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Govindjee

Publications and source records attributed to Govindjee.

At least 55 records · Page 3Linked to original sources
PubMed1980

Effects of bulk pH and of monovalent and divalent cations on chlorophyll a fluorescence and electron transport in pea thylakoids.

Millimolar concentrations of monovalent cations enhance and divalent cations impede the redistribution (spill-over) of electronic excitation energy from Photosystem (PS) II to PS I in cation-depleted (sucrose-washed) thylakoids; this concept is based on chlorophyll a fluorescence and electron transport measurements over a narrow pH range around 7. We have tested the above concept in pea thylakoids over the pH range 5 to 9 by parallel measurements of various chlorophyll a fluorescence parameters (spectra, transients, and lifetimes at 77 K and 293 K, and polarization at 293 K) and of the rates of partial reactions of PSI and II. Our results provide the following information. (1) Mg2+ enhancement of fluorescence is maximum between 680 and 690 nm and minimum between 710 and 720 nm. (2) The optimum conditions for the observation of the Mg2+-induced enhancement of fluorescence are: wavelength of emission, 685 nm; concentratin of Mg2+, 10 mM, and pH, approximately 7.5. (3) Mg2+ decreases the efficiency of excitation redistribution from PS II to PS I over the pH range 6 to 9. (4) The antagonistic effects between Na+ and Mg2+ hold simultaneously for both the fluorescence intensity and lifetime, at physiological temperatures, only within the pH range 6 to 8. (5) Mg2+ enhances the light-limited electron transport rate through PS II in the pH range 5.4 to 8.2 and decreases that through PS I at pH 7.1 and 8.2. The % increase in PS II is, however, about twice the % decrease in PS I.

Cations↗
PubMed1979

Conformation and activity of chloroplast coupling factor exposed to low chemical potential of water in cells.

(1) Photophosphorylation, Ca2+-ATPase and Mg2+-ATPase activities of isolated chloroplasts were inhibited 55--65% when the chemical potential of water was decreased by dehydrating leaves to water potentials (psi w) of --25 bars before isolation of the plastids. The inhibition could be reversed in vivo by rehydrating the leaves. (2) These losses in activity were reflected in coupling factor (CF1) isolated from the leaves, since CF1 from leaves with low psi w had less Ca2+-ATPase activity than control CF1 and did not recouple phosphorylation in CF1-deficient chloroplasts. In contrast, CF1 from leaves having high psi w only partially recoupled phosphorylation by CF1-deficient chloroplasts from leaves havig low psi w. This indicated that low psi w affected chloroplast membranes as well as CF1 itself. (3) Coupling factor from leaves having low psi w had the same number of subunits, and the same electrophoretic mobility, and could be obtained with the same yields as CF1 from control leaves. However, direct measurements of fluorescence polarization, ultraviolet absorption, and circular dichroism showed that CF1 from leaves having low psi w differed from control CF1. The CF1 from leaves having low psi w also had decreased ability to bind fluorescent nucleotides (epsilon-ATP and epsilon-ADP). (4) Exposure of isolated CF1 to low psi w in vitro by preincubation in sucrose-containing media inhibited the Ca2+-ATPase activity of the protein in subsequent assays without sucrose. Inclusion of 5 or 10 mM Mg2+ in the preincubation medium markedly inhibited Ca2+-ATPase activity. (5) These results show that CF1 undergoes changes in cells which alter its phosphorylating ability. Since low cell psi w changed the spectroscopic properties but not other protein properties of CF1, the changes were most likely caused by altered confurn, photophosphorylation. The inhibition of ATPase activity in CF1 in vitro at low psi w and high ion concentration mimicked the change in activity seen in vivo.

Adenosine Triphosphatases↗
PubMed1978

Nuclear magnetic relaxation by the manganese in aqueous suspensions of chloroplasts.

