caa a22 4500 397520956 CHVBK 20180308164632.0 cr unu---uuuuu 161202e199504 xx s 000 0 eng 10.1093/jxb/46.4.361 doi (NATIONALLICENCE)oxford-10.1093/jxb/46.4.361 White Philip J. Department of Cell Physiology, Horticulture Research International, East Mailing, West Mailing, Kent MEW 6BJ, UK and Department of Botany, University of Cambridge, Downing Street, Cambridge CB2 3EA, UK Separation of K+-and Cl −-selective ion channels from rye roots on a continuous sucrose density gradient [Elektronische Daten] [Philip J. White] Microsomal membranes from rye (Secale cereale L.) roots were separated by isopycnic sucrose density gradient centrifugation. The ion channels present in gradient fractions were assayed by reconstitution into planar 1-palmitoyl-2-oleoyl phosphatidylethanolamine bilayers (PLB) and the distributions of ion channel activities were compared with membrane markerenzyme activities. A number of ion channel activities were observed and could be distinguished on the combined bases of their conductance, selectivity, kinetics and pharmacology. A voltage-dependent maxi (498 pS) cation-channel, a voltage-dependent 199-pS cationchannel, 48-pS and 18-pS K+ channels, and a 148-pS Cl− channel (all unitary conductances determined in asymmetrical cis trans 325:100mM KCl) colocalized with the plasma membrane marker-enzyme, vanadatesensitive ATPase. A weakly K +-selective (108 pS) channel, a 1249-pS cation-channel and a 98-pS K + channel colocalized with the tonoplast markerenzyme, nitrate-sensitive ATPase. A 706-pS K+ channel colocalized with the expected distribution of intact plastids and a 38-pS Cl− channel colocalized with either plastid or ER membranes. The membrane location of several other channels including a hypervoltage-sensitive maxi (497 pS) cation-channel, a 270-pS K+ channel, an 8-pS K+ channel and a 4-pS K+ channel was equivocal, but they were tentatively assigned to the Golgi. Thus, the plasma membrane and tonoplast origin of ion channels previously characterized following the incorporation of plasma membrane prepared by aqueous-polymer two-phase partitioning or tonoplast derived from isolated vacuoles into PLB was confirmed and the ion channel complement of previously unassayed membranes was defined. This demonstrates the usefulness of PLB in identifying and characterizing ion channels from plant cell membranes, in particular, those of membranes which are inaccessible to patch-clamp electrodes. © Oxford University Press Articles nationallicence Chloride (Cl−) channel nationallicence potassium (K+) channel nationallicence planar lipid bilayer nationallicence root nationallicence rye nationallicence Secale cerealeL nationallicence Journal of Experimental Botany Oxford University Press 46/4(1995-04), 361-376 0022-0957 46:4<361 1995 46 exbotj https://doi.org/10.1093/jxb/46.4.361 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1093/jxb/46.4.361 text/html Onlinezugriff via DOI NATIONALLICENCE 100 1- White Philip J. Department of Cell Physiology, Horticulture Research International, East Mailing, West Mailing, Kent MEW 6BJ, UK and Department of Botany, University of Cambridge, Downing Street, Cambridge CB2 3EA, UK NATIONALLICENCE 773 0- Journal of Experimental Botany Oxford University Press 46/4(1995-04), 361-376 0022-0957 46:4<361 1995 46 exbotj Metadata rights reserved CC BY-NC-4.0 http://creativecommons.org/licenses/by-nc/4.0 nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-oxford