David J. Goodchild

Composition of the photosystems and chloroplast structure in extreme shade plants

Abstract Chloroplasts were isolated from leaves of three species of tropical rainforest plants, Alocasia macrorrhiza, Cordyline rubra and Lomandra longifolia; these species are representative of extreme “shade” plants. It was found that shade plant chloroplasts contained 4–5 times more chlorophyll than spinach chloroplasts. Their chlorophyll a/chlorophyll b ratio was 2.3 compared with 2.8 for spinach. Electron micrographs of leaf sections showed that the shade plant chloroplasts contained very large grana stacks. The total length of partitions relative to the total length of stroma lamellae was much higher in Alocasia than in spinach chloroplasts. Freeze-etching of isolated chloroplasts revealed both the small and large particles found in spinach chloroplasts. Despite their increased chlorophyll content, low chlorophyll a/chlorophyll b ratio, and large grana, the shade plant chloroplasts were fragmented with digitonin to yield small fragments (D-144) highly enriched in Photosystem I, and large fragments (D-10) enriched in Photosystem II. The degree of fragmentation of the shade plant chloroplasts was remarkably similar to that of spinach chloroplasts, except that the subchloroplast fragments from the shade plants had lower chlorophyll a/chlorophyll b ratios than the corresponding fragments from spinach. The D-10 fragments from the shade plants had chlorophyll a/chlorophyll b ratios of 1.78-2.00 and the D-144 fragments ratios of 3.54–4.07. We conclude that Photosystems I and II of the shade plants have lower proportions of chlorophyll a to chlorophyll b than the corresponding photosystems of spinach. The lower chlorophyll a/chlorophyll b ratio of shade plant chloroplasts is not due to a significant increase in the ratio of Photosystem II to Photosystem I in these chloroplasts. The extent of grana formation in higher plant chloroplasts appears to be related to the total chlorophyll content of the chloroplast. Grana formation may simply be an means of achieving a higher density of light-harvesting assemblies and hence a more efficient collection of light quanta.

Immunocytochemical localization of the cytochrome b/f complex of chloroplast thylakoid membranes.

Antibodies directed against purified cytochrome f, isolated from the cytochrome b/f complex of spinach chloroplasts, were used in on-grid immunogold labelling studies of spinach leaf tissue. Our results show unambiguously that cytochrome f, and hence the cytochrome b/f complex, is located in both appressed and non-appressed thylakoid membranes.

Transbilayer organization of the main chlorophyll a/b-protein of photosystem II of thylakoid membranes

To probe the location of the carboxyl‐terminus of the 28 kDa apoprotein of the light‐harvesting chlorophyll a/b‐protein complex of PS II (LHCII), an antibody was generated against a synthetic octapeptide corresponding to the C‐terminal region of LHCII. The high specificity of the octapeptide antiserum was deonstrated by immunoblots and immunogold labelling. The octapeptide antiserum agglutinated destacked thylakoid membranes, but no significant agglutination occurred with inside‐out vesicles suggesting that the COOH‐terminus is located at the outer, stroma‐exposed surface where the NH2‐terminus is also located [(1983) J. Biol. Chem. 258, 9941‐9948]. Our results support a model for LHCII with four transmembrane‐spanning domains.

The lateral displacement of intramembraneous particles in chloroplast membranes as a function of light intensity

not received Freeze-fracture Intramembrane particle distribution Chloroplast membrane Light intensity

Lateral Distribution of the Photosystem I Complex between the Appressed and Non-Appressed Regions of Spinach Thylakoid Membranes: An Immunocytochemical Study

A marked structural and functional heterogeneity occurs in the thylakoid membranes of higher plants and green algal chloroplasts that contain chlorophyll b. Although the concept of lateral heterogeneity in the distribution of thylakoid complexes between the appressed and non-appressed thylakoids is generally accepted (1–4) the evidence is nevertheless indirect. Biochemical fractionation studies (2–4) and ultrastructural freeze-fracture analyses (cf. 5) indicate that most, but not all, PSII complexes and LHCII are located in the non-appressed membranes, while all ATP synthetase (6), most PSI complexes and small amounts of PSII complexes and LHCII are located in the non-appressed membranes. Conflicting results have been obtained for the distribution of the cytochrome b/f complex.

The Granal Margins of Plant Thylakoid Membranes : An Important Nonappressed Domain

The thylakoid membranes of mature higher plant chloroplasts show a remarkable structural and functional differentiation into appressed and nonappressed domains (Fig. 1). The closely appressed membranes of the granal compartments (whose outer surfaces do not have direct contact with the stroma) contain the PSIIα units consisting of core PSII complexes surrounded by heterogenous Chl a/b-proteins of LHCII (1,2). The planar nonappressed domains of the interconnecting stroma thylakoids and the end-grana membranes have direct access to the stroma: they contain PSI complex (1), ATP synthase (3) and few PSIIβ complexes (2) with smaller light-harvesting antennae. Only the Cyt b/f complex is thought to be present in both membrane domains (4). However, a nonappressed domain, the margins of the granal compartments (Fig. 1), has until recently been ignored. We suggest the granal margins form an important functional and structural domain.

A METHOD OF ESTIMATING RADIAL DISTRIBUTION FUNCTION OF PROTEIN PARTICLES IN MEMBRANES FROM FREEZE-FRACTURE ELECTRON MICROGRAPHS

A computational method for estimating the radial distribution function of particles seen in electron micrographs of freeze-fractured membranes is presented. The method overcomes the previous difficulty of measurements of this quantity from freeze-fracture electron micrographs--the systematic error due to the finite size of the sample areas in the freeze-fracture. By multiplying the measurements by weights specific to the shape of the sample area, the method lessens the errors due to the boundaries of the area and allows a synthesis of the average radial distribution function from counts over many small separate areas.