Immunoelectron microscopic evidence for the extended conformation of light chains in clathrin trimers.
Clathrin is a major component of the basket-like network of hexagons and pentagons that forms the "coat" on the cytoplasmic face of the plasma membrane and the trans Golgi network during the invagination of coated pits. Soluble clathrin is a three-legged structure (triskelion) comprising three identical heavy chains and three different light chains located toward the center of the triskelion on the proximal segment of the leg. All mammalian light chains contain a central domain of 10 heptad repeats, which is necessary for the interaction with heavy chain. Because the repeats are characteristic of alpha helical coiled coils, we proposed that the central domain had an extended conformation (Kirchhausen, T., Scarmato, P., and Harrison, S. C. et al. (1987) Science 236, 320-324). However, an alternative model has recently been proposed (Nathke, I. S., Heuser, J., Lupas, A., Stock, J., Turck, C. W., and Brodsky, F. M. (1992) Cell 68, 899-910). Here, we use single-molecule electron microscopy of clathrin decorated with monoclonal antibodies directed against different epitopes on light chains to show that the light chain central domain has an extended conformation and reaches along most of the proximal segment of the heavy chain leg.
Kirchhausen
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Department of Anatomy and Cellular Biology, Harvard Medical School, Boston, Massachusetts 02115.
Toyoda
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D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001666:Binding Sites, Antibody; D001921:Brain / Q000378:metabolism; D002417:Cattle; D002645:Chickens; D002966:Clathrin / Q000737:chemistry / Q000235:genetics / Q000648:ultrastructure*; D003094:Collagen / Q000737:chemistry; D000939:Epitopes / Q000378:metabolism; D008099:Liver / Q000378:metabolism; D046911:Macromolecular Substances; D016253:Microscopy, Immunoelectron / Q000379:methods; D008961:Models, Structural; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009838:Oligodeoxyribonucleotides; D011487:Protein Conformation; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000648:ultrastructure
D002966:Clathrin; D000939:Epitopes; D046911:Macromolecular Substances; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D003094:Collagen