Femtosecond Spontaneous Fluorescence Studies of Photosynthetic Bacterial Reaction Centers.

The mechanism of the initial electron transfer step in the reaction center of photosynthetic bacteria has been the subject of intense study over the past 10 years. This initial step is ultrafast, occurring in about 3 ps at room temperature [1]. As the understanding of the reaction center improves the need arises for more precise kinetic data. In particular, questions arise to the exponentiality of the observed kinetic signals [2], the possibility of differing behavior at different wavelengths [3], the existence of oscillatory components [2], and of spectral shifts accompanying the excitation and subsequent electron transfer processes. We measured the spontaneous fluorescence decay of P* in Rb. Sphaeroidis R26, Rb. Capsulatus, and mutantsof Rb. Capsulatus using the upconversion technique. For all samples QA was chemically reduced with the exception of the Rb. Sphaeroidis R26 sample for which the quinone was removed. The Rb. Spaeroidis R26 quinone removed measurements with both direct excitation of P at 850 nm and indirect excitation through internal conversion from PQX and energy transfer from BChl excited at 608 nm. All other samples were excited @ 850 nm. Excitation at 608 nm was provided by an antiresonant ring dye laser amplified at 100 Khz by a YAG regen yielding 60 fs pulses [4]. Experiments using 850 nm excitation were performed with a Coherent Mira 900 F Ti sapphire laser operating at 76 MHz with 95 fs pulses [5]. The instrument response functions at 940 nm emission for the two apparatus is ~180 fs. A typical data set is shown in Figure 1. All the samples showed nonexponential decay (see Figure 1) which could be adequately fit by a sum of two exponentials. The shorter component compares very well with the single components determined by stimulated emission [6]. However, the improved dynamic range and absence of other components (excited state absorption and ground state bleaching) make the nonexponentiality very clear. With 850 nm excitation we were unable to detect a risetime for fluorescence at 940 nm. With 610 nm excitation a risetime of 200 fs is apparent (Figure 2). This risetime results from a combination of energy transfer from the accessory pigments and electronic relaxation with the special pair manifold.