Seigneuret M, Kainosho M
Université Paris 7, Laboratoire de Biophysique Cellulaire, URA 526, France.
FEBS Lett. 1993 Jul 19;327(1):7-12. doi: 10.1016/0014-5793(93)81027-w.
High-resolution 13C-NMR experiments have been performed on bacteriorhodopsin biosynthetically labeled with carbonyl-13C amino acids and solubilized in the detergent dodecylmaltoside. 13C-NMR spectra showing good resolution were obtained in the case of labeled amino acids moderately represented in the BR sequence. For BR labeled with [13C]carbonyl methionine, several sequence-specific assignment could be performed by co-labeling with 15N amino acids or proteolysis. These assignments were used to obtain structural data on BR. Water-exposure of methionine side chains in the protein was assessed by studying, using NMR, their oxidation by hydrogen peroxide. Local secondary structure at the level of methionine residues was monitored through the effect of 1H-2H exchange on NMR spectra. It was concluded that Met32, Met68 and Met163 are peripheral while all 6 other methionine residues are deeply embedded within hydrophobic alpha-helices. These results confirm the current model of the BR folding and secondary structure.
