The sites in the I-Ak histocompatibility molecule photoaffinity labeled by an immunogenic lysozyme peptide.

Détails

ID Serval
serval:BIB_6FAA1689047A
Type
Article: article d'un périodique ou d'un magazine.
Collection
Publications
Titre
The sites in the I-Ak histocompatibility molecule photoaffinity labeled by an immunogenic lysozyme peptide.
Périodique
The Journal of biological chemistry
Auteur(s)
Luescher I.F., Crimmins D.L., Schwartz B.D., Unanue E.R.
ISSN
0021-9258
Statut éditorial
Publié
Date de publication
1990
Peer-reviewed
Oui
Volume
265
Numéro
19
Pages
11177-11184
Langue
anglais
Notes
Publication types: Journal Article ; Research Support, Non-U.S. Gov't ; Research Support, U.S. Gov't, P.H.S. Publication Status: ppublish
Résumé
The class II histocompatibilty molecule I-Ak was photoaffinity labeled by NH2- and COOH-terminal photoreactive conjugates of an immunogenic hen egg white lysozyme (HEL) peptide. The labeled alpha and beta chains were digested with protease from Staphylococcus aureus strain V-8 (protease V-8) and/or trypsin, and the proteolytic fragments were separated by high performance liquid chromatography (HPLC) (peptide mapping). Reproducible peptide maps containing a major labeled component were obtained from the three conjugates reported here whose photoreactive group was attached via short spacers of limited flexibility. The COOH-terminal conjugate N-acetyl HEL-(49-61)-iodo-4-azidosalicyloyl thioester (compound 1) labeled hydrophilic tryptic digest fragments on both chains of I-Ak. The labeled digest fragments were homogeneous in reverse-phase and anion-exchange HPLC, indicating that the photoaffinity labeling was site-specific. Conversely, the NH2-terminal conjugate iodo-4-azidosalicyloyl HEL-(46-61) (compound 2: IASA-(46-61)) labeled exceptionally hydrophobic sequences on both chains of I-Ak. The labeling was also site-specific because reverse-phase HPLC of primary digests with protease V-8 and secondary digests with trypsin showed single major labeled components. The labeling of I-Ak by IASA-(46-61) was fully inhibitible by HEL-(46-61). In contrast, IASA attached to the smallest immunogenic peptide 52-61 (compound 3) labeled a distinctly different hydrophilic tryptic fragment. The site of the I-Ak molecule that was photoaffinity labeled by IASA-(46-61) (compound 2) was determined. IASA-(46-61) labeled selectively at Pro-118 of a primary alpha chain fragment most likely encompassing residues 115-134. It labeled Thr-121 of a primary beta chain fragment most likely encompassing residues 109-138. We also obtained evidence that IASA-(46-61) occupied the antigen-specific site; the conjugate stimulated a T-cell hybridoma that recognizes the sequence 52-61 and also competed for the binding of this smaller peptide to I-Ak. Thus, peptides that bind to the allele-specific binding site and are long enough to extend beyond it can interact with a hydrophobic area of class II molecules. This area is formed by sequences of the first halves of the second domain of both alpha and beta chains.
Mots-clé
Affinity Labels, Amino Acid Sequence, Binding Sites, Chromatography, High Pressure Liquid, Histocompatibility Antigens Class II/metabolism, Molecular Sequence Data, Muramidase/metabolism, Peptide Fragments/isolation & purification, Peptide Fragments/metabolism, Peptide Mapping, Photochemistry, Serine Endopeptidases/metabolism, Trypsin/metabolism
Pubmed
Web of science
Création de la notice
28/01/2008 11:20
Dernière modification de la notice
20/08/2019 14:28
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