Differences between two tight ADP binding sites of the chloroplast coupling factor 1 and their effects on ATPase activity.
Purified chloroplast ATP synthase (CF1) contains 1.2-2 mol of tightly bound ADP/mol of enzyme that resists removal by gel filtration or dialysis. CF1 was depleted of its endogenous nucleotide by treatment with alkaline phosphatase. Tightly bound nucleotide was demonstrated not to have an essential structural role. CF1 depleted of endogenous nucleotide retains its ability to catalyze Ca2+- and Mg2+-dependent ATPase activity and is not more sensitive to cold inactivation than untreated CF1. 2'(3')-O-Trinitrophenyladenosine 5'-diphosphate (TNP-ADP) binds tightly to two sites on nucleotide-depleted CF1, binding to either site at a faster rate than that of exchange of bound nucleotide for medium nucleotide. The nucleotide-depleted enzyme binds about one additional mol of TNP-ADP/mol of CF1, indicating that there is a tight TNP-ADP binding site that does not exchange readily with medium nucleotide. It is MgADP in this nonexchanging site, not the easily exchanging ADP binding site, that is responsible for the MgADP-induced inhibition of the ATPase activity. The rate of exchange of tightly bound ADP from CF1 matches the rate at which the Mg2+ATPase activity of CF1 is activated but is not itself responsible for the activation.
Digel
J G
JG
Department of Biology, The Johns Hopkins University, Baltimore, Maryland 21218, USA.
Kishinevsky
A
A
Ong
A M
AM
McCarty
R E
RE
D000244:Adenosine Diphosphate / Q000031:analogs & derivatives / Q000378:metabolism*; D001665:Binding Sites; D000252:Calcium-Transporting ATPases / Q000378:metabolism*; D002736:Chloroplasts / Q000201:enzymology*; D002851:Chromatography, High Pressure Liquid; D003080:Cold Temperature; D008274:Magnesium / Q000378:metabolism; D006180:Proton-Translocating ATPases / Q000378:metabolism*; D018724:Spinacia oleracea; D013816:Thermodynamics
D000244:Adenosine Diphosphate; D006180:Proton-Translocating ATPases; D000252:Calcium-Transporting ATPases; D008274:Magnesium