Spectroscopic properties, amino acid sequence, electron transfer kinetics, and crystal structures of the oxidized (at 1.7 Å resolution) and reduced form (at 1.8 Å resolution) of a novel plastocyanin from the fern Dryopteris crassirhizoma are presented. Kinetic studies show that the reduced form of Dryopteris plastocyanin remains redox-active at low pH, under conditions where the oxidation of the reduced form of other plastocyanins is inhibited by the protonation of a solvent-exposed active site residue, His 87 (equivalent to His 90 in Dryopteris plastocyanin). The x-ray crystal structure analysis of Dryopteris plastocyanin reveals -stacking between Phe 12 and His 90 , suggesting that the active site is uniquely protected against inactivation. Like higher plant plastocyanins, Dryopteris plastocyanin has an acidic patch, but this patch is located closer to the solvent-exposed active site His residue, and the total number of acidic residues is smaller. In the reactions of Dryopteris plastocyanin with inorganic redox reagents, the acidic patch (the "remote" site) and the hydrophobic patch surrounding His 90 (the "adjacent" site) are equally efficient for electron transfer. These results indicate the significance of the lack of protonation at the active site of Dryopteris plastocyanin, the equivalence of the two electron transfer sites in this protein, and a possibility of obtaining a novel insight into the photosynthetic electron transfer system of the first vascular plant fern, including its molecular evolutionary aspects. This is the first report on the characterization of plastocyanin and the first three-dimensional protein structure from fern plant.

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The crystal structure of plastocyanin from a green alga, Ulva pertusa, has been determined at 1.6-Å resolution. At its copper site, U. pertusa plastocyanin has a distorted tetrahedral coordination geometry similar to other plastocyanins. ) bond of poplar and C. reinhardtii plastocyanins by 0.14 and 0.20 Å, respectively. As a result of structural differences, U. pertusa plastocyanin has a less distorted geometry than the other plastocyanins. Thus, the cupric geometry is finely tuned by the interactions between residues 85 and 88 and between residues 83 and 88. This result implies that the copper site is more flexible than reported formerly and that the rack mechanism would be preferable to the entatic theory. The HisMet loop may regulate the electron transfer rate within the complex between plastocyanin and cytochrome f.

The X-ray crystal structures of oxidized and reduced plastocyanin obtained from the fern Dryopteris crassirhizoma have been determined at 1.7 and 1.8 A resolution, respectively. The fern plastocyanin is unique in the longer main chain composed of 102 amino acid residues and in the unusual pH dependence due to the pi-pi stacking interaction around the copper site [Kohzuma, T., et al. (1999) J. Biol. Chem. 274, 11817-11823]. Here we report the structural comparison between the fern plastocyanin and other plastocyanins from cyanobacteria, green algae, and other higher plants, together with the structural changes of fern plastocyanin upon reduction. Glu59 hydrogen bonds to the OH of Tyr83, which is thought to be a possible conduit for electrons, in the oxidized state. However, it moves away from Tyr83 upon reduction like poplar plastocyanin.

Changes in the Chl a/b ratio, electron transport, electron carriers, and photosystem stoichiometries were examined in rice leaves in the present study. The Chl a/b ratio is known to decrease gradually from the top to the bottom leaves, indicating a increase in the abundance of LHC II relative to the reaction center complexes of the two photosystems. We used juvenile rice canopy and obtained the following results: (1) the photosynthetic activity and Chl content per leaf area decreased from the top to the bottom leaves, the Chl a/b ratio also declined from 3.7 to 3.0; (2) when determined on the basis of Chl content, C-550 and Cyt f content decreased, but there was no loss of P-700, Consequently, the PS II/PS I ratio significantly decreased; (3) on the basis of mmol Chl, the levels of Cyt f dramatically decreased and, therefore, no loss was observed for whole chain electron transport per Cyt f; and (4) the percentage abundance of PS IIa increased, but the rate constants of Q A photoreduction and P-700 photooxidation gradually decreased. From these results, we hypothesize that there is a compensatory relationship between the decline in the Chl a/b ratio and that in the PS II/PS I ratio in the lower leaves in rice seedlings.