Surface charges on chloroplast membranes as studied by particle electrophoresis

1. Particle microelectrophoresis mobility studies have been conducted with chloroplast thylakoid membranes and with isolated intact chloroplasts.2. The pH dependence of the electrophoretic mobility indicated that at pH values above 4.3 both membrane systems carry a net negative charge.3. Chemical treatment of thylakoids has shown that neither the sugar residues of the galactolipids in the membrane nor the basic groups of the membrane proteins having pK values between 6 and 10 are exposed at the surface.4. However, treatment with 1-ethyl-3(3-dimethylaminopropyl)carbodiimide, together with glycine methyl ester, neutralized the negative charges on the thylakoid membrane surface indicating the involvement of carboxyl groups which, because of their pH sensitivity, are likely to be the carboxyl groups of aspartic and glutamic acid residues.5. The nature of the protein giving rise to the negative surface charges on the thylakoids is not known but is shown not to involve the coupling factor or the light harvesting $\text{chlorophyl}\text{a}\text{chlorophyll}\text{b}\text{pigment · protein complex}$.6. No significant effect of light was observed on the electrophoretic mobility of either thylakoids or intact chloroplasts.7. The striking difference in the ability of divalent and monovalent cations to screen the surface charges was demonstrated and explained in terms of the Gouy-Chapman theory.8. Calculations of the ζ-potentials for thylakoid membranes gave values for the charge density at the plane of shear to be in the region of one electronic charge per 1500–2000 Ų.9. The significance of the results is discussed in terms of cation distribution in chloroplasts and the effect of cations on photosynthetic phenomena.