tailieunhanh - Báo cáo khoa học: Conformational heterogeneity of transmembrane residues after the Schiff base reprotonation of bacteriorhodopsin 15

bR, N-like and O-like intermediate states of [ 15 N]methionine-labelled wild type and D85N⁄T170C bacteriorhodopsin were accumulated in native membranes by controlling the pH of the preparations. 15 N cross polariza-tion and magic angle sample spinning (CPMAS) NMR spectroscopy allowed resolution of seven out of nine resonances in the bR-state. It was possible to assign some of the observed resonances by using 13 C⁄ 15 N rota | iFEBS Journal Conformational heterogeneity of transmembrane residues after the Schiff base reprotonation of bacteriorhodopsin 15N CPMAS NMR of D85N T170C membranes A. James Mason1 George J. Turner2 and Clemens Glaubitz1 1 Centre for Biomolecular Magnetic Resonance and Institut fur Biophysikalische Chemie . Goethe Universitat Frankfurt Germany 2 Department of Chemistry and Biochemistry Seton HallUniversity South Orange NJ USA Keywords bacteriorhodopsin solid-state NMR N-state O-state Correspondence C. Glaubitz Institut fur Biophysikalische Chemie Centre for Biomolecular Magnetic Resonance . Goethe Universitat Marie-Curie Str. 9 D-60439 Frankfurt Germany Fax 49 69798 29929 Tel 49 69798 29927 E-mail glaubitz@ Received 19 October 2004 revised 10 February 2005 accepted 28 February 2005 bR N-like and O-like intermediate states of 15N methionine-labelled wild type and D85N T170C bacteriorhodopsin were accumulated in native membranes by controlling the pH of the preparations. 15N cross polarization and magic angle sample spinning CPMAS NMR spectroscopy allowed resolution of seven out of nine resonances in the bR-state. It was possible to assign some of the observed resonances by using 13C 15N rotational echo double resonance REDOR NMR and Mn2 quenching as well as D2O exchange which helps to identify conformational changes after the bacteriorhodopsin Schiff base reprotonation. The significant differences in chemical shifts and linewidths detected for some of the resonances in N- and O-like samples indicate changes in conformation structural heterogeneity or altered molecular dynamics in parts of the protein. doi Bacteriorhodopsin 1 is a 26 kDa seven transmembrane helix protein 7TM found in the extremely halo-philic archeaon Halobacterium salinarium 2 . The proton pumping ability of this protein is conferred by the prosthetic retinal attached via a Schiff base to Lys216. The light-induced isomerization from .