The pathway of assembly of intermediate filaments from recombinant alpha-internexin.
The pathway of filament assembly from the neuronal intermediate filament alpha-intermexin was investigated. Optimal assembly occurred in solutions of pH 6.5 to 7 and moderate ionic strength at 37 degrees C. Short filaments formed upon dialysis at 24 degrees C, which elongated further when incubated at 37 degrees C. Soluble forms of alpha-internexin were characterized by analytical ultracentrifugation and electron microscopy. In 10 mM Tris, pH 8, conditions that favor formation of tetramers and other small oligomers for other intermediate filament proteins, alpha-internexin formed 10.5 S particles, apparently unit-length half-filaments in the form of rods 10.6 nm in diameter and 68 nm long. Dialysis vs the same buffer with added 10 mM NaCl yielded 16 S rods, probably unit-length filaments, of the same length but 13.0 nm in diameter. At 50 mM NaCl, rods about 13 nm in diameter and heterogeneous in length were observed in electron micrographs, apparently formed from longitudinal annealing of unit-length rods. The results favor a model of assembly in which coiled coil dimers aggregate laterally to form first "unit-length half-filaments" (Herrmann, H., and Aebi, U. (1998) Curr. Opin. Struct. Biol. 8, 177-185) and then "unit-length filaments," which subsequently elongate by annealing.
Abumuhor
I A
IA
Department of Chemistry and Biochemistry, California State University, Long Beach, California, 90840, USA.
Spencer
P H
PH
Cohlberg
J A
JA
D000595:Amino Acid Sequence; D002352:Carrier Proteins / Q000648:ultrastructure*; D006863:Hydrogen-Ion Concentration; D007381:Intermediate Filament Proteins; D007382:Intermediate Filaments / Q000648:ultrastructure*; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000648:ultrastructure; D009994:Osmolar Concentration; D010316:Particle Size; D011994:Recombinant Proteins / Q000648:ultrastructure; D013696:Temperature; D014461:Ultracentrifugation; D014508:Urea / Q000494:pharmacology
D002352:Carrier Proteins; D007381:Intermediate Filament Proteins; D009419:Nerve Tissue Proteins; D011994:Recombinant Proteins; C046643:alpha-internexin; D014508:Urea