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Counter-current distribution of sonicated inside-out thylakoid vesicles

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Summary

Inside-out thylakoid vesicles were isolated from spinach chloroplasts, and fragmented by sonication. Different fragments were separated by counter-current distribution and analyzed for chlorophyll and P700. The inside-out vesicles had a chlorophyll a/b ratio of 2.2–2.4 (original chloroplasts 2.8–3.0). After further fragmentation of the inside-out vesicles by sonication and separation by countercurrent distribution three populations of vesicles were obtained having chlorophyll a/b ratios of 1.7, 1.9 and 2.5 respectively. The P-700 was depleted in fractions with lower chlorophyll a/b ratio and was nearly absent in the fraction having a chlorophyll a/b ratio of 1.7 (chlorophyll/P700 > 4500 mol/mol). That PSII membrane vesicles, with such a low chlorophyll a/b ratio and lacking PSI, can be prepared by a non-detergent method provides strong support for the notion that PSI and PSII are segregated along the thylakoid membrane.

A plot of P700 per chlorophyll against chlorophyll b/(a+b) fits a straight line connecting the pure PSI membrane (chlorophyll a/b = 6; P700/chlorophyll = 5.6 mmol/mol) with the pure PSII membrane (chlorophyll a/b = 1.7; P700 = 0). These two membranes can be considered as separate phases of a two-dimensional phase system. Models for the thylakoid membrane are discussed.

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Abbreviations

PSI:

Photosystem I

PSII:

Photosystem II

PEG:

Polyethylene Glycol

P700:

Reaction Center of PSI

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Correspondence to Per-Åke Albertsson.

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Albertsson, P., Svensson, P. Counter-current distribution of sonicated inside-out thylakoid vesicles. Mol Cell Biochem 81, 155–163 (1988). https://doi.org/10.1007/BF00219318

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Key words

  • chloroplast
  • thylakoid
  • membrane
  • inside-out vesicles
  • photosystem II
  • photosystem I
  • counter current distribution