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Bacteriopheophytin

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Neal W Woodbury – 1st expert on this subject based on the ideXlab platform

  • electron transfer in bacterial reaction centers with the photoactive Bacteriopheophytin replaced by a bacteriochlorophyll through coordinating ligand substitution
    Biochemistry, 2016
    Co-Authors: Rafael G Saer, Thomas J Beatty, Neal W Woodbury

    Abstract:

    The influence of amino acid substitutions at position M214 (M-subunit, residue 214) on the rate and pathway of electron transfer involving the Bacteriopheophytin cofactor, HA, in a bacterial photosynthetic reaction center has been explored in a series of Rhodobacter sphaeroides mutants. The M214 leucine (L) residue of the wild type was replaced with histidine (H), glutamine (Q), and asparagine (N), creating the mutants M214LH, M214LQ, and M214LN, respectively. As has been reported previously for M214LH, each of these mutations resulted in a bacteriochlorophyll molecule in place of a Bacteriopheophytin in the HA pocket, forming so-called β-type mutants (in which the HA cofactor is called βA). In addition, these mutations changed the properties of the surrounding protein environment in terms of charge distribution and the amino acid side chain volume. Electron transfer reactions from the excited primary donor P to the acceptor QA were characterized using ultrafast transient absorption spectroscopic techniqu…

  • the protein environment of the Bacteriopheophytin anion modulates charge separation and charge recombination in bacterial reaction centers
    Journal of Physical Chemistry B, 2013
    Co-Authors: Rafael G Saer, Thomas J Beatty, Neal W Woodbury

    Abstract:

    The kinetics and pathway of electron transfer has been explored in a series of reaction center mutants from Rhodobacter sphaeroides, in which the leucine residue at M214 near the Bacteriopheophytin cofactor in the A-branch has been replaced with methionine, cysteine, alanine, and glycine. These amino acids have substantially different volumes, both from each other and, except for methionine, from the native leucine. Though the mutation site of M214 is close to the Bacteriopheophytin cofactor, which is involved in the electron transfer, none of the mutations alter the cofactor composition of the reaction center and the primary charge separation reaction is essentially undisturbed. However, the kinetics of electron transfer from HA– → QA becomes both slower and substantially heterogeneous in three of the four mutants. The decreased HA– → QA electron transfer rate allows charge recombination between P+ and HA– to compete with the forward reaction, resulting in a drop in the overall yield of charge separation…

  • b side electron transfer in a rhodobacter sphaeroides reaction center mutant in which the b side monomer bacteriochlorophyll is replaced with Bacteriopheophytin low temperature study and energetics of charge separated states
    Journal of Physical Chemistry B, 2002
    Co-Authors: Evaldas Katilius, Zivile Katiliene, Aileen K W Taguchi, Neal W Woodbury

    Abstract:

    The mutation, HL(M182), in the Rhodobacter sphaeroides reaction center (RC) results in the replacement of the monomer bacteriochlorophyll (BChl) on the inactive side (B side) of the RC with a Bacteriopheophytin (BPheo; the new cofactor is referred to as o B ). In o B -containing RCs, the first excited state of the primary donor (P * ) decays with an accelerated time constant of 2.6 ′ 0.1 ps at room temperature as compared to 3.1 ′ 0.2 ps in wild-type (WT) RCs. At low temperatures, P * decay is essentially the same in the HL(M182) mutant and WT RCs: 1.4 ′ 0.1 ps at 77 K and 1.1 ′ 0.1 ps at 9 K. The yield of the charge-separated P + o B – state decreases from 35% at room temperature to 12% at 77 and 9 K. The decreased P + o B – yield is apparently due to the fact that the rate of the charge separation along the A side of the RC at low temperature increases, while the rate along the B side remains essentially unchanged. From measurements of the long-lived fluorescence decay at room temperature, the standard free energy of the P + o B – state is estimated to be about 0.16 ′ 0.04 eV below P * . Given a difference between the midpoint potentials of BChl and BPheo of 0.26 ′ 0.03 V, the standard free energy of the P + B B – state in WT RC is estimated to be 0.1 ′ 0.07 eV above P * .

Christine Kirmaier – 2nd expert on this subject based on the ideXlab platform

  • temperature dependence of electron transfer to the m side Bacteriopheophytin in rhodobacter capsulatus reaction centers
    Journal of Physical Chemistry B, 2008
    Co-Authors: Jessica I Chuang, Dewey Holten, Steven G Boxer, Christine Kirmaier

    Abstract:

