Nucleotide binding sites on beef heart mitochondrial F1-ATPase. Cooperative interactions between sites and specificity of noncatalytic sites
Milgrom, Y.M.; Cross, R.L.
Journal of Biological Chemistry 268(31): 23179-23185
1993
ISSN/ISBN: 0021-9258
PMID: 8226836
Accession: 009113087
We have studied the properties of beef heart mitochondrial F-1 having inhibitory MgADP bound at one of the three catalytic sites and various levels of occupancy of the three noncatalytic nucleotide sites including zero, two, or three ADP/ATPs or two ADP/ATP plus one GTP. The properties examined include the rate of MgATP-dependent reactivation and the rate of increase in the fraction of F-1 containing transiently bound intermediates. For each form of the enzyme tested, the rate of reactivation closely paralleled the rate of increase in the level of bound intermediates, indicating that when one catalytic site on F-1 is blocked by inhibitory MgADP, the remaining two sites are incapable of residual uni- or bi-site activity. It was also found that the stability of the MgADP-inhibited complex decreases with full occupancy of the noncatalytic sites. This demonstrates that the noncatalytic sites modulate the properties of catalytic sites. Finally, it was found that the noncatalytic sites on mitochondrial F-1 do not, as has long been believed, bind adenine nucleotides exclusively. Evidence is presented that both GTP and PP-i bind tightly at noncatalytic sites.