Interactions of Neurospora crassa plasma membrane H/sup +/-ATPase with N-(ethoxycarbonyl)-2-ethoxy-1,2-dihydroquinoline
The carboxyl group activating reagent N-(ethoxycarbonyl)-2-ethoxy-1,2-dihydroquinoline (EEDQ) interacts with the Neurospora plasma membrane H/sup +/-ATPase in at least three different ways. This reagent irreversibly inhibits ATP hydrolysis with kinetics that are pseudo-first-order at several concentrations of EEDQ, and an appropriate transform of these data suggests that 1 mol of EEDQ inactivates 1 mol of the H/sup +/-ATPase. Inhibition probably involves activation of an ATPase carboxyl group followed by a nucleophilic attack by a vicinal nucleophilic functional group in the ATPase polypeptide chain, resulting in an intra-molecular cross-link. The enzyme is protected against EEDQ inhibition by MgATP in the presence of vanadate, a combination of ligands that has previously been shown to lock the H/sup +/-APTase in a conformation that presumably resembles the transition states of the enzyme phosphorylation and dephosphorylation reactions, but is not protected by the substrate analogue MgADP. The ATPase is also labeled by the exogenous nucleophile (/sup 14/C)glycine ethyl ester in an EEDQ-dependent reaction, and the labeling is diminished in the presence of MgATP plus vanadate. In a third type of interaction, EEDQ mediates the specific cross-linking of ATPase monomers with some other membrane protein, possibly another ATPase monomer, leading to the formation of a product with an apparent molecular weight of about 260,000.
- Research Organization:
- Univ. of North Carolina, Chapel Hill (USA)
- OSTI ID:
- 6583011
- Journal Information:
- Biochemistry; (United States), Journal Name: Biochemistry; (United States) Vol. 25:13; ISSN BICHA
- Country of Publication:
- United States
- Language:
- English
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59 BASIC BIOLOGICAL SCIENCES
ACID ANHYDRASES
ATP-ASE
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BIOCHEMICAL REACTION KINETICS
CARBON 14
CARBON ISOTOPES
CELL CONSTITUENTS
CELL MEMBRANES
ENZYMES
EVEN-EVEN NUCLEI
FUNGI
HYDROLASES
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
LIGHT NUCLEI
MEMBRANES
NEUROSPORA
NUCLEI
PHOSPHOHYDROLASES
PLANTS
RADIOISOTOPES
REACTION KINETICS
STRUCTURAL CHEMICAL ANALYSIS
TRACER TECHNIQUES
YEARS LIVING RADIOISOTOPES