chlorophyll-a and ethylenediamine

Water binding to several Mg(2+) chelates, ethylenediamine, ethylenediamine-N,N'-diacetate, porphyrin, chlorophyll a and bacteriochlorophyll a, to form five- and six-coordinate complexes is studied by means of density functional theory. The results obtained for magnesium chelates are compared with the properties of the respective aqua complexes and the influence of the permittivity of environment on the ligand binding energies is discussed. Although the most common coordination number of Mg(2+) is six, in the tetrapyrrolic chelates it is reduced to five because the accommodation of the sixth water ligand results in no gain in energy. This is in line with the experimental observations made for coordination of chlorophylls in vivo. The binding between Mg(2+) and water is mostly of electrostatic nature, which is supported by the finding that its energy is correlated both with the electron density of the chelator and with electrostatic potential determined at the ligand binding site.

Topics: Bacteriochlorophyll A; Chelating Agents; Chlorophyll; Chlorophyll A; Edetic Acid; Ethylenediamines; Ligands; Magnesium; Models, Molecular; Porphyrins; Quantum Theory; Static Electricity; Water

Chemical modification of plastocyanin was carried out using ethylenediamine plus a water-soluble carbodiimide, which has the effect of replacing a negatively charged carboxylate group with a positively charged amino group at pH 6-8. The conditions were adjusted to produce a series of singly and doubly modified forms of plastocyanin. Differences in charge configuration allowed separation of these forms on a Pharmacia fast protein liquid chromatograph using a Mono Q anion exchange column. These forms were used to study the interaction of plastocyanin with its reaction partner cytochrome f. The rate of cytochrome f oxidation was progressively inhibited upon incorporation of increasing numbers of ethylenediamine moieties indicating a positively charged binding site on cytochrome f. However, differential inhibition was obtained for the various singly modified forms allowing mapping of the binding site on plastocyanin. The greatest inhibition was found for forms modified at negatively charged residues Nos. 42-45 and Nos. 59-61 which comprise a negative patch surrounding Tyr-83. In contrast, the form modified at residue No. 68, on the opposite side of the globular plastocyanin molecule, showed the least inhibition. It can be concluded that the binding site for cytochrome f is located in the vicinity of residues Nos. 42-45 and Nos. 59-61. Modification of plastocyanin at residues Nos. 42-45 showed no effect on the rate of P-700+ reduction, suggesting that these residues are not involved in the binding of Photosystem I. However, an increase in the rate of P-700+ reduction was observed for plastocyanins modified at residue No. 68 or Nos. 59-61, which is consistent with the idea that the reaction domain of Photosystem I is negatively charged and Photosystem I binds at the top of the molecule and accepts electrons via His-87 in plastocyanin. These results raise the possibility that plastocyanin can bind both cytochrome f and Photosystem I simultaneously. The effect of ethylenediamine modification on the formal potential of plastocyanin was also examined. The formal potential of control plastocyanin was found to be +372 +/- 5 mV vs. normal hydrogen electrode at pH 7. All modified forms showed a positive shift in formal potential. Singly modified forms showed increases in formal potentials between +8 and +18 mV with the largest increases being observed for plastocyanins modified at residues Nos. 42-45 or Nos. 59-61.

Topics: Amino Acid Sequence; Chlorophyll; Cytochromes; Cytochromes f; Ethylenediamines; Molecular Sequence Data; Oxidation-Reduction; Plant Proteins; Plastocyanin; Protein Conformation

Plastocyanin treated with tetranitromethane was nitrated at a single location, Tyr-83. Tyr-83 and its neighboring negative charges have been implicated as a binding site for positively charged redox agents (Chapman, S.K., Watson, A.D. and Sykes, A.G. (1983) J. Chem. Soc. Dalton Trans. 1983, 2543-2548). No effect was observed on either the plastocyanin midpoint redox potential or its reaction kinetics with P-700+ and cytochrome f. This makes nitration an ideal spectroscopic probe for monitoring changes in the environment of Tyr-83. The pKa of the nitrotyrosine was 8.6 and 8.3 for reduced and oxidized plastocyanin, respectively, indicating that the charge on the copper atom is 'felt' at Tyr-83. The high pKa value for both forms indicates that Tyr-83 is in a negatively charged environment, near residues Nos. 42-45 and Nos. 59-61. The extinction of the nitrotyrosine chromophore at 360 nm was not affected by a change in redox state. However, the ellipticity of this transition was greater for the oxidized form, indicating that environment of Tyr-83 is dependent upon the charge on the copper atom. This suggests an electrostatically driven conformational change at Tyr-83. A conformational change at Tyr-83 could regulate the binding of plastocyanin with its reaction partners in order to promote smooth electron transport.

Topics: Chlorophyll; Chromatography, High Pressure Liquid; Circular Dichroism; Computers; Cytochromes; Cytochromes f; Ethylenediamines; Isoelectric Point; Kinetics; Nitrates; Plant Proteins; Plastocyanin; Protein Conformation; Spectrophotometry, Ultraviolet; Tyrosine; X-Ray Diffraction