The structure of the V-1-ATPase determined by three-dimensional electron microscopy of single particles

Abstract

We determined the structure of the V-1-ATPase from Manduca sexta to a resolution of 1.8 nm, which for the first time reveals internal features of the enzyme. The V-1-ATPase consists of a headpiece of 13.5 nm in diameter, with six elongated subunits, A(3) and B-3, of approximately equal size, and a stalk of 6 nm in length that connects V-1 with the membrane-bound domain, V-O. At the center of the molecule is a cavity that extends throughout the length of the A(3)B(3) hexamer. Inside the cavity the central stalk can be seen connected to only two of the catalytic A subunits. The structure was obtained by a combination of the Random Conical Reconstruction Technique and angular refinements. Additional recently developed techniques that were used include methods for simultaneous translational rotational alignment of the 0 degrees images, contrast transfer function correction for tilt images, and the Two-Step Radon Inversion Algorithm. (C) 2001 Academic Press.