Per-Åke Albertsson

A quantitative model of the domain structure of the photosynthetic membrane

A model is presented that gives a quantitative picture of the distribution of the photosynthetic components in the photosynthetic membrane of higher plants. A salient feature of the model is that most of the pigments are located in the grana where photosystem I and II carry out linear electron transport, whereas the stroma lamellae, which harbour <20% of the pigments, carry out photosystem-I-mediated cyclic electron transport. This arrangement derives from the observation that more pigments are associated with photosystem I, which therefore captures more quanta than photosystem II. The excess pigments associated with photosystem I are thought to be located in the stroma lamellae.

Partition of cell particles and macromolecules in polymer two-phase systems.

Publisher Summary This chapter discusses the phenomenon of phase separation in polymer–polymer–water mixtures and describes the general behavior of particles and macromolecules in polymer two phase systems. A general problem with all partition methods is that they are highly empirical. Many factors govern the partition behavior. Thus, the physical and chemical nature of the phase polymers, the ionic composition of the phases, the nature of the partitioned substance all influence partition in a complicated manner. It is difficult to predict in detail the behavior of different substances. Time and labor must, therefore, be devoted to find out, more or less by trial and error, suitable solvent systems that can accomplish the desired separations. However, this general problem can be partly overcome by collecting a large number of data on representative substances in a few selected phase systems. If these data can be systematized, then they can be of great help in devising a successful separation experiment on an unknown mixture.

Partition of proteins in aqueous polymer two-phase systems and the effect of molecular weight of the polymer

The partition of substances in aqueous polymer two-phase systems is influenced by the molecular weight of the phase-forming polymers. We investigate how the effect of the molecular weight of the polymers depends on the molecular weight of the partitioned protein. We show that the magnitude of change of the partition is very small for proteins of molecular weights around 10,000, but increases almost linearly up to molecular weights of 250,000.

Interfacial tension of dextran—polyethylene glycol—water two—phase systems

Abstract The interfacial tension between the two aqueous phases of dextran—polyethylene glycol—water systems has been determined by the rotating drop method. Values in the range 5 × 10−4−10−1 dyn/cm have been obtained depending on the polymer concentration. A linear relationship between the logarithm of the interfacial tension and the length of the tie line of the phase diagram was found.

The structure and function of the chloroplast photosynthetic membrane - a model for the domain organization

Recent work on the domain organization of the thylakoid is reviewed and a model for the thylakoid of higher plants is presented. According to this model the thylakoid membrane is divided into three main domains: the stroma lamellae, the grana margins and the grana core (partitions). These have different biochemical compositions and have specialized functions. Linear electron transport occurs in the grana while cyclic electron transport is restricted to the stroma lamellae. This model is based on the following results and considerations. (1) There is no good candidate for a long-range mobile redox carrier between PS II in the grana and PS I in the stroma lamellae. The lateral diffusion of plastoquinone and plastocyanin is severely restricted by macromolecular crowding in the membrane and the lumen respectively. (2) There is an excess of 14±18% chlorophyll associated with PS I over that of PS II. This excess is assumed to be localized in the stroma lamellae where PS I drives cyclic electron transport. (3) For several plant species, the stroma lamellae account for 20±3% of the thylakoid membrane and the grana (including the appressed regions, margins and end membranes) for the remaining 80%. The amount of stroma lamellae (20%) corresponds to the excess (14–18%) of chlorophyll associated with PS I. (4) The model predicts a quantum requirement of about 10 quanta per oxygen molecule evolved, which is in good agreement with experimentally observed values. (5) There are at least two pools of each of the following components: PS I, PS II, cytochrome bf complex, plastocyanin, ATP synthase and plastoquinone. One pool is in the grana and the other in the stroma compartments. So far, it has been demonstrated that the PS I, PS II and cytochrome bf complexes each differ in their respective pools.

Isolation of Photosystem II enriched membrane vesicles from spinach chloroplasts by phase partition

Partition in an aqueous Dextran-polyethylene glycol two-phase system has been used for the separation of chloroplast membrane vesicles obtained by press treatment of a grana-enriched fraction after unstacking in a low salt buffer. The fractions obtained were analysed with respect to chlorophyll, photochemical activities and ultrastructural charasteristics. The results reveal that the material partitioning to the Dextran-rich bottom phase consisted of large membrane vesicles possessing mainly Photosystem II properties with very low contribution from Photosystem I. Measurements of the H2O to phenyl-p-benzoquinone and ascorbate-Cl2Ind to NADP+ electron transport rates indicate a ratio of around six between Photosystem II and I. The total fractionation procedure could be completed within 2-3 h with high recovery of both the Photosystem II water-splitting activity and the Photosystem I reduction of NADP+. These data demonstrate that press treatment of low-salt destabilized grana membranes yields a population of highly Photosystem-II enriched membrane vesicles which can be discriminated by the phase system. We suggest that such membrane vesicles originate from large regions in the native grana membrane which contain virtually only Photosystem II.

