Mutation Induced Modulation of Hydrogen Bonding to P700 Studied Using FTIR Difference Spectroscopy

Wang, Ruili; Sivakumar, Velautham; Redding, Kevin; Hastings, Gary

doi:10.1021/bi034230c

Cited by 17 publications

(42 citation statements)

References 25 publications

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“…2). Very recently, we have provided further evidence that suggests that the interpretation proposed by Witt et al cannot be correct (Wang et al, 2003).…”

Section: Introductionmentioning

confidence: 66%

“…If the 1639(ÿ)/1655(1) cm ÿ1 difference band of WT (in S. 6803) is due to the 13 1 keto C¼O of P A that is strongly H-bonded to Thr-A743, then removal of this H-bond should cause a dramatic upshift of this difference band. We have used FTIR DS to analyze a mutant from C. reinhardtii in which Thr-A739 (Thr-A743 in S. elongatus) is changed to alanine, and we find that the spectra cannot be consistently interpreted if the 1639(ÿ)/1655(1) cm ÿ1 difference band of WT is due to the 13 1 keto C¼O of P A (Wang et al, 2003). In contrast, Witt et al (2002) also produced FTIR DS for mutants from C. reinhardtii in which Thr-A739 was mutated to Y, H, or V. These authors, however, did come to the conclusion that the 1639(ÿ)/1655(1) cm ÿ1 difference band of WT is due to the 13 1 keto C¼O of P A .…”

Section: The Ester C5o Regionmentioning

confidence: 99%

“…Recently, different groups have obtained (P700 1 -P700) FTIR DS using PS I particles from Synechocystis (S.) sp. 6803 (Breton, 2001;Breton et al, 1999Breton et al, , 2002Hastings et al, 2001; Kim and Barry, 2000;Kim et al, 2001) and C. reinhardtii (Breton, 2001;Hastings et al, 2001;Redding et al, 1998;Wang et al, 2003;Witt et al, 2002). From these studies different interpretations of the FTIR DS were proposed: 1), On the basis of the observation of multiple band shifts upon specific 13 4 methyl deuteration of ;68% of the Chls in PS I, it was suggested that the 13 3 ester (Kim and Barry, 2000) and 13 1 keto (Kim et al, 2001) carbonyl (C¼O) modes of the Chls of P700 are heterogeneously distributed in frequency (the mode frequency is different in different PS I subpopulations).…”

Section: Introductionmentioning

confidence: 99%

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FTIR Difference Spectroscopy in Combination with Isotope Labeling for Identification of the Carbonyl Modes of P700 and P700+ in Photosystem I

Wang

Sivakumar

Johnson

et al. 2004

Biophysical Journal

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Room temperature, light induced (P700(+)-P700) Fourier transform infrared (FTIR) difference spectra have been obtained using photosystem I (PS I) particles from Synechocystis sp. PCC 6803 that are unlabeled, uniformly (2)H labeled, and uniformly (15)N labeled. Spectra were also obtained for PS I particles that had been extensively washed and incubated in D(2)O. Previously, we have found that extensive washing and incubation of PS I samples in D(2)O does not alter the (P700(+)-P700) FTIR difference spectrum, even with approximately 50% proton exchange. This indicates that the P700 binding site is inaccessible to solvent water. Upon uniform (2)H labeling of PS I, however, the (P700(+)-P700) FTIR difference spectra are considerably altered. From spectra obtained using PS I particles grown in D(2)O and H(2)O, a ((1)H-(2)H) isotope edited double difference spectrum was constructed, and it is shown that all difference bands associated with ester/keto carbonyl modes of the chlorophylls of P700 and P700(+) downshift 4-5/1-3 cm(-1) upon (2)H labeling, respectively. It is also shown that the ester and keto carbonyl modes of the chlorophylls of P700 need not be heterogeneously distributed in frequency. Finally, we find no evidence for the presence of a cysteine mode in our difference spectra. The spectrum obtained using (2)H labeled PS I particles indicates that a negative difference band at 1698 cm(-1) is associated with at least two species. The observed (15)N and (2)H induced band shifts strongly support the idea that the two species are the 13(1) keto carbonyl modes of both chlorophylls of P700. We also show that a negative difference band at approximately 1639 cm(-1) is somewhat modified in intensity, but unaltered in frequency, upon (2)H labeling. This indicates that this band is not associated with a strongly hydrogen bonded keto carbonyl mode of one of the chlorophylls of P700.

