The Mn complex of photosystem II (PSII) cycles through 4 semi-stable states (S(0) to S(3)). Laser-flash excitation of PSII in the S(2) or S(3) state induces processes with time constants around 350ns, which have been assigned previously to energetic relaxation of the oxidized tyrosine (Y(Z)(ox)). Herein we report monitoring of these processes in the time domain of hundreds of nanoseconds by photoacoustic (or 'optoacoustic') experiments involving pressure-wave detection after excitation of PSII membrane particles by ns-laser flashes. We find that specifically for excitation of PSII in the S(2) state, nuclear rearrangements are induced which amount to a contraction of PSII by at least 30Å(3) (time constant of 350ns at 25°C; activation energy of 285+/-50meV). In the S(3) state, the 350-ns-contraction is about 5 times smaller whereas in S(0) and S(1), no volume changes are detectable in this time domain. It is proposed that the classical S(2)=>S(3) transition of the Mn complex is a multi-step process. The first step after Y(Z)(ox) formation involves a fast nuclear rearrangement of the Mn complex and its protein-water environment (~350ns), which may serve a dual role: (1) The Mn- complex entity is prepared for the subsequent proton removal and electron transfer by formation of an intermediate state of specific (but still unknown) atomic structure. (2) Formation of the structural intermediate is associated (necessarily) with energetic relaxation and thus stabilization of Y(Z)(ox) so that energy losses by charge recombination with the Q(A)(-) anion radical are minimized. The intermediate formed within about 350ns after Y(Z)(ox) formation in the S(2)-state is discussed in the context of two recent models of the S(2)=>S(3) transition of the water oxidation cycle. This article is part of a Special Issue entitled: Photosynthesis Research for Sustainability: From Natural to Artificial.

A novel Fourier-transform infrared (FT-IR) rapid-scan spectrometer has been developed (patent pending EP14194520.4) which yields 1000 times higher time resolution as compared to conventional rapid-scanning spectrometers. The central element to achieve faster scanning rates is based on a sonotrode whose front face represents the movable mirror of the interferometer. A prototype spectrometer with a time resolution of 13 μs was realized, capable of fully automated long-term measurements with a flow cell for liquid samples, here a photosynthetic membrane protein in solution. The performance of this novel spectrometer is demonstrated by recording the photoreaction of bacteriorhodopsin initiated by a short laser pulse that is synchronized to the data recording. The resulting data are critically compared to those obtained by step-scan spectroscopy and demonstrate the relevance of performing experiments on proteins in solution. The spectrometer allows for future investigations of fast, non-repetitive processes, whose investigation is challenging to step-scan FT-IR spectroscopy.

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