Proton and oxygen-17 NMR relaxation rate (T1-1 and T2-1) data are presented for aqueous suspensions of dark-adapted chloroplasts. It is concluded from the dependence of the proton relaxation rates (PRR) upon Mn concentration that T1-1 and T2-1 are determined largely by the loosely bound Mn present in the chloroplast membranes. The frequency and temperature dependences of PRR are characteristic of Mn(II). The effects of oxidants (e.g., ferricyanide) and reductants (e.g., tetraphenylboron) on the PRR indicate that only about one-third to one-fourth of the loosely bound Mn is present in the dark-adapted chloroplasts as Mn(II), the remainder being in a higher oxidation state(s), probably Mn(III). The frequency dependence of the PRR for the chloroplast suspensions was fitted by a simplified form of the Solomon-Bloembergen-Morgan equations, and the following parameters were obtained: tauS = (1.1 +/- 0.1) X 10(-8) S; tauM = (2.2 +/- 0.2) X 10(-8) S; and B = (0.9 +/- 0.09) X 10(19). The oxygen-17 T1 and T2 data for suspensions before and after treatment with a detergent are consistent with the location of the manganese in the interior of the thylakoids. An analysis of the relaxation rates shows that the average lifetime of a water molecule inside a thylakoid is greater than 1 ms.

Chloroplasts↗
PubMed1977

Site of bicarbonate effect in Hill reaction. Evidence from the use of artificial electron acceptors and donors.

Using artificial electron donors and acceptors, it is shown here that the major HCO3- effect in the Hill reaction is after the "primary" electron acceptor (Q) of Photosystem II and before the site of action of 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (at the plastoquinone pool). Chloroplasts in the presence of both 3-(3',4'-dichlorophenyl)-1,1-dimethylurea, which blocks electron flow from the reduced primary acdeptor Q- to the plastoquinone pool, and silicomolybdate, which accepts electrons from Q-, show no significant bicarbonate stimulation of electron flow. However, a 6-7 fold stimulation is clearly observed when oxidized diaminodurene, as an electron acceptor, and dibromothymoquinone, as an inhibitor of electron flow beyond the plastoquinone pool, are used. In the same chloroplast preparation no measurable effect of bicarbonate is observed in a Photosystem I reaction as monitored by electron flow from reduced diaminodurene to methyl viologen in the presence of 3- (3',4'-dichlorophenyl)-1,1-dimethylurea. The insensitivity of the bicarbonate effect to uncouplers of photophosphorylation and the dependence of this effect on the presence of a weak acid anion and on external pH are also reported.

Bicarbonates↗
PubMed1977

The rise in chlorophyll a fluorescence yield and decay in delayed light emission in tris-washed chloroplasts in the 6-100 microseconds time range after an excitation flash.

Parallel measurements of the rise in chlorophyll a fluorescence yield and delayed light emission decay, after a 10 ns saturating excitation flash, have been made in tris (hydroxymethyl)aminomethane-washed chloroplasts. Various electron donor systems (Mn2+; ascorbate; reduced phenylenediamine and benzidine) were used in conjuction with different preillumination regimes to alter [P+-680], the oxidized form of the Photosystem II reaction center chlorophyll a. Conditions giving rise to high [p+ -680] resulted in only a small rise in fluorescence yield, an inhibition of a 6 microseconds component of delayed light emission. These results confirm the hypothesis that P+-680 acts as a quencher of fluorescence and that delayed light emission in the microsecond time range is due to the back reaction of P+-680 and Q-. (Q is the first "stable" electron acceptor of Photosystem II.) Two preillumination flashes are required before the full effect of Tris washing is observed in the delayed light emission decay and fluorescence yield rise; this suggests that a capacity to hold two charges exists between the Tris block and P+-680. Tris washing has no direct effect on the movement of electrons from Z (the first electron donor to P+-680. Finally, Mn2+ donates electrons to P+-680 via Z.

Ascorbic Acid↗
PubMed1976

Anthroyl stearate as a fluorescent probe of chloroplast membranes.

