Molecular mass, stoichiometry, and assembly of 20 S particles Journal Article

  


Authors: Wimmer, C.; Hohl, T. M.; Hughes, C. A.; Müller, S. A.; Söllner, T. H.; Engel, A.; Rothman, J. E.
Article Title: Molecular mass, stoichiometry, and assembly of 20 S particles
Abstract: N-Ethylmaleimide-sensitive factor (NSF), soluble NSF attachment proteins (SNAPs), and SNAP receptor (neuronal SNARE) complexes form 20 S particles with a mass of 788 ± 122 kDa as judged by scanning transmission electron microscopy. A single NSF hexamer and three αSNAP monomers reside within a 20 S particle as determined by quantitative amino acid analysis. In order to study the binding of αSNAP and NSF in solution, to define their binding domains, and to specify the role of oligomerization in their interaction, we fused domains of αSNAP and NSF to oligomerization modules derived from thrombospondin-1, a trimer, and cartilage oligomeric matrix protein, a pentamer, respectively. Binding studies with these fusion proteins reproduced the interaction of αSNAP and NSF N domains in the absence of the hexamerization domain of NSF (D2). Trimeric αSNAP (or its C-terminal half) is sufficient to recruit NSF even in the absence of SNARE complexes. Furthermore, pentameric NSF N domains are able to bind αSNAP in complex with SNAREs, whereas monomeric N domains do not. Our results demonstrate that the oligomerization of both NSF N domains and αSNAP provides a critical driving force for their interaction and the assembly of 20 S particles.
Keywords: carrier protein; unclassified drug; molecular genetics; proteins; animal; electron microscopy; metabolism; animals; microscopy, electron; nerve tissue proteins; protein binding; membrane proteins; protein interaction; scleroprotein; chemistry; amino acid sequence; molecular sequence data; hybrid protein; recombinant fusion proteins; carrier proteins; membrane protein; solutions; amino acids; thrombospondin 1; biochemistry; isolation and purification; nerve protein; molecular weight; glycoproteins; glycoprotein; oligomerization; solution and solubility; macromolecule; macromolecular substances; stoichiometry; amino acid analysis; scanning transmission electron microscopy; vesicular transport proteins; vesicular transport protein; scanning electron microscopy; cartilage; glutaraldehyde; snare protein; oligomers; microscopy, electron, scanning; extracellular matrix proteins; snare proteins; molecular mass; priority journal; article; n-ethylmaleimide-sensitive proteins; soluble n-ethylmaleimide-sensitive factor attachment proteins; n ethylmaleimide; cartilage oligomeric matrix protein; n ethylmaleimide sensitive factor; soluble n ethylmaleimide sensitive factor attachment protein; cartilage matrix protein; glutaral
Journal Title: Journal of Biological Chemistry
Volume: 276
Issue: 31
ISSN: 0021-9258
Publisher: American Society for Biochemistry and Molecular Biology  
Date Published: 2001-08-03
Start Page: 29091
End Page: 29097
Language: English
DOI: 10.1074/jbc.M011292200
PUBMED: 11395481
PROVIDER: scopus
DOI/URL:

http://www.scopus.com/inward/record.url?eid=2-s2.0-0035800765&partnerID=40&md5=072ecb4d3381a33616d65c9cf9b13bb8
Notes: Export Date: 21 May 2015 -- Source: Scopus
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MSK Authors
  1. Thomas H Sollner
    65 Sollner
  2. James E Rothman
    120 Rothman
  3. Christian Georg Wimmer
    5 Wimmer
  4. Christine A Hughes
    4 Hughes
  5. Tobias Martin Hohl
    105 Hohl
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