Question

1) Describe an experimental system used to confirm that proton gradients can power ATP synthesis.

2) Describe the mechanism of ATP synthesis by ATP synthase

Answer 1

1) Boyer predicted [1] that catalysis requires sequential involvement of the three catalytic sites, each of which changes its binding affinity for substrates and products as it proceeds through a cyclical mechanism, the ‘binding change mechanism’. As applied to ATP synthesis, Boyer hypothesized that each site first acts to bind ADP and Pi, then acts to chemically synthesize ATP, then opens to release the ATP, and that all three activities are ongoing simultaneously at the three different sites. The mechanism includes proposals that (a) only one site is catalytically active at any one moment in time, (b) the chemical reaction occurs reversibly (K_eq∼1) at this site, (c) energy input is utilized to bind substrates ADP and Pi into the catalytic sites and to achieve release of ATP, not for actual ADP-O to P bond formation. When the X-ray structures first showed catalytic sites asymmetry, and revolving actin filament videos established rotation at least during ATP hydrolysis, there was an initial rush to conclude that Boyer’s mechanism was proven.

2) The source of energy for ATP synthesis is an electrochemical gradient of protons generated initially by electron transfer complexes across the mitochondrial membrane. Functionally, ATP synthase consists of three parts, (1) a motor in the membrane that converts electrochemical ion gradient energy into subunit rotation, (2) a rotating transmission device, the ‘rotor stalk’, which transmits the energy over a distance of greater than 100 Å to the catalytic sites, and (3) the catalytic sites, three in number, where the mechanical energy of rotation is converted into the chemical bond between the ADP-O and Pi.

[1] doi.org/10.1016/0005-2728(93)90063-L