Implications of threonine hydrogen bonding in the glycophorin A transmembrane helix dimer.
The transmembrane helix of glycophorin A contains a seven-residue motif, LIxxGVxxGVxxT, that mediates protein dimerization. Threonine is the only polar amino acid in this motif with the potential to stabilize the dimer through hydrogen-bonding interactions. Polarized Fourier transform infrared spectroscopy is used to establish a robust protocol for incorporating glycophorin A transmembrane peptides into membrane bilayers. Analysis of the dichroic ratio of the 1655-cm(-1) amide I vibration indicates that peptides reconstituted by detergent dialysis have a transmembrane orientation with a helix crossing angle of <35 degrees. Solid-state nuclear magnetic resonance spectroscopy is used to establish high resolution structural restraints on the conformation and packing of Thr-87 in the dimer interface. Rotational resonance measurement of a 2.9-A distance between the gamma-methyl and backbone carbonyl carbons of Thr-87 is consistent with a gauche- conformation for the chi1 torsion angle. Rotational-echo double-resonance measurements demonstrate close packing (4.0 +/- 0.2 A) of the Thr-87 gamma-methyl group with the backbone nitrogen of Ile-88 across the dimer interface. The short interhelical distance places the beta-hydroxyl of Thr-87 within hydrogen-bonding range of the backbone carbonyl of Val-84 on the opposing helix. These results refine the structure of the glycophorin A dimer in membrane bilayers and highlight the complementary role of small and polar residues in the tight association of transmembrane helices in membrane proteins.
Smith
Steven O
SO
Department of Biochemistry and Cell Biology, Center for Structural Biology, State University of New York at Stony Brook, Stony Brook, New York 11794-5115, USA. steven.o.smith@sunysb.edu
Eilers
Markus
M
Song
David
D
Crocker
Evan
E
Ying
Weiwen
W
Groesbeek
Michel
M
Metz
Guenter
G
Ziliox
Martine
M
Aimoto
Saburo
S
D000595:Amino Acid Sequence; D019281:Dimerization; D006021:Glycophorins / Q000737:chemistry*; D006801:Humans; D006860:Hydrogen Bonding; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000737:chemistry; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D017550:Spectroscopy, Fourier Transform Infrared; D013314:Stress, Mechanical; D013912:Threonine
D006021:Glycophorins; D010446:Peptide Fragments; D013912:Threonine