Abstract
Peptide binding reactions of class II MHC proteins exhibit unusual kinetics, with extremely slow apparent rate constants for the overall association (<100 M(-)(1) s(-)(1)) and dissociation (<10(-)(5) s(-)(1)) processes. Various linear and branched pathways have been proposed to account for these data. Using fluorescence resonance energy transfer between tryptophan residues in the MHC peptide binding site and aminocoumarin-labeled peptides, we measured real-time kinetics of peptide binding to empty class II MHC proteins. Our experiments identified an obligate intermediate in the binding reaction. The observed kinetics were consistent with a binding mechanism that involves an initial bimolecular binding step followed by a slow unimolecular conformational change. The same mechanism is observed for different peptide antigens. In addition, we noted a reversible inactivation of the empty MHC protein that competes with productive binding. The implications of this kinetic mechanism for intracellular antigen presentation pathways are discussed.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Antigens, Differentiation, B-Lymphocyte / chemistry
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Antigens, Differentiation, B-Lymphocyte / genetics
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Antigens, Differentiation, B-Lymphocyte / metabolism
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Energy Transfer
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Escherichia coli / genetics
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Genetic Vectors / chemistry
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Genetic Vectors / metabolism
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HLA-DR1 Antigen / chemistry*
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HLA-DR1 Antigen / genetics
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HLA-DR1 Antigen / metabolism*
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Hemagglutinin Glycoproteins, Influenza Virus / chemistry
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Hemagglutinin Glycoproteins, Influenza Virus / genetics
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Hemagglutinin Glycoproteins, Influenza Virus / metabolism
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Histocompatibility Antigens Class II / chemistry
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Histocompatibility Antigens Class II / genetics
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Histocompatibility Antigens Class II / metabolism
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Humans
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Kinetics
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Models, Chemical
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Oligopeptides / chemistry*
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Oligopeptides / genetics
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Oligopeptides / metabolism*
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Protein Binding / genetics
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Spectrometry, Fluorescence
Substances
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Antigens, Differentiation, B-Lymphocyte
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HLA-DR1 Antigen
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Hemagglutinin Glycoproteins, Influenza Virus
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Histocompatibility Antigens Class II
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Oligopeptides
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invariant chain