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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)
- Source :
- Biochimie. 79:37-42
- Publication Year :
- 1997
- Publisher :
- Elsevier BV, 1997.
-
Abstract
- 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.
- Subjects :
- Alkylation
Stereochemistry
Affinity label
Mitochondria, Liver
Dehydrogenase
Biochemistry
Hydroxybutyrate Dehydrogenase
Membrane Lipids
Animals
Coenzyme binding
Cysteine
Binding site
Phospholipids
Binding Sites
Affinity labeling
Molecular Structure
biology
Chemistry
Active site
Affinity Labels
General Medicine
NAD
Rats
Reagent
Linear Models
biology.protein
NAD+ kinase
Subjects
Details
- ISSN :
- 03009084
- Volume :
- 79
- Database :
- OpenAIRE
- Journal :
- Biochimie
- Accession number :
- edsair.doi.dedup.....a1584ff027c0905c3d8f6fce4e6129af