Further studies of the thylakoid membrane surface charges by particle electrophoresis.

1. Above pH 4.3 the outer surface of thylakoid membranes isolated from pea chloroplasts is negatively charged but below this value it carries an excess of positive charge. 2. Previously the excess negative charge has been attributed to the carboxyl groups of glutamic and aspartic acid residues (Nakatani, H.Y., Barber, J. and Forrester, J.A. (1978), Biochim. Biophys. Acta 504, 215-225) and in this paper it is argued from experiments involving treatments with 1,2-cyclohexanedione that the positive charges are partly due to the guanidino group of arginine. 3. The electrophoretic mobility of granal (enriched in chlorophyll b and PS II activity) and stromal (enriched in PS I activity) lamellae isolated by the French Press technique were found to be the same. 4. Treatment of the pea thylakoids with trypsin or pronase, sufficient to inhibit the salt induced chlorophyll fluorescence changes, increased their electrophoretic mobility indicating that additional negative charges had been exposed at the surface. 5. Polylysine treatment also inhibited the salt induced chlorophyll fluorescence changes but unlike trypsin and pronase, decreased the net negative charge on the surface. 6. The isoelectric point defined as the pH which gave zero electrophoretic mobility (about 4.3) was independent of the nature of the cations in the suspending medium (monovalent vs. divalent).