Picosecond detection of an intermediate in the photochemical reaction of bacterial photosynthesis.

Abstract

Analysis of the spectrum suggests that the formation of pF involves electron transfer from one bacteriochlorophyll molecule to another within the reaction center, or possibly from bacteriochlorophyll to the bacteriopheophytin of the complex. The initial absorbance changes after flash excitation also include a bleaching of an absorption band at 800 nm. The bleaching decays with T AZ 30 psec. The bleaching appears not to be a secondary effect, but rather to reveal another early step in the primary photochemical reaction.The primary photochemical reaction of bacterial photosynthesis is the transfer of an electron from a bacteriochlorophyll complex, P, to an acceptor, X, whose identity is uncertain (1-3). The electron transfer reaction occurs with a quantum yield of essentially 100% (4), and it occurs with great speed, even at temperatures below 40K. It can be studied in preparations of isolated "reaction centers," which contain three different polypeptides, four equivalents of bacteriochlorophyll, two of bacteriopheophytin, and one each of ubiquinone and nonheme iron (1-3).In a search for clues to the mechanism of the electron transfer reaction, Parson et al. (5) State pF is unlikely to be the lowest excited singlet state of the bacteriochlorophyll complex, P*, because measurements of the fluorescence yield indicate that the lifetime of P* is only 20 to 40 psec when X is in the reduced form (and less when X is not reduced) (6, 7). The proposal that the primary reaction involves an intermediate state such as pF, rather than proceeding directly from P*, would explain a longstanding anomaly in the quantitative relationship between the quantum yields of photochemistry and fluorescence (3-5). The possibility has remained, however, that pF is a side-product which forms only if the normal reaction is blocked. A decision on the role of state pF in photosynthesis, therefore, requires information on whether pF is formed under conditions that permit the electron transfer reaction to occur. We report here on an investigation of this point, using picosecond kinetic techniques. The results appear to establish pF as an intermediate in the electron transfer reaction. While this work was in progress, Kaufmann et al. (8) reached the same conclusion, using somewhat different techniques of psec spectroscopy. MATERIALS AND METHODSReaction centers were obtained from cells of Rhodopseudomonas sphaeroides strain R-26 by the method of Clayton and Wang (9). The detergent lauryldimethylamine oxide was replaced by Triton X-100 by dialysis (5, 10).The apparatus used in the psec experiments was essentially that of Magde and Windsor (11). We shall summarize the approach only briefly here for clarity; ref. 11 provides additional details. A single pulse from a mode-locked Nd+3/glass laser was frequency-doubled to provide a 530 nm excitation flash lasting about 8 psec. This illuminated only a small band across the center of the sample. A measuring flash of white light, which was generated by self phase modulation of residual 1060 nm light from th...

EPR properties of the electron carrier intermediate between the reaction center bacteriochlorophylls and the primary acceptor in Chromatium vinosum

“…Evidence for the intermediate comes from picosecond spectroscopy, whichhas suggested that although the (BChl)2 is oxidized within 10 ps of a flash of light [1 ], the primary acceptor is not reduced until some 100-200 ps later [2,3]. During this interval the electron is proposed to reside on the low redox potential I, as part of a transient [(BChl)2÷'I-'] state.…”

“…During this interval the electron is proposed to reside on the low redox potential I, as part of a transient [(BChl)2÷'I-'] state. If normal forward photochemistry is to occur, this state reduces X in 100-200 ps [2,3] but if this is blocked by the prior chemical reduction [4] or removal [5] of X, the transient state has a lifetime of nanoseconds before decaying (t½ ~, 10 ns at 300°K; "~ 30 ns at 80°K [4,6]). It has been suggested [7] that [-" has some spectrophotomeric characteristics of reduced bacteriopheophytin (BPh-").…”

“…sphaeroides and C vinosum was employed to trap the I-" state in C. vinosum at 200°K. Starting with all components reduced except I (top), light generates the intermediary state (center) in a reversible manner, see [1][2][3][4][5][6]. Although the cytochrome Css3 oxidation step at 200°K is 1000-fold slower than the [(BChl ÷ ")2 I-"] decay back reaction(s), its irreversibility, together with prolonged illumination at 200°K, leads to the product (bottom) containing a trapped I-" at the expense of only cytochrome Css3 oxidation, without net redox state changes of (BChl)2 or X.…”

Activationless electron transfer processes in biological systems

“…This pattern has been observed for the DeVault-Chance ET process between cytochrome and the reaction centre in chromatium [ 51. Experimental studies of the primary charge separation steps in bacterial photosynthesis [6,7] have established that several of the elementary ET processes are temperature independent over a wide temperature range [8]. The ET process between bacteriopheophtin (BPh) and ubiquionone-10 (Q) [ 6-81:…”

Light and life III

“…It is my belief that these agonies were counterproductive in the extreme. (33,8). From there it is handed within a fraction of a nanosecond still further to the so-called primary acceptor, now believed in the case of photosynthetic bacteria to be a complex of a nonheme iron and a quinone (31).…”