Dissociation and Reassembly of Soybean Clathrin

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Abstract
Coated vesicles (CVs), approximately 85 nanomolar in diameter, were obtained from etiolated soybean hypocotyls using discontinuous sucrose gradients. The CVs were treated with 4 molar urea and the vesicle membranes were then removed by centrifugation. When the supernatant subsequently was dialyzed against isolation buffer, sedimentable complexes were obtained. Electron microscopic examination of the pelleted complexes shows many spherical, 65 nanometer baskets consisting of a polygonal lattice free of internal membrane. LDS-PAGE reveals that a 185 kilodalton clathrin heavy chain is enriched in both the basket and CV pellets. However, the overall protein pattern is more complex than that of comparable brain fractions. The results are discussed in terms of the similarities between soybean and brain clathrins and the function of CVs in vivo.