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 C-13, 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 CD2 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.
|Number of pages||5|
|Publication status||Published - Dec 2000|