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“…2). Very recently, we have provided further evidence that suggests that the interpretation proposed by Witt et al cannot be correct (Wang et al, 2003).…”

Section: Introductionmentioning

confidence: 66%

Section: The Ester C5o Regionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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FTIR Difference Spectroscopy in Combination with Isotope Labeling for Identification of the Carbonyl Modes of P700 and P700+ in Photosystem I

Wang

Sivakumar

Johnson

et al. 2004

Biophysical Journal

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“…The assignment of the other bands in the FTIR DS is somewhat controversial. However, all of the changes can be rationalized within the framework of a mutation-induced charge redistribution, in which some positive charge on P B is relocated onto P A (40).…”

Section: Figure 3: P 700mentioning

confidence: 99%

Mutation of the Putative Hydrogen-Bond Donor to P₇₀₀ of Photosystem I

Lucas

Konovalova

et al. 2004

Biochemistry

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The primary electron donor of photosystem I (PS1), called P 700 , is a heterodimer of chlorophyll (Chl) a and a′. The crystal structure of photosystem I reveals that the chlorophyll a′ (P A ) could be hydrogenbonded to the protein via a threonine residue, while the chlorophyll a (P B ) does not have such a hydrogen bond. To investigate the influence of this hydrogen bond on P 700 , PsaA-Thr739 was converted to alanine to remove the H-bond to the 13 1 -keto group of the chlorophyll a′ in Chlamydomonas reinhardtii. The PsaA-T739A mutant was capable of assembling active PS1. Furthermore the mutant PS1 contained approximately one chlorophyll a′ molecule per reaction center, indicating that P 700 was still a Chl a/a′ heterodimer in the mutant. However, the mutation induced several band shifts in the visible P 700 + -P 700 absorbance difference spectrum. Redox titration of P 700 revealed a 60 mV decrease in the P 700 /P 700 + midpoint potential of the mutant, consistent with loss of a H-bond. Fourier transform infrared (FTIR) spectroscopy indicates that the ground state of P 700 is somewhat modified by mutation of ThrA739 to alanine. Comparison of FTIR difference band shifts upon P 700 + formation in WT and mutant PS1 suggests that the mutation modifies the charge distribution over the pigments in the P 700 + state, with ∼14-18% of the positive charge on P B in WT being relocated onto P A in the mutant. 1 H-electron-nuclear double resonance (ENDOR) analysis of the P 700 + cation radical was also consistent with a slight redistribution of spin from the P B chlorophyll to P A , as well as some redistribution of spin within the P B chlorophyll. High-field electron paramagnetic resonance (EPR) spectroscopy at 330-GHz was used to resolve the g-tensor of P 700 + , but no significant differences from wild-type were observed, except for a slight decrease of anisotropy. The mutation did, however, provoke changes in the zero-field splitting parameters of the triplet state of P 700 ( 3 P 700 ), as determined by EPR. Interestingly, the mutation-induced change in asymmetry of P 700 did not cause an observable change in the directionality of electron transfer within PS1.In higher plants and green algae, the photosynthetic reaction occurs within two large pigment-protein complexes located in the thylakoid membranes of the chloroplast: photosystems I and II (PS1 and PS2). 1 These two systems exemplify the type 1 ("iron-sulfur") and type 2 ("quinone") reaction centers, respectively, based on their terminal electron acceptors. In oxygenic phototrophs, PS2 and PS1 act in tandem to oxidize water and reduce NADP + , but there are several anoxygenic photosynthetic bacteria that use only one type of reaction center protein.All known reaction centers share a similar structural motif with a core of reaction centers composed of two similar or identical integral membrane subunits, to which the various redox cofactors are bound. Both the protein structural motifs and the cofactor arrangements are characterized by a pseudo-C 2 symmetry ...