    Subpicosecond time-resolved absorption measurements at 77 K on two reaction center (RC) mutants of Rhodobacter capsulatus are reported. In the DLL mutant the D helix of the M subunit has been substituted with the D helix from the L subunit, and in the DLL-FYLFM mutant, three additional mutations are incorporated that facilitate electron transfer to the M side of the RC. In both cases the helix swap has been shown to yield isolated RCs that are devoid of the native Bacteriopheophytin electron carrier HL (Chuang, J. I.; Boxer, S. G.; Holten, D.; Kirmaier, C. Biochemistry 2006, 45, 3845−3851). For DLL, depending whether the detergent Deriphat 160-C or N-lauryl-N,N-dimethylamine-N-oxide (LDAO) is used to suspend the RCs, the excited state of the primary electron donor (P*) decays to the ground state with an average lifetime at 77 K of 330 or 170 ps, respectively; however, in both cases the time constant obtained from single-exponential fits varies markedly as a function of the probe wavelength. These findings…

  • high yield of m side electron transfer in mutants of rhodobacter capsulatus reaction centers lacking the l side Bacteriopheophytin
    Biochemistry, 2006
    Co-Authors: Jessica I Chuang, Dewey Holten, Steven G Boxer, Christine Kirmaier

    Abstract:

    We present studies on a series of photosynthetic reaction center (RC) mutants created in the background of the Rhodobacter capsulatusDLL mutant, in which the D helix of the M subunit has been substituted with that from the L subunit. Previous work on the DLL mutant in chromatophore preparations showed that RCs assembled without the Bacteriopheophytin HL electron acceptor and performed no charge separation following light absorption. We have successfully isolated poly-His-tagged D LL RCs by using the detergent Deriphat 160-C and shown that the RCs are devoid of HL. The excited state of the primary electron donor, P*, is found to have a lifetime of 180 ( 20 ps and to decay exclusively (>95%) via internal conversion to the ground state, with no evidence for formation of any charge-separated intermediates. By additional mutation in the DLL background of two residues that affect the P/P + oxidation potential and one that facilitates M-side electron transfer, we achieve an unprecedented 70% yield of P + HM – , more than doubling the highest yield of this state achieved previously. This result underscores the importance of the relative free energies of P* and the charge-separated states in governing the rates and yields of electron transfer in bacterial RCs and provides a basis for systematically investigating M-side electron transfer without any competition from the native L-side pathway.

  • resonance raman characterization of reaction centers with an asp residue near the photoactive Bacteriopheophytin
    Biochemistry, 1998
    Co-Authors: Christine Kirmaier, Dewey Holten, David F Bocian

    Abstract:

    Qy-excitation resonance Raman (RR) studies are reported for a series of Rhodobacter capsulatus reaction centers (RCs) containing mutations at L-polypeptide residue 121 near the photoactive Bacteriopheophytin (BPhL). The studies focus on the electronic/structural perturbations of BPhL induced by replacing the native Phe with an Asp residue. Earlier work has shown that the electron-transfer properties of F(L121)D RCs are closely related to those of RCs in which BPhL is replaced by bacteriochlorophyll (BChl) (beta-type RCs) or by pheophytin. In addition to the F(L121)D single mutant, RR studies were performed on the F(L121)D/E(L104)L double mutant, which additionally removes the hydrogen bond between BPhL and the native Glu L104 residue. The vibrational signatures of BPhL in the single and double mutants containing Asp L121 are compared with one another and with those of BPhL in both wild-type and F(L121)L RCs. The replacement of the aromatic Phe residue with Leu has no discernible effect on the vibrational properties of BPhL, a finding in concert with the previously reported absence of an effect of the mutation on the electron-transfer characteristics of the RC. In contrast, replacement of Phe with Asp significantly perturbs the vibrational characteristics of BPhL, and in a manner most consistent with Asp L121 being deprotonated and negatively charged. The negative charge of the carboxyl group of Asp L121 interacts with the pi-electron system of BPhL in a relatively nonspecific fashion, diminishing the contribution of charge-separated resonance forms of the C9-keto group to the electronic structure of the cofactor. The presence of a negative charge near BPhL is consistent with the known photochemistry of F(L121)D RCs, which indicates that the free energy of P+BPhL- is substantially higher than in wild-type RCs.

A J Hoff – 3rd expert on this subject based on the ideXlab platform

  • the effect of exchange of Bacteriopheophytin a with plant pheophytin a on charge separation in y m210 w mutant reaction centers of rhodobacter sphaeroides at low temperature
    Biochimica et Biophysica Acta, 2003
    Co-Authors: Anatoli Ya Shkuropatov, P Gast, Sieglinde Neerken, Hjalmar P Permentier, Rik De Wijn, Kristiane A Schmidt, V A Shuvalov, Thijs J Aartsma, A J Hoff

    Abstract:

