title: A reaction-induced fourier transform-infrared spectroscopic study of the lactose permease. A transmembrane potential perturbs carboxylic acid residues.
abstract: In chemiosmotic coupling, a transmembrane ion gradient is used as the source of energy to drive reactions. This process occurs in all cells, but the microscopic mechanism is not understood. Here, Escherichia coli lactose permease was used in a novel spectroscopic method to investigate the mechanism of chemiosmotic coupling in secondary active transporters. To provide a light-triggered electrochemical gradient, bacteriorhodopsin was co-reconstituted with the permease, and reaction-induced Fourier transform-infrared spectra were obtained from the co-reconstituted samples. The bacteriorhodopsin contributions were subtracted from these data to give spectra reflecting permease conformational changes that are induced by an electrochemical gradient. Positive bands in the 1765-1730 cm(-1) region are attributable to carboxylic acid residues in the permease and are consistent with changes of pK(a), protonation state, or environment. This is the first direct information concerning gradient-induced structural changes in the permease at the single amino acid level. Ultimately, these structural changes facilitate galactoside binding and may be involved in the storage of free energy.
authors.affiliation: Department of Biochemistry, Biological Process Technology Institute, University of Minnesota, St. Paul, Minnesota 55108, USA.
authors.forename: J S
authors.initials: JS
authors.lastname: Patzlaff
authors.forename: R J
authors.initials: RJ
authors.lastname: Brooker
authors.forename: B A
authors.initials: BA
authors.lastname: Barry
mesh_terms: D001436:Bacteriorhodopsins / Q000737:chemistry; D029968:Escherichia coli Proteins; D008564:Membrane Potentials; D026901:Membrane Transport Proteins / Q000737:chemistry*; D009004:Monosaccharide Transport Proteins; D017550:Spectroscopy, Fourier Transform Infrared; D027981:Symporters
chemical_list: D029968:Escherichia coli Proteins; C050221:LacY protein, E coli; D026901:Membrane Transport Proteins; D009004:Monosaccharide Transport Proteins; D027981:Symporters; D001436:Bacteriorhodopsins; C021499:lactose permease