On the efficiency of energy transfer and the different pathways of electron transfer in mutant reaction centers of Rhodobacter sphaeroides (original) (raw)
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Febs Letters, 1990
Monomeric bacteriochlorophylls BA and Ba in photosynthetic reaction centers from Rhodobacter sphueroides R26 were exchanged with (132-hydroxy-)bacteriochlorophylls containing a 3-vinyl-or 3-(a-hydroxyethyl)-substituent instead of the 3-acetyl group. The corresponding binding sites must be tolerant to the introduction of the polar residue at C-13* andmodifications of the 3-acetyl group. According to HPLC analysis, the exchange with both pigments amounts to S$ 50% of the total BChl contained in the complex, corresponding to 6 100% of the monomeric BChl aBA,a. The absorption spectra show significant changes in the Qx and Qv-region of the monomeric bacteriochlorophylls. By contrast, the absorption of the primary donor (P870) and reversible photobleaching is retained. The circular dichroism is also unchanged in the 870 nm region. The positive cd band located at around 800 nm in native reaction centers, shifts with the (blue-shifted) QY absorption(s) of BA and/or Ba, whereas the position of the negative one remains nearly unaffected. The data indicate that the latter is the upper excitonic band of the primary donor, and that there is little interaction of the monomeric BA/Bs with the primary donor.
Biochemistry, 1997
Femtosecond transient absorption spectroscopy in the range of 500-1040 nm was used to study electron transfer at 5 K in reaction centers of Rhodobacter sphaeroides R26 in which the bacteriopheophytins (BPhe) were replaced by plant pheophytin a (Phe). Primary charge separation took place with a time constant of 1.6 ps, similar to that found in native RCs. Spectral changes around 1020 nm indicated the formation of reduced bacteriochlorophyll (BChl) with the same time constant, and its subsequent decay in 620 ps. This observation identifies the accessory BChl as the primary electron acceptor. No evidence was found for electron transfer to Phe, indicating that electron transfer from B A -occurs directly to the quinone (Q A ) through superexchange. The results are explained by a model in which the free energy level of P + Phelies above that of P + B A -, which itself is below P*. Assuming that the pigment exchange does not affect the energy levels of P* and P + B A -, our results strongly support a two-step model for primary electron transfer in the native bacterial RC, with no, or very little, admixture of superexchange. Abstract published in AdVance ACS Abstracts, November 15, 1997. 1 Abbreviations: BChl, bacteriochlorophyll; BPhe, bacteriopheophytin; Phe, pheophytin; RC, reaction center; HA,B-exchanged RCs, RCs with BPhe on the A-and B-branches exchanged for plant Phe; HBonly exchanged RCs, RCs with only BPhe on the B-branch exchanged for plant Phe; DADS, decay-associated difference spectrum; SADS, species-associated difference spectrum.
Photosynthesis Research, 1994
The tyrosine-(M)210 of the reaction center of Rhodobacter sphaeroides 2.4.1 has been changed to a tryptophan using site-directed mutagenesis. The reaction center of this mutant has been characterized by low-temperature absorption and fluorescence spectroscopy, time-resolved sub-picosecond spectroscopy, and magnetic resonance spectroscopy. The charge separation process showed bi-exponential kinetics at room temperature, with a main time constant of 36 ps and an additional fast time constant of 5.1 ps. Temperature dependent fluorescence measurements predict that the lifetime of P* becomes 4-5 times slower at cryogenic temperatures. From EPR and absorbance-detected magnetic resonance (ADMR, LD-ADMR) we conclude that the dimeric structure of P is not significantly changed upon mutation. In contrast, the interaction of the accessory bacteriochlorophyll B A with its environment appears to be altered, possibly because of a change in its position.
1998
Low temperature absorption and linear dichroism (LD) measurements were performed on oriented membranes containing wild type Rhodobacter sphaeroides reaction centers, a mutant reaction center with the change Phe M197 to Arg (FM197R), and a double mutant reaction center where, in addition, Gly M203 was replaced by Asp (FM197R/GM203D). The monomeric bacteriochlorophyll band (B), which is highly congested in the wild type reaction center, was separated into two bands in the mutant reaction centers peaking 10 nm (single mutant) or 15 nm (double mutant) apart. This separation arose principally from changes in the interaction of the protein with the L-side monomer bacteriochlorophyll B L .The ability to separate the B bands is extremely useful in spectroscopic studies. The orientations of the two monomer-type transitions contributing to the B band were similar in all three reaction centres studied, and were asymmetric with respect to the orientation axis, with the transition mostly associated with B L making a smaller angle with the C 2 axis. Differences in the LD observed in wild type membrane-bound or isolated reaction centers can be ascribed either to differences in shifts of the B transitions or to differences in the orientation axis.
Biochemistry, 1997
Absorbance difference kinetics were measured on quinone-reduced membrane-bound wild type Rhodobacter sphaeroides reaction centers in the wavelength region from 690 to 1060 nm using 800 nm excitation. Global analysis of the data revealed five lifetimes of 0.18, 1.9, 5.1, and 22 ps and a long-lived component for the processes that underlie the spectral evolution of the system. The 0.18 ps component was ascribed to energy transfer from the excited state of the accessory bacteriochlorophyll (B*) to the primary donor (P*). The 1.9 ps component was associated with a state involving a BChl anion absorbing in the 1020 nm region. This led to the conclusion that primary electron transfer is best described by a model in which the electron is passed from P* to the acceptor bacteriopheophytin (H L) via the monomeric bacteriochlorophyll (B L), with the formation of the radical pair state P + B L-† The investigations were supported by the Life Sciences Foundation (SLW), which is subsidized by the Netherlands Organization for Scientific Research (NWO) and by EC Contracts CT92-0796 and CT93-0278. M.R.J. is a BBSRC Advanced Research Fellow.
