The effects of high hydrostatic pressure (up to 8 kbar) on bacteriochlorophyll a Q y electronic absorption bands of LH2 photosynthetic antenna complexes have been studied at ambient temperature. A variety of samples were studied, including intact membranes and isolated complexes from wild type and mutant photosynthetic bacteria Rhodobacter sphaeroides, Rhodopseudomonas acidophila, and Rhodospirillum molischianum. The spectra of the complexes universally red shift and broaden under elastic compression, while the variations of the integrated intensity remain within the experimental uncertainty. A qualitatively different slope and variation of the slope of the pressure-induced shift is observed for the B800 and B850 absorption bands of LH2 complexes belonging to quasi-monomer and aggregated pigments, respectively. For the complexes from Rhodobacter sphaeroides, e.g., the corresponding slopes are −28 ± 2 and −65 ± 2 cm-1/kbar. The shift rate of the B800 band declines with pressure, while the opposite is observed for the B850 band. The shifts show little if any correlation with hydrogen bonds. Using simple phenomenological arguments and numerical simulations of molecular exciton spectra, it is shown that the shift of the B800 band is governed by pigment−protein interactions, while in addition to that, interpigment couplings (including long-range dipolar and short-range orbital overlap interactions) are instrumental for the B850 band shift. The compressibility of the B800 bacteriochlorophyll binding sites deduced from the B800 band shift at ambient pressure is ∼0.02 kbar-1, and it decreases nonlinearly with pressure. Inter-pigment couplings are responsible for approximately one-third of both the total ambient-pressure solvent shift of the B850 absorption band and its pressure-induced growth. A slight increase with pressure of the B850 band shift due to orbital overlap couplings is predicted.

Using the bacteriochlorophyll a (Bchl) cofactors as intrinsic probes to monitor changes in membrane protein structure, we investigate the response to high-pressure of the LH2 complexes from the photosynthetic bacteria Rhodobacter sphaeroides 2.4.1 and Rhodopseudomonas acidophila 10050. By FT-Raman spectroscopy, we demonstrate that high pressure does not induce significant distortion of the protein-bound 850 nm-absorbing bacteriochlorophyll molecules, or break of the hydrogen bond they are involved in. This indicates in particular that the oligomerization of the polypeptides is not perturbed up to 0.6 GPa. The pressure-induced changes in the Bchl absorption spectra are attributed to pigment-pigment interactions. In contrast, the loss of 800 nm-absorbing bacteriochlorophyll reflects pressure-induced alterations to the tertiary structure of the protein in proximity to the membrane/cytosol interface. This suggests that the LH2 protein does have two independent structural domains. The first domain is pressure independent and comprises mostly the C-terminal domain. The second domain located on the N-terminal side exhibits sensitivity to pressure and pH reminiscent of soluble proteins. The LH2 thus constitutes a suitable model system for studying in detail the stability of membrane-embedded hydrophobic helices and helices located at or close to the solvent/membrane interface.

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In this paper we examine the effect of pressure on the absorption spectrum and binding site of the core antenna complex from the photosynthetic bacterium Rhodospirillum rubrum. Absorption spectra and Raman spectra in preresonance with the Qy transition of the bacteriochlorophyll a were studied at pressures up to 625 MPa. In agreement with previous work we observe a pressure-induced red shift and broadening of the absorption spectrum. We show that at these pressures the pigments within the protein matrix at room temperature experience little if any distortion, and the hydrogen-bonding network involving the C2 and C9 carbonyl groups of the pigment molecules are undisturbed. Having shown the lack of sensitivity to pressure of the binding site interactions, which are known to modulate the absorption spectrum, we feel that it is relatively safe to attribute the pressure-induced red shift broadly to solvatochromic effects and, in particular, to the modulation of the pigment-pigment interactions by the pressure. This paper represents the first vibrational study of photosynthetic complexes at high pressure and the first application of FT Raman spectroscopy to biological molecules at high pressure.