N,N′-dicyclohexylcarbodiimide and 4-chloro-7-nitrobenzofurazan bind to different β subunits of the F 1 ATPase of Escherichia coli

Abstract

The fluorescent thiol reagent 2-(4′-iodoacetamidoanilino)naphthalene-6-sulfonic acid (IAANS) labels the γ, δ, and one of the three β subunits of the F 1 ATPase from Escherichia coli (ECF 1). This is the same β subunit which incorporates 4-chloro-7-nitrobenzofurazan (Nbf) [H. Stan-Lotter and P. D. Bragg (1986) Eur. J. Biochem. 154, 321–327] . After inactivation of ECF 1 with N,N′-dicyclohexylcarbodiimide (DCCD), IAANS labels in addition to the β, γ, and δ subunits also the α subunit. This suggests a conformational change of ECF 1 upon binding of DCCD. The β subunit which incorporates DCCD does not bind IAANS. Likewise, IAANS-modified ECF 1 does not incorporate DCCD into the same β subunit. It is concluded that DCCD and Nbf bind to different β subunits. Since neither of these reagents binds to that β subunit which can be crosslinked to the ϵ subunit by 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide, these data show that there is a difference in the chemical reactivity of each of the three β subunits of ECF 1, despite their identical primary structures. This suggests that there is an asymmetry in the F 1 molecule.