No abstract

Photosystem I has two branches of cofactors down which lightdriven electron transfer (ET) could potentially proceed, each consisting of a pair of chlorophylls (Chls) and a phylloquinone (PhQ). Forward ET from PhQ to the next ET cofactor (F X) is described by two kinetic components with decay times of Ϸ20 and Ϸ200 ns, which have been proposed to represent ET from PhQ B and PhQA, respectively. Immediately preceding each quinone is a Chl (ec3), which receives a H-bond from a nearby tyrosine. To decrease the reduction potential of each of these Chls, and thus modify the relative yield of ET within the targeted branch, this H-bond was removed by conversion of each Tyr to Phe in the green alga Chlamydomonas reinhardtii. Together, transient optical absorption spectroscopy performed in vivo and transient electron paramagnetic resonance data from thylakoid membranes showed that the mutations affect the relative amplitudes, but not the lifetimes, of the two kinetic components representing ET from PhQ to F X. The mutation near ec3 A increases the fraction of the faster component at the expense of the slower component, with the opposite effect seen in the ec3B mutant. We interpret this result as a decrease in the relative use of the targeted branch. This finding suggests that in Photosystem I, unlike type II reaction centers, the relative efficiency of the two branches is extremely sensitive to the energetics of the embedded redox cofactors.Chlamydomonas ͉ directionality ͉ photosynthetic reaction center ͉ pump-probe spectroscopy ͉ transient EPR P hotosynthetic reaction centers (RCs) are the membrane proteins responsible for the capture and storage of light energy in photosynthetic organisms. As with all of the RCs, Photosystem I (PS1) has a C2 symmetrical structure with two virtually identical branches of redox cofactors extending across the membrane (see Fig. 1). The x-ray crystal structure of PS1 from Thermosynechococcus elongatus (1) has allowed identification of amino acid residues interacting with the electron transfer (ET) cofactors, permitting the use of site-directed mutagenesis to investigate to what extent ET occurs in the two branches. Many of these studies (2-10) point toward the possibility that ET takes place in both branches of cofactors (A-branch and Bbranch; see Fig. 1). The data consistently show that the major fraction occurs in the A-branch and that the slow phase of ET from phylloquinone (PhQ) to F x is associated with this branch. The involvement of the B-branch is less clear, but there is mounting evidence to support the assignment of the fast component of PhQ to F x ET to this branch. If ET does indeed occur in both branches in PS1, this behavior would be remarkably different from the type II RCs. In purple bacterial RCs, for example, it is well known that initial ET is biased almost exclusively toward the A-branch, probably because the B-branch quinone is a mobile electron and proton carrier in this type of RC. The strong biasing of the ET in the type II RCs makes it difficult to investigate the fac...

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 ...