Abstract

The mechanism of coupling of ion pumping in the membrane-bound A(O) sector with ATP synthesis in the A(3)B(3) headpiece of the A(1) sector in the A(1)A(O) ATP synthase is a puzzle. Previously, crosstalk between the stalk and nucleotide-binding subunits F(Mm) and B(Mm) of the Methanosarcina mazei Gö1 A-ATP synthase has been observed by nucleotide-dependent cross-link formation of both subunits inside the enzyme. The recently determined NMR solution structure of F(Mm) depicts the protein as a two-domain structure, with a well-folded N-terminus having 78 residues and a flexible C-terminal part (residues 79-101), proposed to become structured after binding to its partner, B(Mm). Here, we detail the crucial interactions between subunits B(Mm) and F(Mm) by determining the NMR structure of the very C-terminus of F(Mm), consisting of 20 residues and hereafter termed F(Mm(81-101)), and performing molecular dynamics simulations on the resulting structure. These data demonstrate that the flexibility of the C-terminus enables F(Mm) to switch between an elongated and retracted state. Docking and MD in conjunction with previously conducted and published NMR results, biochemical cross-linking, and fluorescence spectroscopy data were used to reconstruct a model of a B(Mm)-F(Mm) assembly. The model of the B(Mm)-F(Mm) complex shows the detailed interactions of helices 1 and 2 of the C-terminal domain of B(Mm) with the C-terminal residues of F(Mm). Movements of both helices of B(Mm) accommodate the incoming C-terminus of F(Mm) and connect the events of ion pumping and nucleotide binding in the A(1)A(O) ATP synthase.