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“…Gibasiewicz et al, using femtosecond spectroscopy at low temperature, characterized the exciton coupling in the RC (45). FTIR and EPR have been used to characterize the influence of the mutations on the charge distribution in P700 1 (43,44,46,47). Optical and ADMR measurements have been used to characterize the differences in the optical spectra of the RCs, including triplet-singlet spectra (41)(42)(43)(44).…”

Section: Introductionmentioning

confidence: 99%

Ultrafast Transient Absorption Studies on Photosystem I Reaction Centers from Chlamydomonas reinhardtii. 2: Mutations near the P700 Reaction Center Chlorophylls Provide New Insight into the Nature of the Primary Electron Donor

Holzwarth

Müller

Niklas

et al. 2006

Biophysical Journal

144

113

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The energy transfer and charge separation kinetics in several core Photosystem I particles of Chlamydomonas reinhardtii with point mutations around the PA and PB reaction center chlorophylls (Chls) have been studied using ultrafast transient absorption spectroscopy in the femtosecond to nanosecond time range to characterize the influence on the early electron transfer processes. The data have been analyzed in terms of kinetic compartment models. The adequate description of the transient absorption kinetics requires three different radical pairs in the time range up to approximately 100 ps. Also a charge recombination process from the first radical pair back to the excited state is present in all the mutants, as already shown previously for the wild-type (Müller, M. G., J. Niklas, W. Lubitz, and A. R. Holzwarth. 2003. Biophys. J. 85:3899-3922; and Holzwarth, A. R., M. G. Müller, J. Niklas, and W. Lubitz. 2005. J. Phys. Chem. B. 109:5903-59115). In all mutants, the primary charge separation occurs with the same effective rate constant within the error limits as in the wild-type (>>350 ns(-1)), which implies an intrinsic rate constant of charge separation of <1 ps(-1). The rate constant of the secondary electron transfer process is slowed down by a factor of approximately 2 in the mutant B-H656C, which lacks the ligand to the central metal of Chl PB. For the mutant A-T739V, which breaks the hydrogen bond to the keto carbonyl of Chl PA, only a slight slowing down of the secondary electron transfer is observed. Finally for mutant A-W679A, which has the Trp near the PA Chl replaced, either no pronounced effect or, at best, a slight increase on the secondary electron transfer rate constants is observed. The effective charge recombination rate constant is modified in all mutants to some extent, with the strongest effect observed in mutant B-H656C. Our data strongly suggest that the Chls of the PA and PB pair, constituting what is traditionally called the "primary electron donor P700", are not oxidized in the first electron transfer process, but rather only in the secondary electron transfer step. We thus propose a new electron transfer mechanism for Photosystem I where the accessory Chl(s) function as the primary electron donor(s) and the A0 Chl(s) are the primary electron acceptor(s). This new mechanism also resolves in a straightforward manner the difficulty with the previous mechanism, where an electron would have to overcome a distance of approximately 14 A in <1 ps in a single step. If interpreted within a scheme of single-sided electron transfer, our data suggest that the B-branch is the active branch, although parallel A-branch activity cannot be excluded. All the mutations do affect to a varying extent the energy difference between the reaction center excited state RC* and the first radical pair and thus affect the rate constant of charge recombination. It is interesting to note that the new mechanism proposed is in fact analogous to the electron transfer mechanism in Photosystem II, where the accessory Chl also ...

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Mutation Induced Modulation of Hydrogen Bonding to P700 Studied Using FTIR Difference Spectroscopy

Cited by 17 publications

References 25 publications

FTIR Difference Spectroscopy in Combination with Isotope Labeling for Identification of the Carbonyl Modes of P700 and P700+ in Photosystem I

FTIR Difference Spectroscopy in Combination with Isotope Labeling for Identification of the Carbonyl Modes of P700 and P700+ in Photosystem I

Mutation of the Putative Hydrogen-Bond Donor to P₇₀₀ of Photosystem I

Ultrafast Transient Absorption Studies on Photosystem I Reaction Centers from Chlamydomonas reinhardtii. 2: Mutations near the P700 Reaction Center Chlorophylls Provide New Insight into the Nature of the Primary Electron Donor

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Mutation Induced Modulation of Hydrogen Bonding to P700 Studied Using FTIR Difference Spectroscopy

Cited by 17 publications

References 25 publications

FTIR Difference Spectroscopy in Combination with Isotope Labeling for Identification of the Carbonyl Modes of P700 and P700+ in Photosystem I

FTIR Difference Spectroscopy in Combination with Isotope Labeling for Identification of the Carbonyl Modes of P700 and P700+ in Photosystem I

Mutation of the Putative Hydrogen-Bond Donor to P700 of Photosystem I

Ultrafast Transient Absorption Studies on Photosystem I Reaction Centers from Chlamydomonas reinhardtii. 2: Mutations near the P700 Reaction Center Chlorophylls Provide New Insight into the Nature of the Primary Electron Donor

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Mutation of the Putative Hydrogen-Bond Donor to P₇₀₀ of Photosystem I