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This review explains the potential and the problems, and gives a brief …
Biology Articles » Methods & Techniques » FRAP analysis of photosynthetic membranes » Photosynthetic membranes as model systems for FRAP
One great advantage of photosynthetic membranes is that some of the protein complexes are naturally fluorescent (Table 1), so their diffusion can be observed without any necessity for specific fluorophore binding or GFP gene fusions. In other respects, the majority of thylakoid membranes are far from ideal for FRAP measurements. The membrane systems are enclosed in bacteria or chloroplasts whose overall dimensions are usually rather small. Furthermore, most green plant thylakoid membranes have an intricate, convoluted structure, and extensive lateral heterogeneity on small scales (Mustardy and Garab, 2003). Thus they do not meet the basic requirements for quantitative FRAP, that the membrane geometry is predictable and the membrane environment is uniform over the area of the measurement. While it will not be possible to use FRAP to obtain accurate diffusion coefficients for components of typical green plant thylakoids, it will be possible to see if particular components are mobile or not, and to get a rough idea of time-scales.
The author does not know of any green plant whose thylakoid membrane conformation is as amenable to FRAP as that of Synechococcus. However, some green algae have the advantages of large chloroplast size and low levels of membrane stacking and lateral heterogeneity. Mullineaux and Sarcina (2002) have started a study of Chlamydomonas reinhardtii. It was found that a proportion of the fluorescent chlorophyll in the membrane is mobile. Studies in a range of mutant backgrounds suggest that Photosystem II is immobile, as in cyanobacteria, but a part of the LHCII pool is mobile (M Sarcina, CW Mullineaux, unpublished results). Chloroplast division mutants (Osteryoung and Pyke, 1998) may, potentially, be useful systems for thylakoid FRAP in higher plants.
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