RESONANCE RAMAN STUDIES OF BACTERIORHODOPSIN ANALOGUES |
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| Authors: | R Schiffmiller R H Callender W H Waddell † R Govindjee T G Ebrey H Kakitani † B Honig K Nakanishi |
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| Institution: | Physics Department, City College of The City University of New York, NY 10031, USA;Department of Chemistry, Carnegie-Mellon University Pittsburgh, PA 15213, USA;Department of Physiology and Biophysics, University of Illinois, Urbana, IL 61801, USA;Department of Biochemistry, Columbia University, New York, NY 10032, USA;Department of Chemistry, Columbia University, New York, NY 10027, USA |
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| Abstract: | Abstract— We present the results of resonance Raman measurements on a series of bacteriorhodopsin (bR) analogues formed from synthetic retinals which have replaced the native chromophore in the active site. Specifically, 5,6-dihydro-bR, 13-desmethyl-bR, 10-methyl-bR, 14-methyl-bR, and 10.14-dimethyl-bR have been studied. All five analogues bind and form Schiff base retinal-apoprotein linkages. While the Schiff base linkages of 5,6-dihydro-bR, 13-desmethyl-bR, and 10-methyl-bR are protonated, like the native chromophore, the 14-methyl-bR, and 10,14-dimethyl-bR Schiff bases are unprotonated. These results suggest that the binding site of bacteriorhodopsin near the Schiff base moiety is different from that of rhodopsin. The protonated Schiff base -C=NH- stretching frequency of 5.6-dihydro-bR lies at 1660 cm-1 which is unusually high for a bacteriorhodopsin based pigment. The downward shift upon deuteration is 16 cm-1, essentially identical to that measured for bacteriorhodopsin. This and the other analogue results strongly reinforce our previous arguments that the Schiff base stretching frequency is determined in large part by two factors, the C=N force constant and the stretch interaction with C=N-H bend. On the other hand, the deuterium isotope effect is determined primarily by the stretch-bend interaction. |
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