Physical characterization of gap junction membrane connexons (hemi-channels) isolated from rat liver

Michael Cascio, University of Pittsburgh School of Medicine
Nalin M. Kumar, Scripps Research Institute
Robert Safarik, Scripps Research Institute
Norton B. Gilula, Scripps Research Institute


Enriched subcellular fractions of double membrane gap junctions (plaques) from rat livers were treated under reducing conditions with high salt and non-ionic detergent concentrations at high pH to obtain a preparation of structural 80-90 Å complexes of oligomers (connexons). The isolated oligomers were chromatographically purified, and subsequently characterized immunologically, morphologically by electron microscopy, hydrodynamically by gel filtration and ultracentrifugation, spectroscopically by circular dichroism, and chemically via cross-linking studies. The physical characteristics of these isolated gap junction complexes were compared to those of native membrane-bound gap junctions in rat liver. These analyses indicate that the isolated complex (connexon) principally contains a hexameric arrangement of gap junction protein to form a single membrane hemi-channel.