MACROMOLECULAR STRUCTURE

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The structural and compositional heterogeneity of coated vesicles purified from rat liver has been investigated by compiling mass distributions of LCV and of particles derived from them both by extraction with the non-ionic detergent Triton X-100 and by removal of their clathrin coats by manipulation of pH and ionic strength. The masses of individual particles were determined by computer image processing of dark-field scanning transmission electron micrographs of unstained frozen-dried preparations; these data were compiled into statistical distributions and the resulting data were then correlated with biochemical determinations of the global contents of protein, phospholipid, and cholesterol in the respective preparations. We found that about 30% of "LCV" do not, in fact, contain membrane vesicles (greater than 70% in the case of brain coated vesicles), but have proteinaceous cores. The remaining 70% contain vesicles seemingly of two kinds differentiated according to the relative amounts of material contained within their lumens. Vesicle-containing LCV are markedly heterogeneous, with masses varying over a four-fold range (about 50 to 200 megadaltons-MDa) and a two-fold diameter range (about 80 to 160 nm). The average mass of such particles is 80 MDa, of which 46 MDa is contributed by clathrin and other coat proteins, 10-12 MDa by phospholipid and cholesterol, and 20-22 MDa by vesicle proteins. As estimated by SDS-PAGE the latter constitute an extremely diverse collection of proteins that are present in vary samll amounts, except for a relatively prominent doublet of 46kDa. In addition, structural analyses based on various electron microscopical methods reinforced by digital image processing have been performed on a number of other biological systems, including keratin intermediate filaments, the myofilament lattice of vertebrate skeletal muscle, the fimriae of Bordetella pertussis, and the tail-fibers of bacteriophage T7.