On the state of carotenoids bound to reaction centers of photosynthetic bacteria: a resonance Raman study.

The carotenoids bound to reaction centers of wild, Ga and GIC strains of Rhodopseudomonas spheroides, of Rhodospirrillum rubrum, strain S1 and of Rhodopseudomonas viridis, yield very similar, but unusual resonance Raman spectra. Through a comparison with resonance Raman spectra of 15,15'-cis-beta-carotene, these carotenoids are shown to assume cis conformations, while the corresponding chromatophores contain all-trans forms only. These cis conformations likely are identical for all the carotenoids studied. They remain unaffected by variations of temperature from 20 to 300 K as well as by the redox state of P-870. They are unstable, being rapidly isomerised towards the all-trans forms when extracted from the reaction centers. The possible nature of these conformers is discussed on the basis of their electronic and vibrational spectra.

Dependence of energization of thylakoids on frequency of exciting flashes in intact chloroplasts.

We investigated the frequency-dependence of the flash-induced electrochromic absorbance change, ΔA515, and of the pH-indicating absorbance change of neutral red in isolated intact chloroplasts. The energization pattern of thylakoids depended strongly on the frequency (f) of the exciting flashes, tested between 0.05 and 2 s(-1). When the frequency was increased from 0.1 to 1 s(-1) the total initial change and the slow rise of ΔA515 decreased by about 30% and 70%, respectively, and both the slow rise and decay were considerably accelerated. These changes were fully reversible, even after prolonged excitation at 1 s(-1), if the frequency was decreased again to 0.1 s(-1). Accumulation of an appreciable transmembrane electric field strength could not be detected in any of our experiments, at high frequency, since the decay of ΔA515 was considerably accelerated when the frequency was increased. In contrast, ΔpH significantly increased at higher frequencies of the exciting flashes. In the steady-state (after about 100 flashes) ΔpH was about 0.5-0.8 pH unit higher than in the dark or at low frequencies. In the presence of nigericin or dithionite, both of which prevented accumulation of protons in the lumen, the total initial change in ΔA515 at f=1 s(-1) relative to that at f=0.1 s(-1) decreased to a similar extent as in the control. The proportion of the slow rise relative to the initial amplitude, however, did not decrease. Our data support the suggestion that ΔpH controls the amplitude of the slow rise of ΔA515. However, contrary to a previous statement (B. Bouges-Bouquet (1981) Biochim. Biophys. Acta 535, 327-340), we show that the ΔpH effect cannot be accounted for by variation of the rate of this kinetic component of ΔA515.

Evidence that the variable chlorophyll fluorescence in Chlamydomonas reinhardtii is not recombination luminescence.

Room temperature single photon timing measurements on intact, Chlamydomonas reinhardtii cells at low excitation energies have been analysed using a four exponential kinetic model. Closing the PSII reaction centres produced two major variable lifetime and two minor constant lifetime components. The yield of each component mirrored the changes in lifetime. Such observations indicate the presence of well-connected PSII centres favoring excitation energy transfer. A Chlamydomonas mutant lacking PSII reaction centre proteins exhibited decay components equivalent to those seen at FM in the wild-type. A titration of in vivo fluorescence, in both the mutant and wild-type algae, using DNB, produced decay components similar to those seen on opening PSII reaction centres. Such observations indicate that the luminescence hypothesis for the origin of the long-lived lifetime component is not the case.