Cyclodextrins: a new tool for the controlled lipid depletion of thylakoid membranes

Abstract

Cyclodextrins (CDs) have been used in a controlled lipid depletion of thylakoid membranes avoiding the use of either detergents or lipolytic enzymes. Spinach thylakoid membranes were first treated with different CDs under various conditions. After removal of the CDs by washing, the amounts of mono- and digalactosyldiacylglycerol (MGDG and DGDG), sulfoquinovosyldiacylglycerol (SQDG) and phosphatidylglycerol (PG), protein, pigment and plastoquinone remaining in the membranes were determined. The main results, obtained with α-CD and heptakis-(2,6-di- O-methyl)- β-CD (DM-β-CD), were as follows. (1) Acyl lipids were removed from thylakoid membranes by both CDs (DM-β-CD being more efficient than α-CD); the extent of removal depended on both CD and chlorophyll concentrations. (2) α-CD presented a higher selectivity towards lipid classes than did DM-β-CD, but in both cases the removal order was SQDG > PG > MGDG > DGDG. (3) α-CD showed a preference for those lipids containing saturated 16-carbon acyl chains whereas DM-β-CD was essentially insensitive to the fatty acid composition of the lipids. (4) The protein, chlorophyll and carotenoid contents of thylakoids were not affected by CD treatments. (5) Plastoquinones were removable but in small amounts only and with a low efficiency (DM-β-CD > α-CD). (6) For all lipid classes, the extent of lipid removal was higher at 0° than at 20°C. (7) The presence of MgCl 2 reduced the removal of PG and SQDG but did not affect galactolipid depletion levels. (8) Stable lipid depletion levels in thylakoid membranes were reached after 5–10 min of CD treatment at 0°C. (9) Of the four CDs tested, only three (α-CD, β-CD, and DM-β-CD) promoted lipid depletion whereas one (hydroxypropyl-β-CD) failed completely to do so. It is concluded that CD-mediated lipid removal provides a valuable and versatile tool to achieve controlled and specific lipid depletions in biological membranes. A few examples of the consequences of a CD-induced lipid depletion on fluorescence and electron transport properties of thylakoids are given to show the usefulness of CDs in the investigation of structure-function relationships in photosynthetic membranes.