ASPECTS OF SUBUNIT INTERACTIONS IN THE CHLOROPLAST ATP SYNTHASE .2. CHARACTERIZATION OF A CHLOROPLAST COUPLING FACTOR 1-SUBUNIT-III COMPLEX FROM SPINACH THYLAKOIDS

A complex between chloroplast-coupling factor 1 (CF1) and subunit III of the membrane-spanning portion of the chloroplast ATP synthase (CF0), isolated as described in the accompanying paper (C.M. Wetzel and R.E. McCarty [1993] Plant Physiol 102: 241-249), has been further characterized. A comparison of the ATPase activities of CF1, CF1-subunit III, and the chloroplast ATP synthase (CF1-CF0) holoenzyme revealed that the properties of CF1-subunit III more closely resemble those of CF1-CF0 than those of CF1. In particular, the Ca2+-ATPase activity after reduction of the enzyme with dithiothreitol was much lower in CF1-subunit III and CF1-CF0 than in CF1, suggesting that the association of the inhibitory epsilon subunit is tightened by the presence of either CF0 or subunit III. Cold stability is a property of CF1-CF0 in thylakoid membranes. The ATPase activity of CF1 incubated in the cold in the presence of asolectin liposomes was lost more rapidly than that of either CF1-subunit III or CF1-CF0 incorporated into liposomes. Removal of the epsilon subunit from all three preparations resulted in marked stimulation of their ATPase activity. Although subunit III was also removed during depletion of the epsilon subunit, it is not known whether the two subunits interact directly. CF1 deficient in the epsilon subunit binds to liposomes containing either subunit III or CF0. Taken together, these results provide evidence that the association of CF1 and subunit III of CF0 is specific and may play a role in enzyme regulation.