Characterization of the catalytic and noncatalytic ADP binding sites of the F1-ATPase from the thermophilic bacterium, PS3
Two classes of ADP binding sites at 20 degrees C have been characterized in the F1-ATPase from the thermophilic bacterium, PS3 (TF1). One class is comprised of three sites which saturate with (/sup 3/H)ADP in less than 10 s with a Kd of 10 microM which, once filled, exchange rapidly with medium ADP. The binding of ADP to these sites is dependent on Mg2+. (/sup 3/H)ADP bound to these sites is removed by repeated gel filtrations on centrifuge columns equilibrated with ADP free medium. The other class is comprised of a single site which saturates with (/sup 3/H)ADP in 30 min with a Kd of 30 microM. (/sup 3/H)ADP bound to this site does not exchange with medium ADP nor does it dissociate on gel filtration through centrifuge columns equilibrated with ADP free medium. Binding of (/sup 3/H)ADP to this site is weaker in the presence of Mg2+ where the Kd for ADP is about 100 microM. (/sup 3/H)ADP dissociated from this site when ATP plus Mg2+ was added to the complex while it remained bound in the presence of ATP alone or in the presence of ADP, Pi, or ADP plus Pi with or without added Mg2+. Significant amounts of ADP in the 1:1 TF1.ADP complex were converted to ATP in the presence of Pi, Mg2+, and 50% dimethyl sulfoxide. Enzyme-bound ATP synthesis was abolished by chemical modification of a specific glutamic acid residue by dicyclohexylcarbodiimide, but not by modification of a specific tyrosine residue with 7-chloro-4-nitrobenzofurazan. Difference circular dichroism spectra revealed that the three Mg2+ -dependent, high affinity ADP binding sites that were not stable to gel filtration were on the alpha subunits and that the single ADP binding site that was stable to gel filtration was on one of the three beta subunits.
- Research Organization:
- Tokyo Institute of Technology, Kanagawa-ken, Japan
- OSTI ID:
- 5691020
- Journal Information:
- J. Biol. Chem.; (United States), Vol. 13
- Country of Publication:
- United States
- Language:
- English
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63 RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTANT EFFECTS ON LIVING ORGS. AND BIOL. MAT.
ADP
BIOCHEMICAL REACTION KINETICS
RECEPTORS
AFFINITY
ATP-ASE
BACTERIA
BENZOFURANS
DICHROISM
DMSO
HEAT
IMIDES
MAGNESIUM
TRACER TECHNIQUES
TRITIUM COMPOUNDS
TYROSINE
ACID ANHYDRASES
ALKALINE EARTH METALS
AMINO ACIDS
CARBOXYLIC ACIDS
ELEMENTS
ENERGY
ENZYMES
FURANS
HETEROCYCLIC COMPOUNDS
HYDROLASES
HYDROXY ACIDS
ISOTOPE APPLICATIONS
KINETICS
LABELLED COMPOUNDS
MEMBRANE PROTEINS
METALS
MICROORGANISMS
NUCLEOTIDES
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
ORGANIC OXYGEN COMPOUNDS
ORGANIC SULFUR COMPOUNDS
PHOSPHOHYDROLASES
PROTEINS
REACTION KINETICS
SULFOXIDES
550201* - Biochemistry- Tracer Techniques
560302 - Chemicals Metabolism & Toxicology- Microorganisms- (-1987)