The "second stalk" of Escherichia coli ATP synthase: structure of the isolated dimerization domain

Biochemistry. 2002 May 28;41(21):6875-84. doi: 10.1021/bi025736i.

Abstract

The b subunit of E. coli F(0)F(1)-ATPase links the peripheral F(1) subunits to the membrane-integral F(0) portion and functions as a "stator", preventing rotation of F(1). The b subunit is present as a dimer in ATP synthase, and residues 62-122 are required to mediate dimerization. To understand how the b subunit dimer is formed, we have studied the structure of the isolated dimerization domain, b(62-122). Analytical ultracentrifugation and solution small-angle X-ray scattering (SAXS) indicate that the b(62-122) dimer is extremely elongated, with a frictional ratio of 1.60, a maximal dimension of 95 A, and a radius of gyration of 27 A, values that are consistent with an alpha-helical coiled-coil structure. The crystal structure of b(62-122) has been solved and refined to 1.55 A. The protein crystallized as an isolated, monomeric alpha helix with a length of 90 A. Combining the crystal structure of monomeric b(62-122) with SAXS data from the dimer in solution, we have constructed a model for the b(62-122) dimer in which the two helices form a coiled coil with a right-handed superhelical twist. Analysis of b sequences from E. coli and other prokaryotes indicates conservation of an undecad repeat, which is characteristic of a right-handed coiled coil and consistent with our structural model. Mutation of residue Arg-83, which interrupts the undecad pattern, to alanine markedly stabilized the dimer, as expected for the proposed two-stranded, right-handed coiled-coil structure.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Arginine / chemistry*
  • Arginine / genetics
  • Crystallography, X-Ray
  • Dimerization
  • Escherichia coli / enzymology*
  • Models, Molecular
  • Molecular Sequence Data
  • Mutation
  • Protein Conformation
  • Protein Structure, Tertiary
  • Protein Subunits
  • Proton-Translocating ATPases / chemistry*
  • Proton-Translocating ATPases / genetics

Substances

  • Protein Subunits
  • Arginine
  • Proton-Translocating ATPases

Associated data

  • PDB/1L2P