Document Detail


Interaction of antigenic peptides with MHC class I molecules on living cells studied by photoaffinity labeling.
MedLine Citation:
PMID:  1737923     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Using a direct binding assay based on photoaffinity labeling, we have studied the interaction of antigenic peptides with murine MHC class I molecules on living cells. Photoreactive derivatives were prepared by N-terminal amidation with iodo, 4-azido salicylic acid of the Kd restricted Plasmodium berghei circumsporozoite (P.b. CS) peptide 253-260 (YIPSAEKI) and the Db-restricted Adenovirus 5 early region 1A (Ad5 E1A) peptide 234-243 (SGPSNTPPEI). As assessed in functional competition experiments, both peptide derivatives retained the specific binding activity of the parental peptides for Kd or Dd, respectively. The P.b. CS photoprobe specifically labeled Kd molecules on P815 (H-2d) cells, but failed to label RMA (H-2b) cells. Conversely, the Ad5 E1A photoprobe specifically labeled Db molecules on RMA cells, but failed to label P815 cells. When the two photoprobes were tested on a panel of Con A-activated spleen cells expressing 10 different H-2 haplotypes, significant photoaffinity labeling was observed only on H-2d cells with the P.b. CS photoprobe and on H-2b cells with the Ad5 E1A photoprobe. Labeling of cell-associated Kd or Db molecules with the photoprobes was specifically inhibited by antigenic peptides known to be presented by the same class I molecule. Photoaffinity labeling of Kd with the P.b. CS photoprobe was used to study the dynamics of peptide binding on living P815 cells. Binding increased steadily with the incubation period (up to 8 h) at 37 degrees C and at ambient temperature, but was greatly reduced (greater than 95%) at 0 to 4 degrees C or in the presence of ATP synthesis inhibitors. The magnitude of the labeling was twofold higher at room temperature than at 37 degrees C. In contrast, binding to isolated Kd molecules in solution rapidly reached maximal binding, particularly at 37 degrees C. Dissociation of the photoprobe from either cell-associated or soluble Kd molecules was similar, with a half time of approximately 1 h at 37 degrees C, whereas the complexes were long-lived at 4 degrees C in both instances.
Authors:
I F Luescher; J A Lóez; B Malissen; J C Cerottini
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Publication Detail:
Type:  Journal Article    
Journal Detail:
Title:  Journal of immunology (Baltimore, Md. : 1950)     Volume:  148     ISSN:  0022-1767     ISO Abbreviation:  J. Immunol.     Publication Date:  1992 Feb 
Date Detail:
Created Date:  1992-03-16     Completed Date:  1992-03-16     Revised Date:  2003-11-14    
Medline Journal Info:
Nlm Unique ID:  2985117R     Medline TA:  J Immunol     Country:  UNITED STATES    
Other Details:
Languages:  eng     Pagination:  1003-11     Citation Subset:  AIM; IM    
Affiliation:
Ludwig Institute for Cancer Research, Lausanne Branch, Epalinges, Switzerland.
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MeSH Terms
Descriptor/Qualifier:
Adenosine Triphosphate / physiology
Affinity Labels / metabolism*
Alleles
Amino Acid Sequence
Animals
H-2 Antigens / metabolism
Histocompatibility Antigens Class I / analysis,  metabolism*
Mice
Mice, Inbred C57BL
Molecular Sequence Data
Peptide Fragments / metabolism*
T-Lymphocytes, Cytotoxic / immunology
Temperature
Chemical
Reg. No./Substance:
0/Affinity Labels; 0/H-2 Antigens; 0/Histocompatibility Antigens Class I; 0/Peptide Fragments; 56-65-5/Adenosine Triphosphate

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