RESONANCE RAMAN STUDIES OF BACTERIORHODOPSIN ANALOGS AND RELATED SYSTEMS.
Item
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Title
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RESONANCE RAMAN STUDIES OF BACTERIORHODOPSIN ANALOGS AND RELATED SYSTEMS.
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Identifier
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AAI8103960
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identifier
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8103960
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Creator
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SCHIFFMILLER, RICHARD.
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Contributor
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Robert H. Callender
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Date
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1980
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Language
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English
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Publisher
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City University of New York.
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Subject
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Biophysics, General
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Abstract
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In order to better understand the protein-chromophore interactions in bacteriorhodopsin, five retinal analogs--10,14-dimethyl, 14-methyl, 10-methyl, 5,6-dihydro, and 13-desmethyl--have been regenerated with the opsin apoprotein from the purple membrane of Halobacterium Halobium to form artificial bacteriorhodopsin pigments. These were studied spectroscopically at liquid nitrogen temperatures using resonance Raman scattering techniques. A line appearing at 1010 cm('-1) is assigned to a methyl vibration, while the 1300 cm('-1) vibration is assigned to an interaction mode of adjacent methyl groups. The color of the analog pigments is shown to be due to the degree of (pi)-electron delocalization on the polyene chain of the retinal chromophore. Both 10,14-dimethyl and 14-methyl bacteriorhodopsin are shown to have their chromophores linked to the protein by unprotonated Schiff bases, unlike the other analogs, and their blue-shifted absorption maxima are explained in terms of that observation. It is found that the shift in frequency of the C = C stretching vibration correlates linearly with the absorption maxima of the various analogs, as predicted by (pi)-electron delocalization theory. Moreover, for the first time, the frequency of the C = N stretching vibration is correlated with the extent of delocalization in each analog.;The environment-sensitive lines of the 5,6-dihydro chromophore are shown to be affected less by the protein binding site than those of the other retinals, thus supporting a model which suggests the existence of a counter-ion in the bacterio-opsin protein near the chromophore's ionone ring.;Deuteration of the analogs with deuterium oxide reveals a very small shift in the 13-desmethyl C = NH('+) frequency, and possible implications of this are discussed.;The double beam pump/probe technique is used to obtain the resonance Raman spectrum of the first intermediate of the bacteriorhodopsin photocycle, known as "K". From the significant differences between the fingerprint region vibrations of bacteriorhodopsin and K, we conclude that a trans-cis isomerization has occurred in the photoinduced formation of K. Furthermore, the deuterated spectrum of K confirms that the Schiff base of K is protonated, which argues against a model that proposes that proton tunnelling is the primary event in bacteriorhodopsin photoexcitation.;Finally, resonance Raman spectra of a homologous series of cyanine dyes, diQ - C(,n) - (2m + 1), are presented. These dyes represent an extreme degree of (pi)-electron delocalization and the lack of a shift in the key lines in their spectra with increasing polyene chain length confirms that a change in delocalization is responsible for Raman line shifts in polyene systems.
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Type
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dissertation
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Source
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PQT Legacy CUNY.xlsx
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degree
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Ph.D.
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Program
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Physics
