FI-ATPase is a rotary motor enzyme in which a single ATP molecule drives a
120 degrees rotation of the central gamma subunit relative to the surroundi
ng alpha (3)beta (3) ring. Here, we show that the rotation of F-1-ATPase sp
ontaneously lapses into long (approximate to 30 s) pauses during steady-sta
te catalysis. The effects of ADP-Mg and mutation on the pauses, as well as
kinetic comparison with bulk-phase catalysis, strongly indicate that the pa
used enzyme corresponds to the inactive state of F-1-ATPase previously know
n as the ADP-Mg inhibited form in which F-1-ATPase fails to release ADP-Mg
from catalytic sites. The pausing position of the gamma subunit deviates fr
om the ATP-waiting position and is most likely the recently found intermedi
ate 90 degrees position.