C-deuterated alanine: a new label to study membrane protein structure using site-specific infrared dichroism.
The helix tilt and rotational orientation of the transmembrane segment of M2, a 97-residue protein from the Influenza A virus that forms H(+)-selective ion channels, have been determined by attenuated total reflection site-specific infrared dichroism using a novel labeling approach. Triple C-deuteration of the methyl group of alanine in the transmembrane domain of M2 was used, as such modification shifts the asymmetric and symmetric stretching vibrations of the methyl group to a transparent region of the infrared spectrum. Structural information can then be obtained from the dichroic ratios corresponding to these two vibrations. Two consecutive alanine residues were labeled to enhance signal intensity. The results obtained herein are entirely consistent with previous site-specific infrared dichroism and solid-state nuclear magnetic resonance experiments, validating C-deuterated alanine as an infrared structural probe that can be used in membrane proteins. This new label adds to the previously reported (13)C [double bond] (18)O and C-deuterated glycine as a tool to analyze the structure of simple transmembrane segments and will also increase the feasibility of the study of polytopic membrane proteins with site-specific infrared dichroism.
Torres
Jaume
J
Cambridge Centre for Molecular Recognition, Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge, CB2 1GA, United Kingdom.
Arkin
Isaiah T
IT
D000409:Alanine / Q000737:chemistry*; D002244:Carbon / Q000737:chemistry*; D002942:Circular Dichroism; D004134:Dimyristoylphosphatidylcholine / Q000737:chemistry; D006859:Hydrogen; D008081:Liposomes / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy; D008962:Models, Theoretical; D010100:Oxygen / Q000737:chemistry*; D011487:Protein Conformation; D011506:Proteins / Q000737:chemistry*; D013053:Spectrophotometry; D017550:Spectroscopy, Fourier Transform Infrared
D008081:Liposomes; D011506:Proteins; D002244:Carbon; D006859:Hydrogen; D000409:Alanine; D010100:Oxygen; D004134:Dimyristoylphosphatidylcholine