University of Illinois Urbana-Champaign

Fourier transform infrared difference spectroscopy of bacteriorhodopsin

Dollinger, Gavin David

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Description

Title
Fourier transform infrared difference spectroscopy of bacteriorhodopsin
Author(s)
Dollinger, Gavin David
Issue Date
1985
Doctoral Committee Chair(s)
Frauenfelder, Hans
Department of Study
Physics
Discipline
Physics
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
  • Fourier transform infrared difference spectroscopy
  • photocycle
  • proton pump
  • bacteriorhodopsin
  • chromophores
  • photoisomerization
Language
en
Abstract
We have used the technique of Fourier transform infrared difference spectroscopy to study the photocycle and proton pump of bacteriorhodopsin (bR). By applying this method to the study of native bR and bR with modified chromophores, we have confirmed and extended the earlier resonance Raman results concerning the configuration of the chromophore of bR and its intermediates. These studies also demonstrated that the all-trans to 13-cis photoisomerization of the chromophore is essential to the functioning of the protein. This tecnnique has also allowed us to explore the role of the opsin in the protein's proton pump. Large scale motions of the opsin during the photocycle were detected and a simple model to interpret these motions was formulated. The model involves the rotation of the protein's a helices towards the membrane normal during the formation of the M intermediate. The role of three amino acids, tyrosine, glutamate and aspartate, in the proton pump was investigated by comparing the difference spectra of native bR with those of bR grown on isotopically labelled analogues of these compounds. This comparison revealed that the sidechains of these amino acids undergo changes in protonation and/or environment during the photocycle, functioning, as had been thought, as proton donors and acceptors. However, we were not able to construct a synoptic model of the role of these groups in the proton pump. Finally, we have built and tested a device to be used to take kinetic infrared data on bR. The device is based on the optical spinning sample technique and is placed inside a conventional Fourier transform infrared spectrometer. It can be used to acquire data in the entire midinfrared region and has a maximum time resolution of 40 ps. This time resolution has not yet been achieved. Preliminary data taken with this device are presented.
Type of Resource
text
Permalink
http://hdl.handle.net/2142/25284
Copyright and License Information
1985 Gavin David Dollinger

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