Permeability and gating of lens gap junction channels incorporated into liposomes.
The lens junction protein (MIP26), and its trypsin cleavage product (MIP21), isolated from calf fiber cells, are incorporated into liposomes and the permeability and gating of the resulting channels are studied spectrophotometrically by an osmotic swelling assay. Liposomes incorporated with either protein and loaded with Dextran T-10 swell when placed in isotonic or hypertonic KCl, sucrose or polyethyleneglycol (PEG), indicating the presence of channels permeable to molecules as large as MW 1500. In the absence of calmodulin (CaM), the permeability of either MIP26 or MIP21 channels is not altered by Ca++. On the contrary, MIP26-CaM channels reversibly close in the presence of Ca++ (10(-5)M). Preliminary experiments show channel closure with lowered pH (5.5) as well. While MIP26-CaM channels close to all the permeants tested, MIP21-CaM channels close only partially with Ca++, becoming impermeable to large probes (PEG) while remaining permeable to sucrose and KCl. This indicates that the trypsin-cleaved C-terminal arm of MIP26 is the channel gate. Evidence from spectrophotofluorometry and circular dichroism spectroscopy indicates that activated CaM changes the conformation of isolated MIP26, suggesting that channel occlusion could result from a change in protein configuration.
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D000818:Animals; D020346:Aquaporins; D002417:Cattle; D002942:Circular Dichroism; D003459:Crystallins / Q000378:metabolism; D005136:Eye Proteins / Q000378:metabolism; D005614:Freeze Fracturing; D066298:In Vitro Techniques; D007365:Intercellular Junctions / Q000378:metabolism* / Q000648:ultrastructure; D007473:Ion Channels / Q000378:metabolism* / Q000502:physiology; D007908:Lens, Crystalline / Q000378:metabolism*; D008081:Liposomes / Q000378:metabolism*; D008562:Membrane Glycoproteins; D008854:Microscopy, Electron; D010446:Peptide Fragments / Q000378:metabolism; D010539:Permeability; D013050:Spectrometry, Fluorescence; D013053:Spectrophotometry
D020346:Aquaporins; D003459:Crystallins; D005136:Eye Proteins; D007473:Ion Channels; D008081:Liposomes; D008562:Membrane Glycoproteins; D010446:Peptide Fragments; C030328:aquaporin 0; C046263:lens intrinsic protein MIP 21