Dataset Information

Photon echoes and two dimensional spectra of the amide I band of proteins measured by femtosecond IR - Raman spectroscopy.

ABSTRACT: Infrared (IR) and Raman spectroscopy are fundamental techniques in chemistry, allowing the convenient determination of bond specific chemical composition and structure. Over the last decades, ultrafast multidimensional IR approaches using sequences of femtosecond IR pulses have begun to provide a new means of gaining additional information on molecular vibrational couplings, distributions of molecular structures and ultrafast molecular structural dynamics. In this contribution, new approaches to measuring multidimensional spectra involving IR and Raman processes are presented and applied to the study of the amide I band of proteins. Rephasing of the amide I band is observed using dispersed IR-Raman photon echoes and frequency domain 2D-IR-Raman spectra are measured by use of a mid-IR pulse shaper or over a broader spectral range using a tuneable picosecond laser. A simple pulse shaping approach to performing heterodyned time-domain Fourier Transform 2D-IR-Raman spectroscopy is introduced, revealing that the 2D-IR-Raman spectra distinguish homogeneous and inhomogeneous broadening in the same way as the well-established methods of 2D-IR spectroscopy. Across all datasets, the unique dependence of the amide I data on the IR and Raman strengths, vibrational anharmonicities and inhomogeneous broadening provides a fascinating spectroscopic view of the amide I band.

SUBMITTER: Donaldson PM

PROVIDER: S-EPMC8163424 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:Archaerhodopsin 3 (AR3) is a light driven proton pump from Halorubrum sodomense that has been used as a genetically targetable neuronal silencer and an effective fluorescent sensor of transmembrane potential. Unlike the more extensively studied bacteriorhodopsin (BR) from Halobacterium salinarum, AR3 readily incorporates into the plasma membrane of both E. coli and mammalian cells. Here, we used near-IR resonance Raman confocal microscopy to study the effects of pH and membrane potential on the AR3 retinal chromophore structure. Measurements were performed both on AR3 reconstituted into E. coli polar lipids and in vivo in E. coli expressing AR3 in the absence and presence of a negative transmembrane potential. The retinal chromophore structure of AR3 is in an all-trans configuration almost identical to BR over the entire pH range from 3 to 11. Small changes are detected in the retinal ethylenic stretching frequency and Schiff Base (SB) hydrogen bonding strength relative to BR which may be related to a different water structure near the SB. In the case of the AR3 mutant D95N, at neutral pH an all-trans retinal O-like species (O(all-trans)) is found. At higher pH a second 13-cis retinal N-like species (N(13-cis)) is detected which is attributed to a slowly decaying intermediate in the red-light photocycle of D95N. However, the amount of N(13-cis) detected is less in E. coli cells but is restored upon addition of carbonyl cyanide m-chlorophenyl hydrazone (CCCP) or sonication, both of which dissipate the normal negative membrane potential. We postulate that these changes are due to the effect of membrane potential on the N(13-cis) to M(13-cis) levels accumulated in the D95N red-light photocycle and on a molecular level by the effects of the electric field on the protonation/deprotonation of the cytoplasmic accessible SB. This mechanism also provides a possible explanation for the observed fluorescence dependence of AR3 and other microbial rhodopsins on transmembrane potential.

Dataset Information

Photon echoes and two dimensional spectra of the amide I band of proteins measured by femtosecond IR - Raman spectroscopy.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets