Time-resolved FTIR study of light-driven sodium pump rhodopsins.

Area of Science:

• Biophysics
• Microbial Rhodopsins
• Spectroscopy

Background:

• Microbial rhodopsins are light-driven ion pumps.
• Sodium ion pumps (NaR) represent a novel functional class.
• Previous studies identified K, L/M, and O intermediates in KR2, with limited structural data on later stages.

Purpose of the Study:

• To elucidate the structural dynamics of intermediates in light-driven sodium pumps.
• To investigate the chromophore and protein structural changes during the NaR photocycle.
• To compare the photocycle mechanisms of Krokinobacter eikastus rhodopsin 2 (KR2) and Nonlabens dokdonensis rhodopsin 2 (NdR2).

Main Methods:

• Step-scan time-resolved Fourier-transform infrared (FTIR) spectroscopy.
• Low-temperature static FTIR spectroscopy.
• Isotope labeling studies using 12,14-D2 retinal.

Main Results:

• A K-like intermediate was resolved in both KR2 and NdR2 with minimal spectral differences.
• Unique persistence of hydrogen-out-of-plane (HOOP) vibrations in K, L, and O intermediates suggests similar chromophore structures.
• Isotope labeling confirmed a 13-cis retinal configuration for the O intermediate in NaR.
• Protein vibrations differed among intermediates, correlating with sodium-pumping function.

Conclusions:

• The chromophore structure remains largely conserved across K, L, and O intermediates in NaR.
• The O intermediate in NaR possesses a 13-cis retinal configuration.
• Distinct protein structural changes are linked to the sodium-pumping mechanism in NaR.