Biochemistry, 1997
Absorbance difference kinetics were measured on quinone-reduced membrane-bound wild type Rhodobacter sphaeroides reaction centers in the wavelength region from 690 to 1060 nm using 800 nm excitation. Global analysis of the data revealed five lifetimes of 0.18, 1.9, 5.1, and 22 ps and a long-lived component for the processes that underlie the spectral evolution of the system. The 0.18 ps component was ascribed to energy transfer from the excited state of the accessory bacteriochlorophyll (B*) to the primary donor (P*). The 1.9 ps component was associated with a state involving a BChl anion absorbing in the 1020 nm region. This led to the conclusion that primary electron transfer is best described by a model in which the electron is passed from P* to the acceptor bacteriopheophytin (H L ) via the monomeric bacteriochlorophyll (B L ), with the formation of the radical pair state P + B L -† The investigations were supported by the Life Sciences Foundation (SLW), which is subsidized by the Netherlands Organization for Scientific Research (NWO) and by EC Contracts CT92-0796 and CT93-0278. M.R.J. is a BBSRC Advanced Research Fellow.
1997
ABSTRACT: It is generally accepted that electron transfer in bacterial photosynthesis is driven by the first singlet excited state of a special pair of bacteriochlorophylls (P*). We have examined the first steps of electron transfer in a mutant of the Rhodobacter sphaeroides reaction center in which charge separation from P * is dramatically slowed down. The results provide for the first time clear evidence that excitation of the monomeric bacteriochlorophyll in the active branch of the reaction center (BA) drives ultrafast transmembrane electron transfer without the involvement of P*, demonstrating a new and efficient mechanism for solar energy transduction in photosynthesis. The most abundant charge-separated intermediate state probably is P+BA-, which is formed within 200 fs from BA * and decays with a lifetime of 6.5 ps into P+HA-. We also see evidence for the involvement of a BA+HA- state in the alternative pathway. In photosynthesis, the conversion of light energy into electro...
Biochimica et Biophysica Acta (BBA) - Bioenergetics, 2000
After the light-induced charge separation in the photosynthetic reaction center (RC) of Rhodobacter sphaeroides, the electron reaches, via the tightly bound ubiquinone Q A , the loosely bound ubiquinone Q B . After two subsequent flashes of light, Q B is reduced to ubiquinol Q B H 2 , with a semiquinone anion Q 3 B formed as an intermediate after the first flash. We studied Q B H 2 formation in chromatophores from Rb. sphaeroides mutants that carried ArgCIle substitution at sites 207 and 217 in the L-subunit. While Arg-L207 is 17 A î away from Q B , Arg-L217 is closer (9 A î ) and contacts the Q B -binding pocket. From the pH dependence of the charge recombination in the RC after the first flash, we estimated vG AB , the free energy difference between the Q 3 A Q B and Q A Q 3 B states, and pK 212 , the apparent pK of Glu-L212, a residue that is only 4 A î away from Q B . As expected, the replacement of positively charged arginines by neutral isoleucines destabilized the Q 3 B state in the L217RI mutant to a larger extent than in the L207RI one. Also as expected, pK 212 increased by V0.4 pH units in the L207RI mutant. The value of pK 212 in the L217RI mutant decreased by 0.3 pH units, contrary to expectations. The rate of the Q 3 A Q 3 B CQ A Q B H 2 transition upon the second flash, as monitored by electrometry via the accompanying changes in the 0005-2728 / 00 / $^see front matter ß 2000 Elsevier Science B.V. All rights reserved. PII: S 0 0 0 5 -2 7 2 8 ( 0 0 ) 0 0 1 1 0 -9
Biochemistry, 1997
It is generally accepted that electron transfer in bacterial photosynthesis is driven by the first singlet excited state of a special pair of bacteriochlorophylls (P*). We have examined the first steps of electron transfer in a mutant of the Rhodobacter sphaeroides reaction center in which charge separation from P* is dramatically slowed down. The results provide for the first time clear evidence that excitation of the monomeric bacteriochlorophyll in the active branch of the reaction center (B A ) drives ultrafast transmembrane electron transfer without the involvement of P*, demonstrating a new and efficient mechanism for solar energy transduction in photosynthesis. The most abundant charge-separated intermediate state probably is P + B A -, which is formed within 200 fs from B A * and decays with a lifetime of 6.5 ps into P + H A -. We also see evidence for the involvement of a B A + H A -state in the alternative pathway.
Biochemistry, 1997
It is generally accepted that electron transfer in bacterial photosynthesis is driven by the first singlet excited state of a special pair of bacteriochlorophylls (P*). We have examined the first steps of electron transfer in a mutant of the Rhodobacter sphaeroides reaction center in which charge separation from P* is dramatically slowed down. The results provide for the first time clear evidence that excitation of the monomeric bacteriochlorophyll in the active branch of the reaction center (B A) drives ultrafast transmembrane electron transfer without the involvement of P*, demonstrating a new and efficient mechanism for solar energy transduction in photosynthesis. The most abundant charge-separated intermediate state probably is P + B A-, which is formed within 200 fs from B A * and decays with a lifetime of 6.5 ps into P + H AWe We also see evidence for the involvement of a B A + H A-state in the alternative pathway.