Summary
Isolated rat liver gap junctions were split by two methods. In the first method, isolated gap junctions were stabilized by cross-linking their cytoplasmic surfaces with glutaraldehyde under conditions that prevented the entry of glutaraldehyde into the “gap” region. The “stabilized” junctions were then split in the junctional gap with SDS. In the second procedure, unfixed gap junctions were split by incubation in ureacontaining solutions. Junctional splitting was monitored by electron microscopy of thin sectioned and freeze fractured membrane pellets. Sidedness of the split junctional membranes was defined by labeling their cytoplasmic surfaces with glutaraldehyde-activated ferritin before splitting with urea. Gap junctional splitting did not result in any loss of protein components as determined by SDS-gel electrophoresis. The glutaraldehyde cross-linking procedure was also used to determine the effects of various detergents on the protein-protein interactions in the “gap” region. Of the detergents tested, only SDS caused junctional splitting.
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Manjunath, C.K., Goings, G.E. & Page, E. Detergent sensitivity and splitting of isolated liver gap junctions. J. Membrain Biol. 78, 147–155 (1984). https://doi.org/10.1007/BF01869201
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DOI: https://doi.org/10.1007/BF01869201