Use of a single glycine residue to determine the tilt and orientation of a transmembrane helix. A new structural label for infrared spectroscopy.
Site-directed dichroism is an emerging technique for the determination of membrane protein structure. However, due to a number of factors, among which is the high natural abundance of (13)C, the use of this technique has been restricted to the study of small peptides. We have overcome these problems through the use of a double C-deuterated glycine as a label. The modification of a single residue (Gly) in the transmembrane segment of M2, a protein from the Influenza A virus that forms H(+)-selective ion channels, has allowed us to determine its helix tilt and rotational orientation. Double C-deuteration shifts the antisymmetric and symmetric stretching vibrations of the CD(2) group in glycine to a transparent region of the infrared spectrum where the dichroic ratio of these bands can be measured. The two dichroisms, along with the helix amide I dichroic ratio, have been used to determine the helix tilt and rotational orientation of M2. The results are entirely consistent with previous site-directed dichroism and solid-state NMR experiments, validating C-deuterated glycine (GlyCD(2)) as a structural probe that can now be used in the study of polytopic membrane proteins.
Torres
J
J
Department of Biochemistry, University of Cambridge, Cambridge CB2 1GA, United Kingdom.
Kukol
A
A
Arkin
I T
IT
D002942:Circular Dichroism; D003903:Deuterium; D004134:Dimyristoylphosphatidylcholine; D005998:Glycine; D009980:Influenza A virus; D007473:Ion Channels / Q000737:chemistry*; D008565:Membrane Proteins / Q000737:chemistry; D008958:Models, Molecular; D009949:Orientation; D017433:Protein Structure, Secondary; D013055:Spectrophotometry, Infrared; D014763:Viral Matrix Proteins / Q000737:chemistry*; D000086082:Viroporin Proteins
D003903:Deuterium; D004134:Dimyristoylphosphatidylcholine; D005998:Glycine; D007473:Ion Channels; D008565:Membrane Proteins; D014763:Viral Matrix Proteins; C031540:M-protein, influenza virus; C488937:M2 protein, Influenza A virus; D000086082:Viroporin Proteins