ATP Synthase Consists of Two ComplexesFj and F0

ATP synthase actually consists of two principal complexes. The spheres observed in electron micrographs make up the Fj unit, which catalyzes ATP synthesis. These F1 spheres are attached to an integral membrane protein aggregate called the F0 unit. F1 consists of five polypeptide chains named a, ¡3, y, S, and e, with a subunit stoichiometry a3fi3ySe (Table 21.3). F0 consists of three hydrophobic subunits denoted by a, b, and c, with an apparent stoichiometry of a1b2c9-12. F0 forms the transmembrane pore or channel through which protons move to drive ATP synthesis. The a, ¡3, y, S, and e subunits of F1 contain 510, 482, 272, 146, and 50 amino acids, respectively, with a total molecular mass

for F1 of 371 kD. The a and ¡3 subunits are homologous, and each of these subunits bind a single ATP. The catalytic sites are in the ¡3 subunits; the function of the ATP sites in the a subunits is unknown (deletion of the sites does not affect activity).

John Walker and his colleagues have determined the structure of the Fj complex (Figure 21.24). The F1-ATPase is an inherently asymmetrical structure, with the three ¡3 subunits having three different conformations. In the structure solved by Walker, one of the ¡-subunit ATP sites contains AMP-PNP (a nonhydrolyzable analog of ATP), and another contains ADP, with the third site being empty. This state is consistent with the binding change mechanism for ATP synthesis proposed by Paul Boyer, in which three reaction sites cycle concertedly through the three intermediate states of ATP synthesis (take a look at Figure 21.28 on page 697).

How might such cycling occur? Important clues have emerged from several experiments that show that the y subunit rotates with respect to the a/3 complex. How such rotation might be linked to transmembrane proton flow and ATP synthesis is shown in Figure 21.25. In this model, the c subunits of F0

FIGURE 21.24 • Molecular graphic images (a) side view and (b) top view of the F1-ATP synthase showing the individual component peptides. The y-subunit is the pink structure visible in the center of view (b).

FIGURE 21.24 • Molecular graphic images (a) side view and (b) top view of the F1-ATP synthase showing the individual component peptides. The y-subunit is the pink structure visible in the center of view (b).

FIGURE 21.25 • A model of the F1 and F0 components of the ATP synthase, a rotating molecular motor. The a, b, a, /, and 8 subunits constitute the stator of the motor, and the c, y, and e subunits form the rotor. Flow of protons through the structure turns the rotor and drives the cycle of conformational changes in a and ¡3 that synthesize ATP.

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FIGURE 21.25 • A model of the F1 and F0 components of the ATP synthase, a rotating molecular motor. The a, b, a, /, and 8 subunits constitute the stator of the motor, and the c, y, and e subunits form the rotor. Flow of protons through the structure turns the rotor and drives the cycle of conformational changes in a and ¡3 that synthesize ATP.

c are arranged in a ring. Several lines of evidence suggest that each c subunit consists of a pair of antiparallel transmembrane helices with a short hairpin loop on the cytosolic side of the membrane. A ring of c subunits could form a rotor that turns with respect to the a subunit, a stator consisting of five transmembrane a-helices with proton access channels on either side of the membrane. The y subunit is postulated to be the link between Fx and F0. Several experiments have shown that y rotates relative to the (a/3) 3 complex during ATP synthesis. If y is anchored to the c subunit rotor, then the c rotor-y complex can rotate together relative to the (a/3) 3 complex. Subunit b possesses a single transmembrane segment and a long hydrophilic head domain, and the complete stator may consist of the b subunits anchored at one end to the a subunit and linked at the other end to the (a/3)3 complex via the 8 subunit, as shown in Figure 21.25.

What, then, is the mechanism for ATP synthesis? The c rotor subunits each carry an essential residue, Asp61. (Changing this residue to Asn abolishes ATP synthase activity.) Rotation of the c rotor relative to the stator may depend upon neutralization of the negative charge on each c subunit Asp61 as the rotor turns. Protons taken up from the cytosol by one of the proton access channels in a could protonate an Asp61 and then ride the rotor until they reach the other proton access channel on a, from which they would be released into the matrix. Such rotation would cause the y subunit to turn relative to the three ^-subunit nucleotide sites of Fx, changing the conformation of each in sequence, so that ADP is first bound, then phosphorylated, then released, according to Boyer's binding change mechanism. Paul Boyer and John Walker shared in the 1997 Nobel Prize for chemistry for their work on the structure and mechanism of ATP synthase.

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