PubMed Health⌕ Search

Biomedical subjects

H J van Gorkom

Publications and source records attributed to H J van Gorkom.

At least 19 recordsLinked to original sources
PubMed2004

Energy and electron transfer in photosystem II reaction centers with modified pheophytin composition.

Energy and electron transfer in Photosystem II reaction centers in which the photochemically inactive pheophytin had been replaced by 13(1)-deoxo-13(1)-hydroxy pheophytin were studied by femtosecond transient absorption-difference spectroscopy at 77 K and compared to the dynamics in untreated reaction center preparations. Spectral changes induced by 683-nm excitation were recorded both in the Q(Y) and in the Q(X) absorption regions. The data could be described by a biphasic charge separation. In untreated reaction centers the major component had a time constant of 3.1 ps and the minor component 33 ps. After exchange, time constants of 0.8 and 22 ps were observed. The acceleration of the fast phase is attributed in part to the redistribution of electronic transitions of the six central chlorin pigments induced by replacement of the inactive pheophytin. In the modified reaction centers, excitation of the lowest energy Q(Y) transition produces an excited state that appears to be localized mainly on the accessory chlorophyll in the active branch (B(A) in bacterial terms) and partially on the active pheophytin H(A). This state equilibrates in 0.8 ps with the radical pair. B(A) is proposed to act as the primary electron donor also in untreated reaction centers. The 22-ps (pheophytin-exchanged) or 33-ps (untreated) component may be due to equilibration with the secondary radical pair. Its acceleration by H(B) exchange is attributed to a faster reverse electron transfer from B(A) to. After exchange both and are nearly isoenergetic with the excited state.

Dose-Response Relationship, Radiation↗
PubMed2001

Kinetics of electron transfer from Q(a) to Q(b) in photosystem II.

The oxidation kinetics of the reduced photosystem II electron acceptor Q(A)(-) was investigated by measurement of the chlorophyll fluorescence yield transients on illumination of dark-adapted spinach chloroplasts by a series of saturating flashes. Q(A)(-) oxidation depends on the occupancy of the "Q(B) binding site", where this reaction reduces plastoquinone to plastoquinol in two successive photoreactions. The intermediate, one-electron-reduced plastosemiquinone anion Q(B)(-) remains tightly bound, and its reduction by Q(A)(-) may proceed with simple first-order kinetics. The next photoreaction, in contrast, may find the Q(B) binding site occupied by a plastoquinone, a plastoquinol, or neither of the two, resulting in heterogeneous Q(A)(-) oxidation kinetics. The assumption of monophasic Q(B)(-) reduction kinetics is shown to allow unambiguous decomposition of the observed multiphasic Q(A)(-) oxidation. At pH 6.5 the time constant for Q(A)(-) oxidation was found to be 0.2-0.4 ms with Q(B) in the site, 0.6-0.8 ms with Q(B)(-) in the site, 2-3 ms when the site is empty and Q(B) has to bind first, and of the order of 0.1 s if the site is temporarily blocked by the presence of Q(B)H(2) or other low-affinity inhibitors such as carbonyl cyanide m-chlorophenylhydrazone (CCCP). Effects of pH and H(2)O/D(2)O exchange were found to be remarkably nonspecific. No influence of the S-states could be demonstrated.

Electron Transport↗
PubMed2001

Pigment organization and their interactions in reaction centers of photosystem II: optical spectroscopy at 6 K of reaction centers with modified pheophytin composition.

