Evidence for a common structure for a class of membrane channels.
Electron microscopic analysis of gap-junction-like structures isolated from an anthropod (Nephrops norvegicus) and composed of a 16-kDa polypeptide, show the functional unit to be a star-shaped hexamer of protein arranged around a central channel which runs perpendicular to the plane of the membrane. Estimations of the molecular volume carried out on an averaged projection are consistent with a subunit mass of 16-18 kDa. Fourier transform infrared spectroscopy indicates a high alpha-helical content for the protein, supporting secondary-structure predictions of four transmembrane alpha helices/monomer. The averaged projection shows a close resemblance to a hexamer of the 16-kDa protein built on the basis of a four alpha-helical bundle [Finbow, M. E., Eliopoulos, E. E., Jackson, P. J., Keen, J. N., Meagher, L., Thompson, P., Jones, P. C. & Findlay, J. B. C. (1992) Protein Eng. 5, 7-15]. The reconstructed image is also similar to that obtained for gap-junction-like channels isolated from a related arthropod [Homarus americanus; Sikerwar, S. S., Downing, K. H. & Glaeser, R. M. (1991) J. Struct. Biol. 106, 255-263] whose protein content was unknown but which we demonstrate may be composed of a related 16-kDa protein. Previous studies have shown a high sequence identity of the Nephrops 16-kDa protein with the 16-kDa proteolipid subunit c of the vascular H(+)-ATPase, both of which in turn bear similarity to the 8-kDa proteolipid subunit of the F1F0-ATP synthase. Expression of cDNA coding for the Nephrops 16-kDa protein in Saccharomyces cerevisiae, in which the endogenous gene coding for the V-ATPase proteolipid has been inactivated, restores V-ATPase activity and cell growth.
Holzenburg
A
A
Department of Biochemistry & Molecular Biology, University of Leeds, England.
Jones
P C
PC
Franklin
T
T
Pali
T
T
Heimburg
T
T
Marsh
D
D
Findlay
J B
JB
Finbow
M E
ME
D000595:Amino Acid Sequence; D000818:Animals; D001181:Arthropods; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D005583:Fourier Analysis; D007091:Image Processing, Computer-Assisted; D007365:Intercellular Junctions / Q000737:chemistry / Q000648:ultrastructure; D007473:Ion Channels / Q000737:chemistry* / Q000648:ultrastructure; D008565:Membrane Proteins / Q000737:chemistry* / Q000648:ultrastructure; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D012441:Saccharomyces cerevisiae; D017386:Sequence Homology, Amino Acid; D013055:Spectrophotometry, Infrared
D007473:Ion Channels; D008565:Membrane Proteins