On the site of action of plastocyanin in isolated chloroplasts

Adding plastocyanin to plastocyanin-depleted chloroplast particles, restored both their ability to catalyse the photoinduced electron transfer from ascorbate-DCIP to NADP, and to induce the photooxidation of cytochrome f. It is concluded, therefore, that plastocyanin mediates the photoinduced oxidation of cytochrome f, as previously suggested.     

THE WAVELENGTH DEPENDENCE OF MASSIVE CAROTENE SYNTHESIS IN DUNALIELLA BARDAWIL (CHLOROPHYCEAE)1

Dunaliella bardawil Ben-Amotz & Avron accumulates high concentrations of β-carotene when grown under high light intensity. The β-carotene is composed mainly of 9-cis and all-trans β-carotene. Accumulation of β-carotene and an increase in the ratio of the 9-cis to the all-trans isomer are strongly dependent on the light intensity under which the algae are cultivated but are independent of light quality within the photosynthetically active radiation range. Cells grown under continuous red (>645 nm) or white light of 500 W·m^?2 reach a value of about 32 pg β-carotene·cell^?1 and a ratio of 9-cis to all-trans β-carotene of around 2, whereas cells grown under low red or white light intensity of 25 W·m^?2 contain about 3 pg·cell^?1 and a ratio of isomers of around 0.3.     

Strength and co-operativity of contributions of surface salt bridges to protein stability

Many of the interactions that stabilize proteins are co-operative and cannot be reduced to a sum of pairwise interactions. Such interactions may be analysed by protein engineering methods using multiple thermodynamic cycles comprising wild-type protein and all combinations of mutants in the interacting residues. There is a triad of charged residues on the surface of barnase, comprising residues Asp8, Asp12 and Arg110, that interact by forming two exposed salt bridges. The three residues have been mutated to alanine to give all the single, double and triple mutants. The free energies of unfolding of wild-type and the seven mutant proteins have been determined and the results analysed to give the contributions of the residues in the two salt bridges to protein stability. It is possible to isolate the energies of forming the salt bridges relative to the solvation of the separated ions by water. In the intact triad, the apparent contribution to the stabilization energy of the protein of the salt bridge between Asp12 and Arg110 is -1.25 kcal mol-1, whereas that of the salt bridge between Asp8 with Arg110 is -0.98 kcal mol-1. The strengths of the two salt bridges are coupled: the energy of each is reduced by 0.77 kcal mol-1 when the other is absent. The salt-linked triad, relative to alanine residues at the same positions, does not contribute to the stability of the protein since the favourable interactions of the salt bridges are more than offset by other electrostatic and non-electrostatic energy terms. Salt-linked triads occur in other proteins, for example, haemoglobin, where the energy of only the salt-bridge term is important and so the coupling of salt bridges could be of general importance to the stability and function of proteins.     

Plasma membrane potential of the alga dunaliella, and its relation to osmoregulation

A fluorescent dye sensitive to membrane potential was used to follow the plasma-membrane potential in the unicellular halo-tolerant alga Dunaliella salina. The signal observed during dissipation of the plasma membrane potential by the addition of excess K⁺ and valinomycin, or a protonophore, was taken as a measure of the preexisting potential. A resting potential of −85 to −100 millivolts (negative inside) was calculated. Following a hypertonic shock, the plasma membrane was rapidly hyperpolarized. This hyperpolarization was transient, and the algae resumed their resting potential about 30 minutes after the shock. The resting plasma membrane potential was decreased by vanadate and is concluded to be generated mostly by the plasma membrane ATPase of Dunaliella. The transient hyperpolarization following a hypertonic shock indicates, therefore, a transient activation of the ATPase. This is further corroborated by a rapid transient decrease in the intracellular ATP following a hypertonic shock and its inhibition by vanadate. It is suggested that activation of the plasma membrane ATPase may be the trigger for osmoregulation in Dunaliella.     

Involvement of the Plasma Membrane ATPase in the Osmoregulatory Mechanism of the Alga Dunaliella salina

The unicellular halotolerant alga Dunaliella salina recovers normally from a hypertonic shock even when suspended in NaCl and buffer only. Furthermore, addition of Cu²⁺, valinomycin and KCl, or permeable ions such as methyltriphenylphosphonium or thiocyanate, do not affect the recovery. However, treatment with two specific inhibitors of the plasma membrane adenosine triphosphatase (ATPase), diethylstilbestrol, or vanadate, fully inhibit the recovery. The inhibition is manifested by the inability of the cells to both synthesize glycerol and return to their original volume. The inhibitions are nonlethal, reversible and equally effective in the dark or the light. Since the plasma membrane ATPase is the only enzyme known to be inhibited by both diethylstilbestrol and vanadate, it is concluded that its activity is essential for the recovery of Dunaliella from a hypertonic shock. Mechanisms by which the plasma membrane ATPase may participate in the activation of glycerol production in the algae are discussed.     

Properties of Photosynthetic Mutants Isolated from Euglena gracilis

Four different photosynthetic mutants of Euglena gracilis were characterized as to their lesions in photosynthetic electron transport. Two were defective around photosystem II: one, in electron transport on the oxidizing side of photosystem II, and the second lacked cytochrome 558. The location of the defect in the third mutant was concluded to be in the carbon fixation cycle, since it could catalyse both photosynthetic electron transport and photophosphorylation. The fourth mutant had a defect in its mechanism of photophosphorylation.