Photosystem II reaction centers (RC) with selectively exchanged pheophytin (Pheo) molecules as described in [Germano, M., Shkuropatov, A. Ya., Permentier, H., Khatypov, R. A., Shuvalov, V. A., Hoff, A. J., and van Gorkom, H. J. (2000) Photosynth. Res. 64, 189-198] were studied by low-temperature absorption, linear and circular dichroism, and triplet-minus-singlet absorption-difference spectroscopy. The ratio of extinction coefficients epsilon(Pheo)/epsilon(Chl) for Q(Y) absorption in the RC is approximately 0.40 at 6 K and approximately 0.45 at room temperature. The presence of 2 beta-carotenes, one parallel and one perpendicular to the membrane plane, is confirmed. Absorption at 670 nm is due to the perpendicular Q(Y) transitions of the two peripheral chlorophylls (Chl) and not to either Pheo. The "core" pigments, two Pheo and four Chl absorb in the 676-685 nm range. Delocalized excited states as predicted by the "multimer model" are seen in the active branch. The inactive Pheo and the nearby Chl, however, mainly contribute localized transitions at 676 and 680 nm, respectively, although large CD changes indicate that exciton interactions are present on both branches. Replacement of the active Pheo prevents triplet formation, causes an LD increase at 676 and 681 nm, a blue-shift of 680 nm absorbance, and a bleach of the 685 nm exciton band. The triplet state is mainly localized on the Chl corresponding to B(A) in purple bacteria. Both Pheo Q(Y) transitions are oriented out of the membrane plane. Their Q(X) transitions are parallel to that plane, so that the Pheos in PSII are structurally similar to their homologues in purple bacteria.

Chlorophyll↗
PubMed2001

Ultrahigh field MAS NMR dipolar correlation spectroscopy of the histidine residues in light-harvesting complex II from photosynthetic bacteria reveals partial internal charge transfer in the B850/His complex.

Low-temperature 15N and 13C CP/MAS (cross-polarization/magic angle spinning) NMR has been used to analyze BChl-histidine interactions and the electronic structure of histidine residues in the light-harvesting complex II (LH2) of Rhodopseudomonas acidophila. The histidines were selectively labeled at both or one of the two nitrogen sites of the imidazole ring. The resonances of histidine nitrogens that are interacting with B850 BChl a have been assigned. Specific 15N labeling confirmed that it is the tau-nitrogen of histidines which is ligated to Mg2+ of B850 BChl molecules (beta-His30, alpha-His31). The pi-nitrogens of these Mg2+-bound histidines were found to be protonated and may be involved in hydrogen bond interactions. Comparison of the 2-D MAS NMR homonuclear (13C-13C) dipolar correlation spectrum of [13C6,15N3]-histidines in the LH2 complex with model systems in the solid state reveals two different classes of electronic structures from the histidines in the LH2. In terms of the 13C isotropic shifts, one corresponds to the neutral form of histidine and the other resembles a positively charged histidine species. 15N-13C double-CP/MAS NMR data provide evidence that the electronic structure of the histidines in the neutral BChl a/His complexes resembles the positive charge character form. While the Mg...15N isotropic shift confirms a partial positive charge transfer, its anisotropy is essentially of the lone pair type. This provides evidence that the hybridization structure corresponding to the neutral form of the imidazole is capable of "buffering" a significant amount of positive charge.

Culture Media↗
PubMed2001

Secondary stabilization reactions and proton-coupled electron transport in photosystem II investigated by electroluminescence and fluorescence spectroscopy.

The oxidized primary electron donor in photosystem II, P(680)(+), is reduced in several phases, extending over 4 orders of magnitude in time. Especially the slower phases may reflect the back-pressure exerted by water oxidation and provide information on the reactions involved. The kinetics of secondary electron-transfer reactions in the microseconds time range after charge separation were investigated in oxygen-evolving thylakoids suspended in H2O or D2O. Flash-induced changes of chlorophyll fluorescence yield and electric field-induced recombination luminescence were decomposed into contributions from oxidation states S(0), S(1), S(2), and S(3) of the oxygen-evolving complex and interpreted in terms of stabilization kinetics of the initial charge-separated state S(j)Y(Z)P(680)(+)Q(A)(-)Q(B). In approximately 10% of the centers, only charge recombination took place. Otherwise, no static heterogeneity was involved in the microsecond reduction of P(680)(+) by Y(Z) (stabilization) or Q(A)(-) (recombination). The recombination component in active centers occurs mainly upon charge separation in S(3), and, in the presence of D2O, in S(2) as well and is tentatively attributed to the presence of Y(Z)(ox)S(j-1) in equilibrium with Y(Z)S(j). A 20-30 micros stabilization occurs in all S-states, but to different extents. Possible mechanisms for this component are discussed. D2O was found to decrease: (i) the rate of the reaction Y(Z)(ox)S(1) to Y(Z)S(2), (ii) the equilibrium constant between P680(+)Y(Z)S(2) and P(680)Y(Z)(ox)S(2), (iii) the rate of the slow phase of P(680)(+) reduction for the S(3) --> S(0) transition, and (iv) the rate of electron transfer from Q(A)(-) to Q(B) /Q(B)(-). The increased 'miss probability' in D2O is due to (iii).

