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Free keywords:
H+V-ATPase; vacuolar-type ATPase; Manduca sexta; three-dimensional reconstruction; electron microscopy; simultaneous alignment; Radon transforms
CTF correction
Abstract:
We determined the structure of the V1-ATPase from Manduca sexta to a resolution of 1.8 nm, which for the first time reveals internal features of the enzyme. The V1-ATPase consists of a headpiece of 13.5 nm in diameter, with six elongated subunits, A3 and B3, of approximately equal size, and a stalk of 6 nm in length that connects V1 with the membrane-bound domain, V0. At the center of the molecule is a cavity that extends throughout the length of the A3B3 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° images, contrast transfer function correction for tilt images, and the Two-Step Radon Inversion Algorithm.
