Dehydration affects the stability of primary charge separation in bacterial reaction centers: Studies by optical and differential FTIR spectroscopy

The photosynthetic reaction center (RC) of Rb. sphaeroides catalyzes light-induced electron transfer events which are connected to the conformational dynamics of the protein. The light-induced charge separation between the primary donor (P) and the quinone acceptor (QA) is stabilized by solvent/protein conformational rearrangements. After a laser pulse P+QA recombination, which occurs with a lifetime t~100 ms in room temperature solutions, is accelerated (t~20 ms) at cryogenic temperatures [1] and in dehydrated glassy matrices at room temperature [2]. After prolonged photoexcitation, a slow phase of recombination (t~250 s) is observed, attributed to additional conformational changes [3]. Differential FTIR bands of water associated with the QA/QA transition have been observed upon continuous illumination, leading to propose that weakly bound water molecules plays a role in P+QA stabilization [4]. By controlling the hydration level of RC-detergent films, through equilibration at given relative humidities (r), a strong inhibition of the P+QA conformational stabilization has been observed at low hydration [5]. We compared FTIR light-minus-dark (P+QA/ PQA) differential spectra in hydrated (r=76%) and dehydrated (r=11%) RC films over the 4000- 1000 cm-1. The spectra differ significantly in the 3750-3550 cm-1 range, the band attributed to weakly hydrogen bonded water molecules [5] being strongly reduced in the dried film. Dehydration also affects the 1800-1200 cm-1 range, which includes contributions from P, the quinones and the peptide. Optical absorption measurements performed under the same photoexcitation regime reveal a slow (t~5 s) kinetic component of P+QA recombination which disappears in the dehydrated sample, indicating at low r a destabilization of the charge separated state. As a whole the data suggest a correlation between the hydration shell dynamics and the conformational RC dynamics which stabilize the charge separated state.