Benzoquinones↗
PubMed2000

Photochemically induced nuclear spin polarization in reaction centers of photosystem II observed by 13C-solid-state NMR reveals a strongly asymmetric electronic structure of the P680(.+) primary donor chlorophyll.

We report (13)C magic angle spinning NMR observation of photochemically induced dynamic nuclear spin polarization (photo-CIDNP) in the reaction center (RC) of photosystem II (PS2). The light-enhanced NMR signals of the natural abundance (13)C provide information on the electronic structure of the primary electron donor P(680) (chlorophyll a molecules absorbing around 680 nm) and on the p(z) spin density pattern in its oxidized form, P(680)(.+). Most centerband signals can be attributed to a single chlorophyll a (Chl a) cofactor that has little interaction with other pigments. The chemical shift anisotropy of the most intense signals is characteristic for aromatic carbon atoms. The data reveal a pronounced asymmetry of the electronic spin density distribution within the P(680)(.+). PS2 shows only a single broad and intense emissive signal, which is assigned to both the C-10 and C-15 methine carbon atoms. The spin density appears shifted toward ring III. This shift is remarkable, because, for monomeric Chl a radical cations in solution, the region of highest spin density is around ring II. It leads to a first hypothesis as to how the planet can provide itself with the chemical potential to split water and generate an oxygen atmosphere using the Chl a macroaromatic cycle. A local electrostatic field close to ring III can polarize the electronic charge and associated spin density and increase the redox potential of P(680) by stabilizing the highest occupied molecular orbital, without a major change of color. This field could be produced, e.g., by protonation of the keto group of ring V. Finally, the radical cation electronic structure in PS2 is different from that in the bacterial RC, which shows at least four emissive centerbands, indicating a symmetric spin density distribution over the entire bacteriochlorophyll macrocycle.

Carbon Isotopes↗
PubMed2000

Exploring the calcium-binding site in photosystem II membranes by solid-state (113)Cd NMR.

Calcium (Ca(2+)) is an essential cofactor for photosynthetic oxygen evolution. Although the involvement of Ca(2+) at the oxidizing side of photosystem II of plants has been known for a long time, its ligand interactions and mode of action have remained unclear. In the study presented here, (113)Cd magic-angle spinning solid-state NMR spectroscopy is used to probe the Ca(2+)-binding site in the water-oxidizing complex of (113)Cd(2+)-substituted PS2. A single NMR signal 142 ppm downfield from Cd(ClO(4))(2).2H(2)O was recorded from Cd(2+) present at the Ca(2+)-binding site. The anisotropy of the signal is small, as indicated by the absence of spinning side bands. The signal intensity is at its maximum at a temperature of -60 degrees C. The line width of the proton signal in a WISE (wide-line separation) two-dimensional (1)H-(113)Cd NMR experiment demonstrates that the signal arises from Cd(2+) in a solid and magnetically undisturbed environment. The chemical shift, the small anisotropy, and the narrow line of the (113)Cd NMR signal provide convincing evidence for a 6-fold coordination, which is achieved partially by oxygen and partially by nitrogen or chlorine atoms in otherwise a symmetric octahedral environment. The absence of a (113)Cd signal below -70 degrees C suggests that the Ca(2+)-binding site is close enough to the tetramanganese cluster to be affected by its electron spin state. To our knowledge, this is the first report for the application of solid-state NMR in the study of the membrane-bound PS2 protein complex.

Binding Sites↗
PubMed1999

Activating anions that replace Cl- in the O2-evolving complex of photosystem II slow the kinetics of the terminal step in water oxidation and destabilize the S2 and S3 states.

