Assembly properties of two CNBr fragments of avian desmin that correspond to the headpiece domain and helix 1B.
To study how different domains of the muscle-specific intermediate filament protein, desmin, contribute to its polymerization, two of its CNBr fragments were examined as to their oligomeric structure under assembly conditions. One of these, D88, covers residues 1-88 and represents almost the entire headpiece; the other, D109, covers residues 145-254, and includes the entire Helix 1B and part of linker L12 of the intact molecule. Chemical cross-linking followed by SDS-PAGE, and analytical gel filtration, revealed that in 10 mM Tris-HCl, pH 8.5, conditions that favor tetramerization of intact desmin D88 formed only dimers. D109, on the other hand, formed primarily a dimeric species but low levels of trimeric and tetrameric species were also detectable. These data are consistent with the proposal that, during assembly of intact protein molecules into IF, the headpiece and Helix 1 contribute to dimerization of two polypeptides into a parallel, in-register coiled-coil. However, additional interactions, including headpiece-to-rod binding and hydrophobic interaction along the entire rod domain, are required to stabilize the tetramers and full-size IF.
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Department of Anatomy and Cell Biology, University of Cincinnati College of Medicine, Ohio 45267-0521.
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D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002645:Chickens; D002850:Chromatography, Gel; D003432:Cross-Linking Reagents; D003488:Cyanogen Bromide; D003893:Desmin / Q000302:isolation & purification; D004120:Dimethyl Suberimidate; D004591:Electrophoresis, Polyacrylamide Gel; D005022:Ethyldimethylaminopropyl Carbodiimide; D005895:Gizzard, Avian / Q000032:analysis; D046911:Macromolecular Substances; D008970:Molecular Weight; D010446:Peptide Fragments / Q000302:isolation & purification; D013388:Succinimides
D000596:Amino Acids; D003432:Cross-Linking Reagents; D003893:Desmin; D046911:Macromolecular Substances; D010446:Peptide Fragments; D013388:Succinimides; D004120:Dimethyl Suberimidate; C001426:N-hydroxysuccinimide; D003488:Cyanogen Bromide; D005022:Ethyldimethylaminopropyl Carbodiimide