria LiverDehydrogenaseBiochemistryHydroxybutyrate DehydrogenaseMembrane LipidsAnimalsCoenzyme bindingCysteineBinding sitePhospholipidsBinding SitesAffinity labelingMolecular StructurebiologyChemistryActive siteAffinity LabelsGeneral MedicineNADRatsReagentLinear Modelsbiology.proteinNAD+ kinasedescription
The structure of the rat liver's D-3-hydroxybutyrate dehydrogenase (BDH) active site has been investigated using an affinity alkylating reagent, the 3-chloroacetyl pyridine adenine dinucleotide (3-CAPAD). This NAD+ analogue reagent strongly inactivates the enzyme following a concentration- and time-dependent process with a stoichiometry of approximately 1. The reagent reacts at the coenzyme binding site as revealed by the efficient protection by NADH. The effect of 3-CAPAD is stronger with the enzyme into its natural membrane environment than with the lipid-free purified apoBDH or with the reconstituted apoBDH-mitochondrial phospholipid complex. The pH-dependent effect on the inactivation process is in agreement with the participation of protons in the catalytic mechanism of BDH. Furthermore, this study exhibits the phospholipid activating role in BDH catalytic activation.
| year | journal | country | edition | language |
|---|---|---|---|---|
| 1997-01-01 | Biochimie |