Photosystem II, the multisubunit protein complex that oxidizes water to O2, requires the inorganic cofactors Ca2+ and Cl- to exhibit optimal activity. Chloride can be replaced functionally by a small number of anionic cofactors (Br-, NO3-, NO2-, I-), but among these anions, only Br- is capable of restoring rates of oxygen evolution comparable to those observed with Cl-. UV absorption difference spectroscopy was utilized in the experiments described here as a probe to monitor donor side reactions in photosystem II in the presence of Cl- or surrogate anions. The rate of the final step of the water oxidation cycle was found to depend on the activating anion bound at the Cl- site, but the kinetics of this step did not limit the light-saturated rate of oxygen evolution. Instead, the lower oxygen evolution rates supported by surrogate anions appeared to be correlated with an instability of the higher oxidation states of the oxygen-evolving complex that was induced by addition of these anions. Reduction of these states takes place not only with I- but also with NO2- and to a lesser extent even with NO3- and Br- and is not related to the ability of these anions to bind at the Cl- binding site. Rather, it appears that these anions can attack higher oxidation states of the oxygen evolving complex from a second site that is not shielded by the extrinsic 17 and 23 kDa polypeptides and cause a one-electron reduction. The decrease of the oxygen evolution rate may result from accumulated damage to the reaction center protein by the one-electron oxidation product of the anion.

Anions↗
PubMed1998

S-state dependence of chloride binding affinities and exchange dynamics in the intact and polypeptide-depleted O2 evolving complex of photosystem II.

The Cl- binding properties in the successive oxidation states of the O2 evolving complex of photosystem II were investigated by measurements of UV absorbance changes, induced by a series of saturating flashes, that monitor manganese oxidation state transitions. In dark-adapted, intact photosystem II, Cl- can be replaced by NO3- in minutes, in an exchange reaction that depends on the NO3- concentration and that is not rate-limited by dissociation of Cl- from its binding site. Preillumination of dark-adapted photosystem II by one or two flashes accelerated the NO3- substitution reaction by an order of magnitude. A quantitative analysis of the Cl- concentration dependence of UV absorbance changes, measured in photosystem II preparations depleted of extrinsic 17 and 23 kDa polypeptides, shows that the Cl- binding properties of photosystem II change with the oxidation state of the oxygen evolving complex. Although the affinity for the individual S-states could not be determined with precision, it is shown that the affinity is an order of magnitude lower in the S2 state than in the S1 state. Comparison of the results obtained using intact photosystem II and preparations depleted of the 17 and 23 kDa extrinsic polypeptides suggests that these proteins constitute a diffusion barrier, which prevents fast equilibration of the Cl- binding site with the medium, but does not change the Cl- affinity of the binding site.

Binding Sites↗
PubMed1997

Bicarbonate may Be required for ligation of manganese in the oxygen-evolving complex of photosystem II.

It was previously shown in the photosystem II membrane preparation DT-20 that photoxidation of the oxygen-evolving manganese cluster was blocked by 0.1 mM formate, unless 0.2 mM bicarbonate was present as well [Wincencjusz, H., Allakhverdiev, S. I., Klimov, V. V., and Van Gorkom, H. J. (1996) Biochim. Biophys. Acta 1273, 1-3]. Here it is shown by measurements of EPR signal II that oxidation of the secondary electron donor, YZ, is not inhibited. However, the reduction of is greatly slowed and occurs largely by back reaction with reduced acceptors. Bicarbonate is shown to prevent the loss of fast electron donation to . The release of about one or two free Mn2+ per photosystem II during formate treatment, and the fact that these effects are mimicked by Mn-depletion, suggests that formate may act by replacing a bicarbonate which is essential for Mn binding. Irreversible light-induced rebinding in an EPR-silent form of Mn2+ that was added to Mn-depleted DT-20 was indeed found to depend on the presence of bicarbonate, as did the reconstitution in such material of both the fast electron donation to and the UV absorbance changes characteristic of a functional oxygen-evolving complex. It is concluded that bicarbonate may be an essential ligand of the functional Mn cluster.

Bicarbonates↗
PubMed1997

The photosynthetic oxygen evolving complex requires chloride for its redox state S2-->S3 and S3-->S0 transitions but not for S0-->S1 or S1-->S2 transitions.

