Dissociation-reconstitution experiments support the presence of two catalytic beta-subunits in mitochondrial F1.

Mitochondrial F1, inactivated to various extents with 4-chloro-7-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl), was dissociated with LiCl and reconstituted after removal of the salt. This procedure resulted in a reactivation that corresponded with a reactivation theoretically expected on the basis of the assumption that the reassociation of beta-subunits into native F1 molecules is random and that two out of the three beta-subunits are directly involved in catalysis. Repeated inactivation of such reactivated F1, followed by the same dissociation-association procedure, resulted in similar data. After inactivation of F1 by covalent binding of 2-N-AT(D)P to one catalytic site, no reactivation upon dissociation-reassociation was obtained due to the fact that such modified F1 did not dissociate under the experimental conditions used.