sup 1 H and sup 31 P nuclear magnetic resonance and kinetic studies of the active site structure of chloroplast CF sub 1 ATP synthase
- Univ. of Virginia, Charlottesville (USA)
The interaction of nucleotides and nucleotide analogues and their complexes with Mn{sup 2+} bound to both the latent and dithiothreitol-activated CF{sub 1} ATP synthase has been examined by means of steady-state kinetics, water proton relaxation rate (PRR) measurements, and {sup 1}H and {sup 31}P nuclear relaxation measurements. Titration of both the latent and activated Mn{sup 2+}-CF{sub 1} complexes with ATP, ADP, P{sub i}, Co(NH{sub 3}){sub 4}ATP, Co(NH{sub 3}){sub 4}ADP, and Co(NH{sub 3}){sub 4}AMPPCP leads to increases in the water relaxation enhancement, consistent with enhanced metal binding and a high ternary complex enhancement. Steady-state kinetic studies are consistent with competitive inhibition of CF{sub 1} by Co(NH{sub 3}){sub 4}AMPPCP with respect to CaATP. {sup 1}H and {sup 31}P nuclear relaxation measurements in solutions of CF{sub 1} and Co(NH{sub 3}){sub 4}AMPPCP were used to determine the conformation of the bound substrate analogue and the arrangement with respect to this structure of high- and low-affinity sites for Mn{sup 2+}. The bound nucleotide analogue adopts a bent conformation, with the low-affinity sites for Mn{sup 2+}. The bound nucleotide analogue adopts a bent conformation, with the low-affinity Mn{sup 2+} site situated between the adenine and triphosphate moieties and the high-affinity metal site located on the far side of the triphosphate chain. The low-affinity metal forms a distorted inner-sphere complex with the {beta}-P and {gamma}-P of the substrate. The distances from Mn{sup 2+} to the triphosphate chain are too large for first coordination sphere complexes but are appropriate for second-sphere complexes involving, for example, intervening hydrogen-bonded water molecules or residues from the protein.
- OSTI ID:
- 6290743
- Journal Information:
- Biochemistry; (USA), Vol. 29:26; ISSN 0006-2960
- Country of Publication:
- United States
- Language:
- English
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ATP-ASE
BIOCHEMICAL REACTION KINETICS
CHLOROPHYLL-BINDING PROTEINS
NUCLEAR MAGNETIC RESONANCE
ALGORITHMS
CATIONS
CHLOROPLASTS
ENZYME ACTIVITY
MANGANESE COMPOUNDS
MOLECULAR STRUCTURE
NUCLEOTIDES
PHOSPHORUS 31
PHOSPHORUS 32
PROTONS
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ACID ANHYDRASES
BARYONS
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
CELL CONSTITUENTS
CHARGED PARTICLES
DAYS LIVING RADIOISOTOPES
ELEMENTARY PARTICLES
ENZYMES
FERMIONS
HADRONS
HYDROLASES
IONS
ISOTOPES
KINETICS
LIGHT NUCLEI
MAGNETIC RESONANCE
MATHEMATICAL LOGIC
NUCLEI
NUCLEONS
ODD-EVEN NUCLEI
ODD-ODD NUCLEI
ORGANIC COMPOUNDS
PHOSPHOHYDROLASES
PHOSPHORUS ISOTOPES
PHOTOSYNTHETIC REACTION CENTERS
PROTEINS
RADIOISOTOPES
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TRANSITION ELEMENT COMPOUNDS
550601* - Medicine- Unsealed Radionuclides in Diagnostics