The Cl- requirement in the redox cycle of the oxygen-evolving complex (OEC) was determined by measurements of flash-induced UV absorbance changes in Cl(-)-depleted and Cl(-)-reconstituted photosystem II membranes. On the first flash after dark adaptation the spectrum and amplitude of those changes, known to reflect the oxidation of MnIII to MnIV on the S1-->S2 transition, were the same in the presence or absence of Cl-. On the second and later flashes, however, absorbance changes in Cl(-)-depleted samples revealed only electron transfer from tyrosine to quinone which reversed slowly in the dark by charge recombination and did not produce the S3-state. A rapid method was developed to remove Cl- after producing the S3-state by two flashes. The lifetime of the S3-state was found to be unaffected by Cl(-)-depletion, in contrast to the 20-fold stabilization of the S2 lifetime by Cl- removal, and the Cl(-)-depleted S3-state did not proceed to S0 on flash illumination. However, when the same Cl(-)-depletion procedure was applied after producing the S0-state by three flashes, further advance to S2 by two additional flashes was not impaired by the absence of Cl-. The requirement for Cl- only on the S2-->S3 and S3-->S0 transitions can be rationalized by the hypothesis that Cl- is required for electron transfer between manganese ions within the oxygen-evolving complex.

Chlorides↗
PubMed1987

Electron transfer in the water-oxidizing complex of Photosystem II.

An overview is presented of secondary electron transfer at the electron donor side of Photosystem II, at which ultimately two water molecules are oxidized to molecular oxygen, and the central role of manganese in catalyzing this process is discussed. A powerful technique for the analysis of manganese redox changes in the water-oxidizing mechanism is the measurement of ultraviolet absorbance changes, induced by single-turnover light flashes on dark-adapted PS II preparations. Various interpretations of these ultraviolet absorbance changes have been proposed. Here it is shown that these changes are due to a single spectral component, which presumably is caused by the oxidation of Mn(III) to Mn(IV), and which oscillates with a sequence +1, +1, +1, -3 during the so-called S0----S1----S2----S3----S0 redox transitions of the oxygen-evolving complex. This interpretation seems to be consistent with the results obtained with other techniques, such as those on the multiline EPR signal, the intervalence Mn(III)-Mn(IV) transition in the infrared, and EXAFS studies. The dark distribution of the S states and its modification by high pH and by the addition of low concentrations of certain water analogues are discussed. Finally, the patterns of proton release and of electrochromic absorbance changes, possibly reflecting the change of charge in the oxygen-evolving system, are discussed. It is concluded that nonstoichiometric patterns must be considered, and that the net electrical charge of the system probably is the highest in state S2 and the lowest in state S1.

Chlorophyll↗
PubMed1981

External electric field effects on prompt and delayed fluorescence in chloroplasts.

An electric field pulse was applied to a suspension of osmotically swollen spinach chloroplasts after illumination with a saturating flash in the presence of DCMU. In addition to the stimulation of delayed fluorescence by the electric field, discovered by Arnold and Azzi (Arnold, W.A. and Azzi, R. (1971) Photochem. Photobiol. 14, 233-240) a sudden drop in fluorescence yield was observed. The kinetics of this fluorescence change were identical to those of the integrated delayed fluorescence emission induced by the pulse. The S-state dependence of the stimulated emission was very similar to that of the normal luminescence. We assume that the membrane potential generated by the pulse changes the activation energy for the back reaction in Photosystem II. On this basis, and making use of data we obtained earlier from electrochromic absorbance changes induced by the pulse, the kinetics of the field-induced prompt and delayed fluorescence changes, and also the amplitude of the fluorescence decrease, which was about 12% for a nearly saturating pulse, are explained. Our results indicate that in those reaction centers where a decrease of the activation energy occurs the effect of a pulse can be quite spectacular: the back reaction, which normally takes seconds, is completed in a few hundred microseconds when a sufficiently strong pulse is applied. Measurements of the polarization of the stimulate luminescence supported the interpretation given above. Only 2.8% of the back reaction was found to proceed via transition of reexcited chlorophyll to the ground state, both during the field pulse and in the absence of the field.

