Multiple distinct coiled-coils are involved in dynamin self-assembly
Department of Cell Biology
Animals; COS Cells; Centrifugation, Density Gradient; Circular Dichroism; Dynamin I; Dynamins; Endocytosis; GTP Phosphohydrolases; Microtubules; Mutagenesis, Site-Directed; Phenotype; Protein Binding; Protein Conformation; Protein Denaturation; *Protein Folding; Protein Structure, Secondary; Structure-Activity Relationship; Ultracentrifugation; Yeasts
Life Sciences | Medicine and Health Sciences
Dynamin, a 100-kDa GTPase, has been implicated to be involved in synaptic vesicle recycling, receptor-mediated endocytosis, and other membrane sorting processes. Dynamin self-assembles into helical collars around the necks of coated pits and other membrane invaginations and mediates membrane scission. In vitro, dynamin has been reported to exist as dimers, tetramers, ring-shaped oligomers, and helical polymers. In this study we sought to define self-assembly regions in dynamin. Deletion of two closely spaced sequences near the dynamin-1 C terminus abolished self-association as assayed by co-immunoprecipitation and the yeast interaction trap, and reduced the sedimentation coefficient from 7.5 to 4.5 S. Circular dichroism spectroscopy and equilibrium ultracentrifugation of synthetic peptides revealed coiled-coil formation within the C-terminal assembly domain and at a third, centrally located site. Two of the peptides formed tetramers, supporting a role for each in the monomer-tetramer transition and providing novel insight into the organization of the tetramer. Partial deletions of the C-terminal assembly domain reversed the dominant inhibition of endocytosis by dynamin-1 GTPase mutants. Self-association was also observed between different dynamin isoforms. Taken altogether, our results reveal two distinct coiled-coil-containing assembly domains that can recognize other dynamin isoforms and mediate endocytic inhibition. In addition, our data strongly suggests a parallel model for dynamin subunit self-association.
DOI of Published Version
J Biol Chem. 1999 Apr 9;274(15):10277-86.
The Journal of biological chemistry
Okamoto, Patricia M.; Tripet, Brian P.; Litowski, Jennifer R.; Hodges, Robert S.; and Vallee, Richard B., "Multiple distinct coiled-coils are involved in dynamin self-assembly" (1999). Open Access Articles. 768.