Folding and stability of the b subunit of the F(1)F(0) ATP synthase.
The F(1)F(0) ATP synthase is a reversible molecular motor that employs a rotary catalytic cycle to couple a chemiosmotic membrane potential to the formation/hydrolysis of ATP. The multisubunit enzyme contains two copies of the b subunit that form a homodimer as part of a narrow, peripheral stalk structure that connects the membrane (F(0)) and soluble (F(1)) sectors. The three-dimensional structure of the b subunit is unknown making the nature of any interactions or conformational changes within the F(1)F(0) complex difficult to interpret. We have used circular dichroism and analytical ultracentrifugation analyses of a series of N- and C-terminal truncated b proteins to investigate its stability and structure. Thermal denaturation of the b constructs exhibited distinct two-state, cooperative unfolding with T(m) values between 30 and 40 degrees C. CD spectra for the region comprising residues 53-122 (b(53-122)) showed theta;(222)/theta;(208) = 0.99, which reduced to 0.92 in the presence of the hydrophobic solvent trifluoroethanol. Thermodynamic parameters for b(53-122) (DeltaG, DeltaH and DeltaC(p)) were similar to those reported for several nonideal, coiled-coil proteins. Together these results are most consistent with a noncanonical and unstable parallel coiled-coil at the interface of the b dimer.
Revington
Matthew
M
Department of Biochemistry, The University of Western Ontario, London, Ontario, Canada, N6A 5C1.
Dunn
Stanley D
SD
Shaw
Gary S
GS
D000595:Amino Acid Sequence; D002942:Circular Dichroism; D008969:Molecular Sequence Data; D011487:Protein Conformation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D006180:Proton-Translocating ATPases / Q000737:chemistry* / Q000378:metabolism; D016415:Sequence Alignment; D012965:Sodium Chloride; D013696:Temperature; D013816:Thermodynamics; D014270:Trifluoroethanol; D014461:Ultracentrifugation
D012965:Sodium Chloride; D014270:Trifluoroethanol; D006180:Proton-Translocating ATPases