Dimeric and monomeric organization of photosystem II - Distribution of five distinct complexes in the different domains of the thylakoid membrane

The supramolecular organization of photosystem II (PSII) was characterized in distinct domains of the thylakoid membrane, the grana core, the grana margins, the stroma lamellae, and the so-called Y100 fraction. PSII supercomplexes, PSII core dimers, PSII core monomers, PSII core monomers lacking the CP43 subunit, and PSII reaction centers were resolved and quantified by blue native PAGE, SDS-PAGE for the second dimension, and immunoanalysis of the D1 protein. Dimeric PSII (PSII supercomplexes and PSII core dimers) dominate in the core part of the thylakoid granum, whereas the monomeric PSII prevails in the stroma lamellae. Considerable amounts of PSII monomers lacking the CP43 protein and PSII reaction centers (D1-D2-cytochrome b559 complex) were found in the stroma lamellae. Our quantitative picture of the supramolecular composition of PSII, which is totally different between different domains of the thylakoid membrane, is discussed with respect to the function of PSII in each fraction. Steady state electron transfer, flash-induced fluorescence decay, and EPR analysis revealed that nearly all of the dimeric forms represent oxygen-evolving PSII centers. PSII core monomers were heterogeneous, and a large fraction did not evolve oxygen. PSII monomers without the CP43 protein and PSII reaction centers showed no oxygen-evolving activity.

Separation and characterization of stroma and grana membranes: evidence for heterogeneity in antenna size of both photosystem I and photosystem II

A rapid procedure to fractionate the thylakoid membrane into two well-separated vesicle populations, one originating from the grana and the other from the stroma-membrane region, has been developed. This was achieved by sonication of thylakoids present in an aqueous two-phase system followed by partitioning either by countercurrent distribution or by a batch procedure in three steps. The membrane populations were analysed according to their composition and photochemical activities. The grana membranes comprise, on chlorophyll basis, about 60% of the thylakoid material and are enriched in PS II, but also contain some PS I, while the stroma membranes comprise about 40% and are enriched in PS I, but also contain some PS II. Cytochrome f was slightly enriched in the grana-derived vesicle fraction. The properties of both PS I and PS II differ between the two populations. The PS I of the grana fraction (PS Iα) reached half-saturation at about half the light intensity of the PS I in the stroma-membrane fraction (PS Iβ). The rate of P-700 photooxidation under low light illumination was higher for PS Iα than for PS Iβ (30% larger rate constant), showing that PS Iα has a larger antenna. The PS II of the grana fraction (PS IIα) reached half-saturation at half the light intensity compared to the PS II of the stroma-membrane fraction (PS IIβ). The results show that the grana-derived membranes contain PS Iα and PS IIα which have larger functional antenna sizes than the corresponding PS Iβ and PS IIβ of the stroma membranes. The results suggest that the photosystems of the grana are designed to allow effective electron transport both at low and high light intensities, while the stroma-membrane photosystems mainly work at high light intensities as a supplement to the grana systems.

Partition between polymer phases

Abstract Separation of biopolymers and cell particles can be accomplished by partition between two or more immiscible, liquid, aqueous, polymer-containing phases. The phase systems are obtained by mixing water solutions of different polymers, such as dextran and polyethylene glycol. This review describes the various factors which determine the partition. By binding a biospecific ligand covalently to one of the phase polymers, biospecific affinity partition is obtained. Applications on the separation of chloroplasts and membrane vesicles are described. Partition can also be used to detect and study interaction between macromolecules. Formulae for the calculation of the dissociation constant for a 1:1 protein-protein complex are presented.

Separation of subchloroplast membrane particles by counter-current distribution

Counter-current distribution in an aqueous Dextran-polyethylene glycol two-phase system has been used to fractionate membrane fragments obtained by press treatment of Class II chloroplasts. By the counter-current distribution technique membrane particles are separated according to their surface properties such as charge and hydrophobicity. The fractions obtained were analysed with respect to photochemical activities, chlorophyll and P-700 contents. The Photosystem II enrichment after counter-current distribution was better than that obtained by differential centrifugation of the disrupted chloroplasts. However, the best separation of Photosystem I and II enriched particles could be achieved if differential centrifugation was combined with the counter-current distribution technique. Each centrifugal fraction could be further separated into Photosystems I and II enriched fractions since the Photosystem II particles preferred the dextran-rich bottom phase while the Photosystem I particles preferred the polyethylene glycol-rich top phase. By this procedure it was possible, without the use of detergents, to obtain vesicles which were more enriched in Photosystem II as compared to intact grana stacks. The partition behaviour of undisrupted Class II chloroplasts and the Photosystem I centrifugal fraction was the same. This similarity indicated that the membrane which is exposed to the surrounding polymers by the Class II chloroplasts is the Photosystem I rich membrane of the stroma lamellae.