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PMID: 1955465 Published · ppublish English Journal Article

Translocation of peptides through microsomal membranes is a rapid process and promotes assembly of HLA-B27 heavy chain and beta 2-microglobulin translated in vitro.

The Journal of cell biology ·Vol. 115 ·No. 4 ·1991-11-00 ·Pages 959-70

Lévy F, Larsson R, Kvist S

Abstract

We have translated major histocompatibility complex (MHC) class I heavy chains and human beta 2-microglobulin in vitro in the presence of microsomal membranes and a peptide from the nucleoprotein of influenza A. This peptide stimulates assembly of HLA-B27 heavy chain and beta 2-microglobulin about fivefold. By modifying this peptide to contain biotin at its amino terminus, we could precipitate HLA-B27 heavy chains with immobilized streptavidin, thereby directly demonstrating class I heavy chain-peptide association under close to physiological conditions. The biotin-modified peptide stimulates assembly to the same extent as the unmodified peptide. Both peptides bind to the same site on the HLA-B27 molecule. Immediately after synthesis of the HLA-B27 heavy chain has been completed, it assembles with beta 2-microglobulin and peptide. These interactions occur in the lumen of the microsomes (endoplasmic reticulum), demonstrating that the peptide must cross the microsomal membrane in order to promote assembly. The transfer of peptide across the microsomal membrane is a rapid process, as peptide binding to heavy chain-beta 2-microglobulin complexes is observed in less than 1 min after addition of peptide. By using microsomes deficient of beta 2-microglobulin (from Daudi cells), we find a strict requirement of beta 2-microglobulin for detection of peptide interaction with the MHC class I heavy chain. Furthermore, we show that heavy chain interaction with beta 2-microglobulin is likely to precede peptide binding. Biotin-modified peptides are likely to become a valuable tool in studying MHC antigen interaction and assembly.

MeSH Terms
Amino Acid Sequence Biological Transport Biotin Cell-Free System Culture Techniques Endoplasmic Reticulum/metabolism HLA-B27 Antigen/biosynthesis Histocompatibility Antigens Class I/biosynthesis Humans Intracellular Membranes/metabolism Kinetics Microsomes/metabolism Molecular Sequence Data Nucleoproteins/metabolism Peptides/metabolism Protein Biosynthesis beta 2-Microglobulin/biosynthesis
Chemicals
HLA-B27 Antigen Histocompatibility Antigens Class I Nucleoproteins Peptides beta 2-Microglobulin Biotin
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Lévy F
Ludwig Institute for Cancer Research, Stockholm, Sweden.
Larsson R
Kvist S
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Article Info
Journal
The Journal of cell biology
Abbr.
J Cell Biol
ISSN
0021-9525
DOI
10.1083/jcb.115.4.959
Published
1991-11-00
Pages
959-70
Language
English
Region
United States
NLM ID
0375356
PMCID
PMC2289957
Subset
IM
External Links
PubMed source DOI: 10.1083/jcb.115.4.959 PMC Full Text View journal details
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