Chloroplasts↗
PubMed1981

Quantum efficiency and antenna size of photosystems II alpha, II beta and I in tobacco chloroplasts.

Reaction center concentrations were determined in chloroplasts of tobacco, cv John William's Broadleaf, and its mutants Su/su and Su/su var. Aurea. Quantum yields of the primary reactions of Photosystems I, II alpha and II beta (Melis, A. and Homann, P.H. (1975) Photochem. Photobiol. 21, 431--437) were obtained by measurement of their rate constants and the absorbed energy, under conditions where all three photosystems operated simultaneously and produced almost irreversibly a single charge separation. The concentrations and reaction rates of the photosystems were different in chloroplasts from the wild type and the mutants, but in chloroplasts of each type of plant used essentially all quanta absorbed by chlorophyll caused a charge separation in PS I, PS II alpha or PS II beta. since the quantum efficiency of each photosystem was close to one, kinetic differences between the photosystems and between different kinds of chloroplasts were only due to differences in antenna size. From the rate constants the number of chlorophyll molecules in the antenna of each photosystem could be calculated. It is argued that PS II alpha and PS II beta must be different, independent structures.

Chlorophyll↗
PubMed1981

Chlorophyll composition of photosystems II alpha, II beta and I in tobacco chloroplasts.

The antenna composition of the Photosystems II alpha, II beta and I was studied in tobacco chloroplasts. Absorbance spectra, recorded at 4 K, were analyzed for the wild type and the mutants Su/su and Su/su var. Aurea, containing higher concentrations of the photosystems. With chloroplasts of Su/su we measured the action spectra of the three photosystems from 625 to 690 nm. Above 675 nm absorption by Photosystem I dominated. This system had a maximum at 678 nm and a shoulder at 660 nm. Of the long-wavelength chlorophyll a forms, absorbing at 690, 697 and 705 nm at 4 K, which are generally assigned to Photosystem I, the 697 nm form occurred in an amount of four molecules per reaction center of Photosystem I in each type of chloroplast. The Photosystem II alpha spectrum was characterized by maxima at 650 and 672 nm, showing clearly the participation of the chlorophyll a and b containing light-harvesting complex. In the mutants the light-harvesting complex has a chlorophyll a to chlorophyll b ratio of more than 1; the amount of the 672 nm chlorophyll a was normal, whereas the amount of chlorophyll b was markedly decreased in the mutants relative to the wild type. The Photosystem II beta spectrum mainly consisted of a band at 683 nm.

Chlorophyll↗
PubMed1980

Electrochromic absorbance changes in spinach chloroplasts induced by an external electrical field.

Absorbance changes induced by electrical field pulses were studied in osmotically swollen spinach chloroplasts. The results and their interpretation on the basis of the geometry and electrical properties of the material may be summarized as follows: 1. The spherical vesicles, 'blebs', formed upon dilution of a chloroplast suspension consist of only a single membrane, while part of the thylakoid system remains concentrated in a few patches on its surface. 2. When an electrical field pulse is applied, an up to 3000-fold enhanced field is built up in the membrane, with a time constant of about 20 mus. From this the specific capacitance of the bleb wall was found to be 2 microF . CM-2. 3. The electrical field in the membrane causes several absorbance changes of the photosynthetic pigments with different dependencies on the direction of polarization of the measuring light. Some of these are due to field-induced changes in orientation, in particular of chlorophyll alpha, and have a relaxation time of less than 100 mus. Most of the absorbance changes directly reflect the kinetics of the membrane potential and can be ascribed to electrochromic shifts of photosynthetic pigments, mainly of carotenoids. 4. The carotenoid absorbance changes depend quadratically on the membrane potential; an apparent saturation at high applied field strengths is ascribed to dielectric breakdown at a membrane potential of about 1 V. 5. All carotenoids in the membrane contribute to the absorbance changes induced by an externally applied field, whereas the well-known light-induced electrochromic absorbance change at 518 nm is mainly caused by a minor fraction of permanently polarized and spectrally red-shifted carotenoids. A computer simulation showed that this interpretation quantitatively explains the results and requires no unreasonable values of the various parameters involved.

Carotenoids↗