The buffering capacity of the internal phase of thylakoids and the magnitude of the pH changes inside under flashing light

Junge, Wolfgang; Ausländer, Winfried; McGeer, Allison; Runge, Thomas M.

doi:10.1016/0005-2728(79)90175-0

Cited by 161 publications

(85 citation statements)

References 27 publications

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“…It is noted that in vivo the buffer capacity of the lumen of chloroplasts was estimated to be 0.8-1.0 mM (pH 6.4-8.1) and suggested to be due mostly to stationary groups such as proteins and lipid head groups (34), and not to mobile buffers such as MES added here at low concentrations to minimize the bulk pH changes. In vivo the role of HCO − 3 for the activity of PSII may therefore be even more pronounced than observed during the experiments shown in Fig.…”

Section: Discussion Hco −mentioning

confidence: 94%

Mobile hydrogen carbonate acts as proton acceptor in photosynthetic water oxidation

Koroidov

Shevela

Shutova

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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Cyanobacteria, algae, and plants oxidize water to the O 2 we breathe, and consume CO 2 during the synthesis of biomass. Although these vital processes are functionally and structurally well separated in photosynthetic organisms, there is a long-debated role for CO 2 /HCO − 3 in water oxidation. Using membrane-inlet mass spectrometry we demonstrate that HCO − 3 acts as a mobile proton acceptor that helps to transport the protons produced inside of photosystem II by water oxidation out into the chloroplast's lumen, resulting in a light-driven production of O 2 and CO 2 . Depletion of HCO − 3 from the media leads, in the absence of added buffers, to a reversible down-regulation of O 2 production by about 20%. These findings add a previously unidentified component to the regulatory network of oxygenic photosynthesis and conclude the more than 50-y-long quest for the function of CO 2 / HCO − 3 in photosynthetic water oxidation.O xygenic photosynthesis in cyanobacteria, algae, and higher plants leads to the reduction of atmospheric CO 2 to energy-rich carbohydrates. The electrons needed for this process are extracted in a cyclic, light-driven process from water that is split into dioxygen (O 2 ) and protons. This reaction is catalyzed by a penta-μ-oxo bridged tetra-manganese calcium cluster (Mn 4 CaO 5 ) within the oxygen-evolving complex (OEC) of photosystem II (PSII) (1-4). The possible roles of inorganic carbon, C i ðC i = CO 2 ; H 2 CO 3 ; HCO − 3 ; CO 2− 3 Þ, in this process have been a controversial issue ever since Otto Warburg and Günter Krippahl (5) reported in 1958 that oxygen evolution by PSII strictly depends on CO 2 and therefore has to be based on the photolysis of H 2 CO 3 ("Kohlensäure") and not of water. These first experiments were indirect and, as became apparent later, were wrongly interpreted (6-8). Several research groups followed up on these initial results and identified two possible sites of C i interaction within PSII (reviewed in refs. 9-12). Functional and spectroscopic studies showed that HCO − 3 facilitates the reduction of the secondary plastoquinone electron acceptor (Q B ) of PSII by participating in the protonation of Q 2− B . Binding of HCO − 3 (or CO 2− 3 ) to the nonheme Fe between the quinones Q A and Q B was recently confirmed by X-ray crystallography (3,13,14). Despite this functional role at the acceptor side, the very tight binding of HCO − 3 to this site makes it impossible for the activity of PSII to be affected by changing the C i level of the medium; instead inhibitors such as formate need to be added to induce the acceptor-side effect (15). Consequently, the watersplitting electron-donor side of PSII has also been studied intensively (for recent reviews, see refs. 11 and 12). Although a tight binding of C i near the Mn 4 CaO 5 cluster is excluded on the basis of X-ray crystallography (3, 14), FTIR spectroscopy (16), and mass spectrometry (17, 18), the possibility that a weakly bound HCO − 3 affects the activity of PSII at the donor side remains a viable option (reviewed i...

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Section: Discussion Hco −mentioning

confidence: 94%

Mobile hydrogen carbonate acts as proton acceptor in photosynthetic water oxidation

Koroidov

Shevela

Shutova

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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“…Flash-induced charge transfer across the thylakoid membrane was detected by electrochromic absorption changes of intrinsic pigments at a wavelength of 522 nm [26,27], proton release into the suspending medium by the dye Cresol red [24], and proton intake from the thylakoid lumen by Neutral red [28]. The pH-transients were calculated as the difference of two original recordings obtained with and without the respective indicator dye.…”

Section: Methodsmentioning

confidence: 99%

ATP synthase: activating versus catalytic proton transfer

Groth

Junge

1995

FEBS Letters

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ATP synthase (F-ATPase) of chloroplasts, CFoCF~, is both activated and driven by transmembrane protonmotive force. We dichotomized between activating and driving proton transfer by specific inhibitors, tentoxin and venturicidin. Thylakoids membranes were submitted to voltage steps (by flashing light) superimposed to a steady pH-difference. Transient proton intake, transfer and release by CFoCF~ was monitored by spectroscopic probes. Both activities, activation and catalysis, required all three partial reactions of the proton, however, activating proton transfer rose first (monophasically, z!12 ~ 15 ms) followed by another phase of equal magnitude with a time lag of about 15 ms. Both types of consecutive proton transfer reactions contribute free energy for ATP synthesis.

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“…Abbreviations: DCMU, 3 (3,ldimethylurea;DBMIB,2, iments with dark-adapted chloroplasts the chloroplasts were not exposed to light (while thawing and preparing the suspension) until the measuring light was gated open, typically 10 ms before the actual sampling and averaging (to allow for relaxation of the photomultiplier in response to light gating). The total measuring light energy/sample was 3 pJ/cm2, which excited <5% of the reaction centres [8].…”

Section: Methodsmentioning

confidence: 99%

“…Absorption changes of neutral red which indicate pH-changes inside thylakoids were recorded as in [3]. Signals at 548 nm were recorded in the presence and in the absence of this dye.…”

Section: Methodsmentioning

confidence: 99%

“…The subtraction of the latter from the former ones yielded a response solely to pH-changes inside (note the strong buffering of the external phase by bovine serum albumin). This response is practically artefact free, quantitative and kinetically highly resolving [3]. Fig.1 shows the absorption changes of neutral red under excitation of chloroplasts which were totally dark adapted (according to [S]) with a series of 4 short laser flashes.…”

Section: Methodsmentioning

confidence: 99%

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A cyclic protolytic reaction around photosystem II at the inside of the thylakoid membrane in DCMU‐poisoned chloroplasts

1981

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The buffering capacity of the internal phase of thylakoids and the magnitude of the pH changes inside under flashing light

Cited by 161 publications

References 27 publications

Mobile hydrogen carbonate acts as proton acceptor in photosynthetic water oxidation

Mobile hydrogen carbonate acts as proton acceptor in photosynthetic water oxidation

ATP synthase: activating versus catalytic proton transfer

A cyclic protolytic reaction around photosystem II at the inside of the thylakoid membrane in DCMU‐poisoned chloroplasts

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