Proteolytic processing of ovalbumin and β-galactosidase by the proteasome to yield antigenic peptides

Lawrence R. Dick, Carla Aldrich, Stephen C. Jameson, Carolyn R. Moomaw, Bikash C. Pramanik, C. Kuyler Doyle, George N. DeMartino, Michael J. Bevan, James M. Forman, Clive A. Slaughter

Research output: Contribution to journalArticle

168 Citations (Scopus)

Abstract

The identification of genes in the class II region of the MHC that are homologous to genes encoding subunits of the proteasome has led to intense interest in the possible role of this enzyme in the proteolytic processing of polypeptide Ags. We have tested the ability of the 20S proteasome to produce peptides that can be presented by class I molecules as targets for killing by OVA-specific and β-galactosidase-specific CTL clones. Samples of intact OVA and β-galactosidase were subjected to digestion in vitro by 20S proteasome purified from bovine red cells and the resulting peptide mixtures were fractionated by reverse-phase HPLC. The fractions were tested for their ability to sensitize appropriate mouse target cells for lysis by specific CTL clones. In both cases, components that under all chromatographic conditions eluted with retention times indistinguishable from synthetic peptides representing known epitopes of the naturally processed proteins were found to be able to sensitize the target cells. Moreover, in the case of OVA, the presence of the expected target peptides was demonstrated directly by amino acid sequence and mass spectrometric analysis. The results demonstrate that the pure 20S proteasome is capable of generating antigenic peptides from two proteins for presentation by class I molecules without the participation of additional components of the protein degradation system. This finding is consistent with the hypothesis of proteasome involvement in Ag processing in vivo.

Original languageEnglish (US)
Pages (from-to)3884-3894
Number of pages11
JournalJournal of Immunology
Volume152
Issue number8
StatePublished - Apr 15 1994
Externally publishedYes

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Galactosidases
Ovalbumin
Proteasome Endopeptidase Complex
Peptides
Clone Cells
MHC Class II Genes
Proteolysis
Epitopes
Digestion
Amino Acid Sequence
Proteins
Peptide Hydrolases
High Pressure Liquid Chromatography
Genes

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology

Cite this

Dick, L. R., Aldrich, C., Jameson, S. C., Moomaw, C. R., Pramanik, B. C., Doyle, C. K., ... Slaughter, C. A. (1994). Proteolytic processing of ovalbumin and β-galactosidase by the proteasome to yield antigenic peptides. Journal of Immunology, 152(8), 3884-3894.

Proteolytic processing of ovalbumin and β-galactosidase by the proteasome to yield antigenic peptides. / Dick, Lawrence R.; Aldrich, Carla; Jameson, Stephen C.; Moomaw, Carolyn R.; Pramanik, Bikash C.; Doyle, C. Kuyler; DeMartino, George N.; Bevan, Michael J.; Forman, James M.; Slaughter, Clive A.

In: Journal of Immunology, Vol. 152, No. 8, 15.04.1994, p. 3884-3894.

Research output: Contribution to journalArticle

Dick, LR, Aldrich, C, Jameson, SC, Moomaw, CR, Pramanik, BC, Doyle, CK, DeMartino, GN, Bevan, MJ, Forman, JM & Slaughter, CA 1994, 'Proteolytic processing of ovalbumin and β-galactosidase by the proteasome to yield antigenic peptides', Journal of Immunology, vol. 152, no. 8, pp. 3884-3894.
Dick LR, Aldrich C, Jameson SC, Moomaw CR, Pramanik BC, Doyle CK et al. Proteolytic processing of ovalbumin and β-galactosidase by the proteasome to yield antigenic peptides. Journal of Immunology. 1994 Apr 15;152(8):3884-3894.
Dick, Lawrence R. ; Aldrich, Carla ; Jameson, Stephen C. ; Moomaw, Carolyn R. ; Pramanik, Bikash C. ; Doyle, C. Kuyler ; DeMartino, George N. ; Bevan, Michael J. ; Forman, James M. ; Slaughter, Clive A. / Proteolytic processing of ovalbumin and β-galactosidase by the proteasome to yield antigenic peptides. In: Journal of Immunology. 1994 ; Vol. 152, No. 8. pp. 3884-3894.
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