    Abstract The Bacteriopheophytin a molecules at the HA and HB binding sites of reaction centers (RCs) of the Y(M210)W mutant of Rhodobacter sphaeroides were chemically exchanged with plant pheophytin a. The Y(M210)W mutation slows down the formation of HA−, presumably by raising the free energy level of the P+BA− state above that of P* due to increasing the oxidation potential of the primary electron donor P and lowering the reduction potential of the accessory bacteriochlorophyll BA. Exchange of the Bacteriopheophytins with pheophytin a on the contrary lowers the redox potential of HA, inhibiting its reduction. A combination of the mutation and pigment exchange was therefore expected to make the A-side of the RC incapable of electron transfer and cause the excited state P* to deactivate directly to the ground state or through the B-side, or both. Time-resolved absorption difference spectroscopy at 10 K on the RCs that were modified in this way showed a lifetime of P* lengthened to about 500 ps as compared to about 200 ps measured in the original Y(M210)W RCs. We show that the decay of P* in the pheophytin-exchanged preparations is accompanied by both return to the ground state and formation of a new charge-separated state, the absorption difference spectrum of which is characterized by bleachings at 811 and 890 nm. This latter state was formed with a time constant of ca. 1.7 ns and a yield of about 30%, and lasted a few nanoseconds. On the basis of spectroscopic observations these bands at 811 and 890 nm are tentatively attributed to the presence of the P+BB− state, where BB is the accessory bacteriochlorophyll in the “inactive” B-branch of the cofactors. The BB molecules in Y(M210)W RCs are suggested to be spectrally heterogeneous, absorbing in the Qy region at 813 or 806 nm. The results are discussed in terms of perturbation of the free energy level of the P+BB− state and absorption properties of the BB bacteriochlorophyll in the mutant RCs due to a long-range effect of the Y(M210)W mutation on the protein environment of the BB binding pocket.

  • efficient conditions for the photoaccumulation of ha in the photosynthetic reaction centre of rhodobacter sphaeroides r26 with uniformly labelled Bacteriopheophytin monitored by fourier transform infrared difference spectroscopy
    Vibrational Spectroscopy, 1999
    Co-Authors: T A Egorovazachernyuk, P Gast, Andre Remy, Anatoly Ya Shkuropatov, A J Hoff, Klaus Gerwert, Huub J M De Groot

    Abstract:

    Abstract The native Bacteriopheophytins (BPheo) a at sites HA and HB in the reaction centre (RC) of Rhodobacter sphaeroides R26 have been exchanged with uniformly 13 C , 15 N labelled BPheo a. Exchange at the HA site was >50% as monitored with light-induced FTIR-difference spectroscopy indicated by the shift of the band at 1590 cm−1. The photoreduction of HA has been monitored by light-induced FTIR-difference spectroscopy at 22°C in the presence of reductant and mediator: either sodium dithionite and cytochrome c, or sodium dithionite and dyes (potassium indigotetrasulfonate, neutral red). New experimental conditions are described for HA− photoaccumulation, and light-induced FTIR-difference spectra characteristic of HA− are reported. The HA−/HA FTIR-difference spectra of Rb. sphaeroides R26 RCs in which uniformly 13 C , 15 N labelled BPheo a replaces the photoactive BPheo will allow the BPheo modes to be assigned and to be discriminated from those of the RC protein.

  • photosynthetic rcs of rb sphaeroides r26 with uniformly 13 c 15 n labelled Bacteriopheophytin monitored with ftir and solid state mas nmr
    , 1998
    Co-Authors: Tatiana Egorovazachernyuk, P Gast, Andre Remy, Anatoly Ya Shkuropatov, A J Hoff, Klaus Gerwert, Huub J M De Groot

    Abstract:

    Although the crystal structure of the photosynthetic reaction center (RC) complex has been determined in great detail, with up to 2.65 A resolution [1, 2] several fundamental questions regarding the charge separation mechanism and electron pathway are still unresolved. For example, the precise role of the protein-cofactor interactions in assisting electron transport and prevention of wasteful back reactions is still poorly understood. The RC of Rhodobacter (Rb.) sphaeroides contains two Bacteriopheophytins (BPheo) that are chemically identical but only one, HA, is known to function as an intermediary electron acceptor while the other, HB, appears not to be actively involved in electron transfer [3,4]. Selective isotopic labelling of the cofactors provides a tool for probing the RC structure and the cofactor-protein binding sites using complementary spectroscopic techniques like MAS NMR and FTIR [5-7]. We recently introduced a novel concept of multispin labelling and assignment of 13C CP/MAS NMR resonances of uniformly 13C labelled pheophytin (Pheo) a in Rb. sphaeroides R26 RCs [8]. The present investigation aims at providing a next step into the study of the protein-Bacteriopheophytin (BPheo) interactions in Rb. sphaeroides R26 RCs through the uniform isotope labelling of BPheo (Fig. 1) followed by 1-D 13C CP/MAS NMR and FTIR difference spectroscopy. Photoaccumulation of HA- under reversible conditions required for FTIR studies was reported for the first time by Nabedryk et al. [9], however the procedure for the photoaccumulation of HA appearedto be difficult to handle. Therefore a new method for efficient generating HA- in a film of Rb. sphaeroides R26 RCs was developed [10].