Abstract
A method is described that uses trypsin digestion combined with collagenase-hyaluronidase which produces a population of gap junction vesicles. The hexagonal lattice of subunits ("connexons") comprising the gapjunctions appears unaltered by various structural criteria and by buoyant density measurements. The gap junction vesciles are closed by either a single or a double profile of nonjunctional "membrane," which presents a smooth, particle-free fracture face. Horseradish peroxidase and cytochrome c studies have revealed that about 20% of the gap junction vesicles are impermeable to proteins 12,000 daltons or larger. The increased purity of the trypsinized junction preparation suggests that one of the disulfide reduction products of the gap- junction principal protein may be a nonjunctional contaminating peptide. The gap junction appears to be composed of a single 18,000- dalton protein, connexin, which may be reduced to a single 9,000-dalton peak. The number of peptides in this reduced peak are still unknown.
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- Benedetti E. L., Emmelot P. Electron microscopic observations on negatively stained plasma membranes isolated from rat liver. J Cell Biol. 1965 Jul;26(1):299–305. doi: 10.1083/jcb.26.1.299. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Benedetti E. L., Emmelot P. Hexagonal array of subunits in tight junctions separated from isolated rat liver plasma membranes. J Cell Biol. 1968 Jul;38(1):15–24. doi: 10.1083/jcb.38.1.15. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Evans W. H., Gurd J. W. Preparation and properties of nexuses and lipid-enriched vesicles from mouse liver plasma membranes. Biochem J. 1972 Jul;128(3):691–700. doi: 10.1042/bj1280691. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Goodenough D. A. Bulk isolation of mouse hepatocyte gap junctions. Characterization of the principal protein, connexin. J Cell Biol. 1974 May;61(2):557–563. doi: 10.1083/jcb.61.2.557. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Goodenough D. A., Gilula N. B. The splitting of hepatocyte gap junctions and zonulae occludentes with hypertonic disaccharides. J Cell Biol. 1974 Jun;61(3):575–590. doi: 10.1083/jcb.61.3.575. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Goodenough D. A., Revel J. P. The permeability of isolated and in situ mouse hepatic gap junctions studied with enzymatic tracers. J Cell Biol. 1971 Jul;50(1):81–91. doi: 10.1083/jcb.50.1.81. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Goodenough D. A., Stoeckenius W. The isolation of mouse hepatocyte gap junctions. Preliminary chemical characterization and x-ray diffraction. J Cell Biol. 1972 Sep;54(3):646–656. doi: 10.1083/jcb.54.3.646. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Graham R. C., Jr, Karnovsky M. J. The early stages of absorption of injected horseradish peroxidase in the proximal tubules of mouse kidney: ultrastructural cytochemistry by a new technique. J Histochem Cytochem. 1966 Apr;14(4):291–302. doi: 10.1177/14.4.291. [DOI] [PubMed] [Google Scholar]
- Karnovsky M. J., Rice D. F. Exogenous cytochrome c as an ultrastructural tracer. J Histochem Cytochem. 1969 Nov;17(11):751–753. doi: 10.1177/17.11.751. [DOI] [PubMed] [Google Scholar]
- McNutt N. S., Weinstein R. S. The ultrastructure of the nexus. A correlated thin-section and freeze-cleave study. J Cell Biol. 1970 Dec;47(3):666–688. doi: 10.1083/jcb.47.3.666. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Pappas G. D., Asada Y., Bennett M. V. Morphological correlates of increased coupling resistance at an electrotonic synapse. J Cell Biol. 1971 Apr;49(1):173–188. doi: 10.1083/jcb.49.1.173. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Payton B. W., Bennett M. V., Pappas G. D. Permeability and structure of junctional membranes at an electrotonic synapse. Science. 1969 Dec 26;166(3913):1641–1643. doi: 10.1126/science.166.3913.1641. [DOI] [PubMed] [Google Scholar]
- Zampighi G., Robertson J. D. Fine structure of the synaptic discs separated from the goldfish medulla oblongata. J Cell Biol. 1973 Jan;56(1):92–105. doi: 10.1083/jcb.56.1.92. [DOI] [PMC free article] [PubMed] [Google Scholar]