Site-Directed Mutagenesis of Threonine M222 and Tryptophan M252 in the Photosynthetic Reaction Center of Rhodobacter sphaeroides

Stilz, Hans Ulrich; Finkele, Ulrich; Holzapfel, W. B.; Lauterwasser, Christoph; Oesterhelt, Dieter

doi:10.1007/978-3-642-61297-8_26

Cited by 13 publications

(7 citation statements)

References 11 publications

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“…sphaeroides RC structures [12]. Several site-specific mutants at position M250 confirmed its proposed role for electron transfer and established its extraordinary importance for the binding of QA [40,41]. Only the aromatic residues phenylalanine and tyrosine can replace the tryptophan at position M250, only to a certain extent.…”

Section: Electron Transfermentioning

confidence: 81%

Structure of the photosynthetic reaction centre from Rhodobacter sphaeroides at 2.65 å resolution: cofactors and protein-cofactor interactions

et al. 1994

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Section: Electron Transfermentioning

confidence: 81%

Structure of the photosynthetic reaction centre from Rhodobacter sphaeroides at 2.65 å resolution: cofactors and protein-cofactor interactions

et al. 1994

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“…Nevertheless, experiments on other mutants in which similar amounts of QA were lost showed that the loss of quinone did not affect the kinetics of the initial electron-transfer step (Stilz et al, 1990). In addition, a heterogeneity at the site of YM210 cannot be excluded.…”

Section: Discussionmentioning

confidence: 98%

Role of tyrosine M210 in the initial charge separation of reaction centers of Rhodobacter sphaeroides

Finkele¹,

Lauterwasser²,

Zinth³

et al. 1990

Biochemistry

133

113

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ABSTRACT:Femtosecond spectroscopy was used in combination with site-directed mutagenesis to study the influence of tyrosine M210 (YM210) on the primary electron transfer in the reaction center of Rhodobacter sphaeroides. The exchange of YM210 to phenylalanine caused the time constant of primary electron transfer to increase from 3.5 f 0.4 ps to 16 f 6 ps while the exchange to leucine increased the time constant even more to 22 f 8 ps. The results suggest that tyrosine M210 is important for the fast rate of the primary electron transfer.x e primary photochemical event during photosynthesis of bacteriochlorophyll-(Bchl-) containing organisms is a lightinduced charge separation within a transmembrane protein complex called the reaction center (RC). The crystal structures of RC's from Rhodopseudomonas (Rps.) viridis and Rhodobacter (Rb.) sphaeroides have been solved to high resolution [reviewed in Deisenhofer and Michel (1989), Chang et al. (1986), Tiede et al. (1988), andRees et al. (1989)l. The RC from Rb. sphaeroides contains three protein subunits referred to as L, M, and H, according to their respective mobilities in SDS-polyacrylamide gels. Associated with the L and M subunits are the cofactors, consisting of four Bchl a, two bacteriopheophytin (Bph) a, one atom of non-heme ferrous iron, two quinones (QA and Qe), and in some species one carotenoid [reviewed in Parson (1987) and Feher et al.' Financial support was from the Deutsche Forschungsgemeinschaft,

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“…In addition to this purely structural role, it is probable that the protein plays some specific function in assisting the electron transport and in preventing wasteful back reactions. Aromatic residues such as Tyr M208 located between P, B A , and H A or Trp M250 situated in van der Waals contact distance of both H A and Q A ( 9 ; Figure ) have been pinpointed as playing a key role in the electron-transfer reactions ( − ). Structural rearrangements of the cofactors and of the RC protein are expected to contribute to the reorganization energy associated with long-range electron transfer.…”

mentioning

confidence: 99%

Conformational Heterogeneity of the Bacteriopheophytin Electron Acceptor H_A in Reaction Centers from Rhodopseudomonas viridis Revealed by Fourier Transform Infrared Spectroscopy and Site-Directed Mutagenesis

et al. 1999

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The light-induced Fourier transform infrared (FTIR) difference spectra corresponding to the photoreduction of either the HA bacteriopheophytin electron acceptor (HA-/HA spectrum) or the QA primary quinone (QA-/QA spectrum) in photosynthetic reaction centers (RCs) of Rhodopseudomonas viridis are reported. These spectra have been compared for wild-type (WT) RCs and for two site-directed mutants in which the proposed interactions between the carbonyls on ring V of HA and the RC protein have been altered. In the mutant EQ(L104), the putative hydrogen bond between the protein and the 9-keto C=O of HA should be affected by changing Glu L104 to a Gln. In the mutant WF(M250), the van der Waals interactions between Trp M250 and the 10a-ester C=O of HA should be modified. The characteristic effects of both mutations on the FTIR spectra support the proposed interactions and allow the IR modes of the 9-keto and 10a-ester C=O of HA and HA- to be assigned. Comparison of the HA-/HA and QA-/QA spectra leads us to conclude that the QA-/QA IR signals in the spectral range above 1700 cm-1 are largely dominated by contributions from the electrostatic response of the 10a-ester C=O mode of HA upon QA photoreduction. A heterogeneity in the conformation of the 10a-ester C=O mode of HA in WT RCs, leading to three distinct populations of HA, appears to be related to differences in the hydrogen-bonding interactions between the carbonyls of ring V of HA and the RC protein. The possibility that this structural heterogeneity is related to the observed multiexponential kinetics of electron transfer and the implications for primary processes are discussed. The effect of 1H/2H exchange on the QA-/QA spectra of the WT and mutant RCs shows that neither Glu L104 nor any other exchangeable carboxylic residue changes appreciably its protonation state upon QA reduction.

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Site-Directed Mutagenesis of Threonine M222 and Tryptophan M252 in the Photosynthetic Reaction Center of Rhodobacter sphaeroides

Cited by 13 publications

References 11 publications

Structure of the photosynthetic reaction centre from Rhodobacter sphaeroides at 2.65 å resolution: cofactors and protein-cofactor interactions

Structure of the photosynthetic reaction centre from Rhodobacter sphaeroides at 2.65 å resolution: cofactors and protein-cofactor interactions

Role of tyrosine M210 in the initial charge separation of reaction centers of Rhodobacter sphaeroides

Conformational Heterogeneity of the Bacteriopheophytin Electron Acceptor H_A in Reaction Centers from Rhodopseudomonas viridis Revealed by Fourier Transform Infrared Spectroscopy and Site-Directed Mutagenesis

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Site-Directed Mutagenesis of Threonine M222 and Tryptophan M252 in the Photosynthetic Reaction Center of Rhodobacter sphaeroides

Cited by 13 publications

References 11 publications

Structure of the photosynthetic reaction centre from Rhodobacter sphaeroides at 2.65 å resolution: cofactors and protein-cofactor interactions

Structure of the photosynthetic reaction centre from Rhodobacter sphaeroides at 2.65 å resolution: cofactors and protein-cofactor interactions

Role of tyrosine M210 in the initial charge separation of reaction centers of Rhodobacter sphaeroides

Conformational Heterogeneity of the Bacteriopheophytin Electron Acceptor HA in Reaction Centers from Rhodopseudomonas viridis Revealed by Fourier Transform Infrared Spectroscopy and Site-Directed Mutagenesis

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Conformational Heterogeneity of the Bacteriopheophytin Electron Acceptor H_A in Reaction Centers from Rhodopseudomonas viridis Revealed by Fourier Transform Infrared Spectroscopy and Site-Directed Mutagenesis