Lateral Diffusion of Plastocyanin and Plastoquinol in Thylakoid Membranes

Identification of the plastocyanin binding subunit of photosystem I

“…(iii) The excess of 7 positively charged residues is consistent with studies of the surface charge density at the oxidizing site of PSI which provided first evidence that subunit III is positively charged [10]. The positive charges may not only compensate the high negative surface charge density at the electron transfer site to P700 but also provide a suitable protein environment for rapid binding [7] while the bulk of the repulsing negatively charged lipid membrane may facilitate the diffusion of plastocyanin in the thylakoid lumen [12].…”

“…The time of the electron transfer from crosslinked plastocyanin to P700 + of 13/~s ( fig.3C) is the same as that of the complex in vivo [11,12]. This is evidence that cross-linking with EDC conserved the orientation of the complex and that subunit III provides the conformation of PS I necessary for the extremely rapid electron transfer from plastocyanin to P700 +.…”

“…In this study the subunit has been shown to be positively charged which enables the fast electron transfer to the oxidizing site of the PSI reaction center of extremely negative surface charge density. Plastocyanin bound to PSI in situ reduces P700 + with a halftime of 12-14/zs [11,12] which has also been observed in vitro [13]. In a recent study Wynn and Malkin [14] have crosslinked plastocyanin to a PSI subunit of 19 kDa and demonstrated that the carboxyl groups of plastocyanin are necessary for this reaction.…”

“…Electroblotting to nitrocellulose, immuno-gold staining and characterization of the monospecific antibodies raised against plastocyanin has been described [12]. Preparative SDS-PAGE was as described in [17] except that a 2-mm gel was loaded over the full width with isolated PSI containing 350/~g chl.…”

“…The latter gel was used for electroblotting [12] to an Immobilon membrane (Millipore) in 20 mM CAPS buffer, pH 10, and 20070 methanol for 4.5 h at 0.2 A and 4°C. The membrane was stained with 0.001070 (w/v) Coomassie brilliant blue G-250 in methanol/acetic acid (40: 4, v/v).…”