Crosstalk along the stalk: Dynamics of the interaction of subunits B and F in the A_1A_O ATP synthase of methanosarcina mazei G?1

摘要

The mechanism of coupling of ion pumping in the membrane-bound A_O sector with ATP synthesis in the A_3B_3 headpiece of the A_1 sector in the A_1A_O ATP synthase is a puzzle. Previously, crosstalk between the stalk and nucleotide-binding subunits F_(Mm) and B_(Mm) of the Methanosarcina mazei G?1 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_1A_O ATP synthase.