1. A reversible light-induced enhancement of the fluorescence of a "hydrophobic fluorophore", 12-(9-anthroyl)-stearic acid (anthroyl stearate), is observed with chloroplasts supporting phenazine methosulfate, cyclic or 1,1'-ethylene-2,2'-dipyridylium dibromide (Diquat) pseudo-cyclic electron flow; no fluorescence change is observed when methyl viologen or ferricyanide are used as electron acceptors. The stearic acid moiety of anthroyl stearate is important for its localization and fluorescence response in the thylakoid membrane, since structural analogs of anthroyl stearate lacking this group do not show the same response. 2. This effect is decreased under phosphorylating conditions (presence of ADP, Pi, Mg2+), and completely inhibited by the uncoupler of phosphorylation NH4Cl(5-10mM), as well as the ionophores nigericin and gramicidin-D (both at 5 - 10(-8)M). The MgCl2 concentration dependence of the anthroyl stearate enhancement effect is identical to that previously observed for cyclic photophosphorylation, as well as for the formation of a "high energy intermediate". The anthroyl stearate fluorescence enhancement is inhibited by increasing concentrations of ionophores in parallel with the decrease in ATP synthesis, but is essentially unaffected by specific inhibitors (Dio-9 and phlorizin) of photophosphorylation; thus, it appears that anthroyl stearate monitors a component of the "high energy state" of the thylakoid membrane rather than a terminal phosphorylation step. 3. The light-induced anthroyl stearate fluorescence enhancement is suggested to monitor a proton gradient in the energized chloroplast because (a) similar enhancement can be produced by sudden injection of hydrogen ions in a solution of anthroyl stearate; (b) when the proton gradient is dissipated by gramicidin or nigericin light-induced anthroyl stearate fllorescence is eliminated; (c) when the proton gradient is dissipated by tetraphenylboron, light-induced anthroyl stearate fluorescence decreases, and (d) light-induced anthroyl stearate fluorescence change as a function of pH is qualitatively similar to that observed with other probes for a proton gradient (e.g. 9-aminoacridine). Furthermore, anthroyl stearate does not monitor H+ uptake per se because (a) the pH dependence of H+ transport is different from that of the anthroyl stearate fluorescence change, and (b) tetraphenylboron, which does not inhibit H+ uptake, reduces anthroyl stearate fluorescence. Thus, anthroyl stearate appears to be a useful probe of a proton gradient supported by phenazine methosulfate of Diquat catalyzed electron flow and is the first "non-amine" fluorescence probe utilized for this purpose in chloroplasts.

Anthracenes↗
PubMed1976

Inhibition of the reoxidation of the secondary electron acceptor of photosystem II by bicarbonate depletion.

In bicarbonate-depleted chloroplasts, the chlorophyll a fluorescence decayed with a halftime of about 150 ms after the third flash, and appreciably faster after the first and second flash of a series of flashes given after a dark period. After the fourth to twentieth flashes, the decay was also slow. After addition of bicarbonate, the decay was fast after all the flashes of the sequence. This indicates that the bicarbonate depletion inhibits the reoxidation of the secondary acceptor R2- by the plastoquinone pool; R is the secondary electron acceptor of pigment system II, as it accepts electrons from the reduced form of the primary electron acceptor (Q-). This conclusion is consistent with the measurements of the DCMU (3-(3,4-dichlorophenyl)-),)-dimethylurea)- induced chlorophyll a fluorescence after a series of flashes in the presence and the absence of bicarbonate, if it is assumed that DCMU not only causes reduction of Q if added in the state QR-, but also if added in the state QT2-.

Bicarbonates↗
PubMed1976

A major site of bicarbonate effect in system II reaction. Evidence from ESR signal IIvf, fast fluorescence yield changes and delayed light emission.

In order to determine the major site of bicarbonate action in the electron transport complex of Photosystem II, the following experimental techniques were used: electron spin resonance measurements of Signal IIvf, measurements of chlorophyll a fluorescence yield rise and decay kinetics, and delayed light emission decay. From data obtained using these experimental techniques the following conclusions were made: (1) absence of bicarbonate causes a reversible inactivation of up to 40% of Photosystem II reaction center activity; (2) there is no significant effect of bicarbonate on electron flow from the charge accumulating S state to Z; (3) there is no significant effect of bicarbonate on electron flow from Z to P-680+; (4) electron flow from Q-- to the intersystem electron transport pool is inhibited by from 4- to 6-fold under bicarbonate depletion conditions.

Bicarbonates↗
PubMed1975

Water proton relaxation as a monitor of membrane-bound manganese in spinach chloroplasts.

First measurements of proton relaxation on chloroplast membranes are presented here. Experiments show that the water proton spin-lattice relaxation rate in chloroplast thylakoid membrane suspensions can be used to monitor membrane-bound manganese. The relaxation effect is reduced to 0.4 of its original value upon manganese extraction by washing with either alkaline Tris buffer or NH2OH/EDTA solution. Large increases in the proton relaxation rate are measured in the presence of reductants such as tetraphenylboron and NH2OH; oxidants such as potassium ferricyanide or 2,6-dichlorophenolindophenol lead to an decrease in this rate. These results suggest that manganese exists as a mixture of oxidation states in dark-adapted chloroplasts.

2,6-Dichloroindophenol↗