Conformational Complexity in the LH2 Antenna of the Purple Sulfur Bacterium Allochromatium vinosum Revealed by Hole-Burning Spectroscopy

Abstract

This work discusses the protein conformational complexity of the B800-850 LH2 complexes from the purple sulfur bacterium Allochromatium vinosum, focusing on the spectral characteristics of the B850 chromophores. Low-temperature B850 absorption and the split B800 band shift blue and red, respectively, at elevated temperatures, revealing isosbestic points. The latter indicates the presence of two (unresolved) conformations of B850 bacteriochlorophylls (BChls), referred to as conformations 1 and 2, and two conformations of B800 BChls, denoted as B800 and B800. The energy differences between average site energies of conformations 1 and 2, and B800 and B800 are similar (∼200 cm), suggesting weak and strong hydrogen bonds linking two major subpopulations of BChls and the protein scaffolding. Although conformations 1 and 2 of the B850 chromophores, and B800 and B800, exist in the ground state, selective excitation leads to 1 → 2 and B800 → B800 phototransformations. Different static inhomogeneous broadening is revealed for the lowest energy exciton states of B850 (fwhm ∼195 cm) and B800 (fwhm ∼140 cm). To describe the 5 K absorption spectrum and the above-mentioned conformations, we employ an exciton model with dichotomous protein conformation disorder. We show that both experimental data and the modeling study support a two-site model with strongly and weakly hydrogen-bonded B850 and B800 BChls, which under illumination undergo conformational changes, most likely caused by proton dynamics.

Contribution of low-temperature single-molecule techniques to structural issues of pigment–protein complexes from photosynthetic purple bacteria

“…Interface 15: 20170680 experiments, the situation appears to be static because the proteins are locked into a conformation with distinct electronic couplings among the B800 BChl a molecules, giving rise to the observed dimerization. In the hole-burning experiments reported in [43], which are conducted at lower intensities, only a part of the proteins are driven into this conformation. Then the average over the sub-ensemble of proteins, in particular including those that are only exposed briefly to the laser light due to spectral diffusion and/or non-resonant excitation, yields a mixture of dimerized/non-dimerized B800 molecules.…”

“…Using hole-burning spectroscopy, it has been shown unambiguously that the LL LH2 complex from Alc. vinosum undergoes a light-induced conformational change [43]. In single-molecule spectroscopy, a minimum excitation intensity is required to generate a signal that can be discriminated from the background.…”

“…Clearly, both techniques, single-molecule spectroscopy and spectral hole burning, have their strengths and weaknesses, but together they can be exploited for obtaining complementary information. Comparing the intensities used in the hole-burning and in the single-molecule experiments, it is clear that for the latter experiments the proteins are driven predominantly into one of the light-induced conformations found in [43]. Nevertheless, from the single-molecule experiments clear evidence is obtained for a heterogeneous apoprotein composition in a single ring (whether driven into an out-of-equilibrium population of the two conformations or not), and the molischianum-like pigment arrangement is confirmed in agreement with the sequence analysis.…”

“…Given that it takes about 30 s for recording a fluorescence-excitation spectrum from a single complex, it becomes clear that these spectra represent time-averaged spectra. The hole-burning data presented in [43] have been collected from many minutes up to several hours. Hence, these represent averages over both, time and sub-ensembles of fluctuating composition.…”

“…Interface 15: 20170680 experiments, the situation appears to be static because the proteins are locked into a conformation with distinct electronic couplings among the B800 BChl a molecules, giving rise to the observed dimerization. In the hole-burning experiments reported in [43], which are conducted at lower intensities, only a part of the proteins are driven into this conformation.…”