Abstract
| Originalsprog | Engelsk |
|---|---|
| Tidsskrift | European Journal of Immunology |
| Vol/bind | 31 |
| Udgave nummer | 10 |
| Sider (fra-til) | 2986-96 |
| Antal sider | 10 |
| ISSN | 0014-2980 |
| Status | Udgivet - 2001 |
Bibliografisk note
Keywords: Animals; Disulfides; Escherichia coli; Histocompatibility Antigens Class I; Humans; Hydrogen-Ion Concentration; Mice; Peptides; Protein Folding; Recombinant Proteins; T-Lymphocytes; beta 2-MicroglobulinCitationsformater
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Efficient assembly of recombinant major histocompatibility complex class I molecules with preformed disulfide bonds. / Ostergaard Pedersen, L; Nissen, Mogens Holst; Hansen, N J; Nielsen, L L; Lauenmøller, S L; Blicher, T; Nansen, A; Sylvester-Hvid, C; Thomsen, Allan Randrup; Buus, S.
I: European Journal of Immunology, Bind 31, Nr. 10, 2001, s. 2986-96.Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › peer review
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TY - JOUR
T1 - Efficient assembly of recombinant major histocompatibility complex class I molecules with preformed disulfide bonds.
AU - Ostergaard Pedersen, L
AU - Nissen, Mogens Holst
AU - Hansen, N J
AU - Nielsen, L L
AU - Lauenmøller, S L
AU - Blicher, T
AU - Nansen, A
AU - Sylvester-Hvid, C
AU - Thomsen, Allan Randrup
AU - Buus, S
N1 - Keywords: Animals; Disulfides; Escherichia coli; Histocompatibility Antigens Class I; Humans; Hydrogen-Ion Concentration; Mice; Peptides; Protein Folding; Recombinant Proteins; T-Lymphocytes; beta 2-Microglobulin
PY - 2001
Y1 - 2001
N2 - The expression of major histocompatibility class I (MHC-I) crucially depends upon the binding of appropriate peptides. MHC-I from natural sources are therefore always preoccupied with peptides complicating their purification and analysis. Here, we present an efficient solution to this problem. Recombinant MHC-I heavy chains were produced in Escherichia coli and subsequently purified under denaturing conditions. In contrast to common practice, the molecules were not reduced during the purification. The oxidized MHC-I heavy chain isoforms were highly active with respect to peptide binding. This suggests that de novo folding of denatured MHC-I molecules proceed efficiently if directed by preformed disulfide bond(s). Importantly, these molecules express serological epitopes and stain specific T cells; and they bind peptides specifically. Several denatured MHC-I heavy chains were analyzed and shown to be of a quality, which allowed quantitative analysis of peptide binding. The analysis of the specificity of the several hundred human MHC haplotypes, should benefit considerably from the availability of pre-oxidized recombinant MHC-I.
AB - The expression of major histocompatibility class I (MHC-I) crucially depends upon the binding of appropriate peptides. MHC-I from natural sources are therefore always preoccupied with peptides complicating their purification and analysis. Here, we present an efficient solution to this problem. Recombinant MHC-I heavy chains were produced in Escherichia coli and subsequently purified under denaturing conditions. In contrast to common practice, the molecules were not reduced during the purification. The oxidized MHC-I heavy chain isoforms were highly active with respect to peptide binding. This suggests that de novo folding of denatured MHC-I molecules proceed efficiently if directed by preformed disulfide bond(s). Importantly, these molecules express serological epitopes and stain specific T cells; and they bind peptides specifically. Several denatured MHC-I heavy chains were analyzed and shown to be of a quality, which allowed quantitative analysis of peptide binding. The analysis of the specificity of the several hundred human MHC haplotypes, should benefit considerably from the availability of pre-oxidized recombinant MHC-I.
M3 - Journal article
C2 - 11592075
VL - 31
SP - 2986
EP - 2996
JO - European Journal of Immunology
JF - European Journal of Immunology
SN - 0014-2980
IS - 10
ER -