Chloroplast thylakoid proteins associated with sequestered proton-buffering domains. Plastocyanin contributes buffering groups to localized proton domains

Journal Article · Tue Aug 01 04:00:00 EDT 1989 · Journal of Bioenergetics and Biomembranes; (USA)

DOI:https://doi.org/10.1007/BF00762525· OSTI ID:5046652

Allnutt, F C; Atta-Asafo-Adjei, E; Dilley, R A ^[1]

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1. Purdue Univ., West Lafayette, IN (USA)

Thylakoid membrane proteins are organized so as to shield 30-50 nmol H+ (mg Chl)-1 from freely equilibrating with either the external or the lumen aqueous phases. Amine groups provide binding sites for this metastable buffering array and can be quantitatively measured by acetylation using (3H)acetic anhydride. The principle of the assay is that a metastable acidic domain will have relatively less of the reactive neutral form of the amine compared to the amount present after addition of an uncoupler. The extent of the acetylation reaction is strongly influenced by whether the lumen pH comes to complete equilibrium with the external pH prior to adding the acetic anhydride. Determination of the lumen pH by (14C)methylamine distribution after the standard 3 or 5 min equilibration in pH 8.6 buffer indicated that the lumen may have been 0.2 to 0.3 pH more acidic than the external phase. This effect was taken into account by determining the pH dependence, in the pH 8.2-8.6 range, of acetylation of the membrane proteins studied, and the labeling data were conservatively corrected for this possible contribution. Experiments were carried out to identify the thylakoid proteins that contribute such metastable domain amine groups, using the above conservative correction. Surprisingly, plastocyanin contributes buried amine groups, but cytochrome f did not give evidence for such a contribution, if the conservative correction in the labeling was applied. If the correction was too conservative, cytochrome f may contribute amines to the sequestered domains. The new methodology verified earlier results suggesting that three Tris-releasable photosystem II-associated proteins also contribute significantly to the sequestered amine-buffering array.

• OSTI ID: 5046652
• Journal Information: Journal of Bioenergetics and Biomembranes; (USA), Journal Name: Journal of Bioenergetics and Biomembranes; (USA) Vol. 21:4; ISSN 0145-479X; ISSN JBBID
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ACETYLATION
ACYLATION
ANHYDRIDES
BARYONS
BIOCHEMICAL REACTION KINETICS
BUFFERS
CARBON 14 COMPOUNDS
CELL CONSTITUENTS
CHEMICAL REACTIONS
CHLOROPLASTS
CYTOCHROMES
ELEMENTARY PARTICLES
FERMIONS
HADRONS
HYDROGEN COMPOUNDS
ISOTOPE APPLICATIONS
KINETICS
LABELLED COMPOUNDS
MEMBRANE PROTEINS
NUCLEONS
ORGANIC COMPOUNDS
PHOTOSYNTHETIC REACTION CENTERS
PIGMENTS
PROTEINS
PROTONS
RADIOASSAY
REACTION KINETICS
RECEPTORS
TRACER TECHNIQUES
TRITIUM COMPOUNDS