Alloreactive cytotoxic T lymphocytes recognize epitopes determined by both the α helices and β sheets of the class I peptide binding site

H. D. Hunt, T. I. Munitz, Larry R Pease

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Abstract

A chimeric class I glycoprotein was created to investigate the functional contribution of the α helices and the β-pleated sheets in forming the antigen recognition site (ARS) of antigen-presenting molecules. This novel molecule was generated by replacing the DNA sequences encoding the α helices of the L(d) gene with the corresponding sequences from the Kb gene. Serologic analysis of transfected L cells that expressed the chimeric molecule (Kb/(α)L(d)/(β)) revealed that the engineered class I glycoprotein retains two conformational epitopes associated with the α helices of Kb, as defined by monoclonal antibodies K10.56 and 28-13-3. These results demonstrate that the α helices of Kb can associate with the β- pleated sheets of L(d) to form a stable structure, which is expressed on the cell surface. To address the role of the α helices of the ARS in determining T cell crossreactivity, alloreactive cytotoxic T lymphocytes (CTL) were used to analyze L cells expressing Kb/(α)L(d)/(β). CTL raised against Kb or L(d) as alloantigens showed little, if any, ability to lyse L cells expressing Kb/(α)L(d)/(β). Thus, alloreactive CTL did not recognize structures determined by the α helices alone or by the β sheets of the ARS alone. However, bulk and cloned alloreactive CTL that were generated against the mutant Kb glycoprotein Kbm8 reacted strongly with Kb/(α)L(d)/(β). In addition to the Kb α helices, the Kbm8 ARS shares a single polymorphic amino acid at position 24 with Kb/(α)L(d)/(β). Amino acid 24 is located on the β2 strand that forms part of the floor of the ARS and has been identified as a component of pocket B in the HLA class I ARS. The substitution of Glu to Ser at this position was shown previously to be the central determinant of the Kbm8 mutant alloantigenicity. The functional significance of this position in determining crossreactivity between bm8 and Kb/(α)L(d)/(β) identifies pocket B as a strong anchor for allogenic self- peptides. These findings demonstrate that determinants recognized by CTL on class I alloantigens are formed by interactions involving both the α helices and β sheets of the ARS. These interactions are best explained by the influence of the α helices and β sheets on the peptide-binding properties of these antigen-presenting molecules.

Original languageEnglish (US)
Pages (from-to)821-829
Number of pages9
JournalJournal of Experimental Medicine
Volume175
Issue number3
StatePublished - 1992

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T-Lymphocyte Epitopes
Cytotoxic T-Lymphocytes
Binding Sites
Antigens
Peptides
Glycoproteins
Isoantigens
Amino Acids
Histocompatibility Antigens Class I
HLA Antigens
Genes
Epitopes
Monoclonal Antibodies
T-Lymphocytes

ASJC Scopus subject areas

  • Immunology

Cite this

@article{9784646472b947de9a07c6f86f4b2d5a,
title = "Alloreactive cytotoxic T lymphocytes recognize epitopes determined by both the α helices and β sheets of the class I peptide binding site",
abstract = "A chimeric class I glycoprotein was created to investigate the functional contribution of the α helices and the β-pleated sheets in forming the antigen recognition site (ARS) of antigen-presenting molecules. This novel molecule was generated by replacing the DNA sequences encoding the α helices of the L(d) gene with the corresponding sequences from the Kb gene. Serologic analysis of transfected L cells that expressed the chimeric molecule (Kb/(α)L(d)/(β)) revealed that the engineered class I glycoprotein retains two conformational epitopes associated with the α helices of Kb, as defined by monoclonal antibodies K10.56 and 28-13-3. These results demonstrate that the α helices of Kb can associate with the β- pleated sheets of L(d) to form a stable structure, which is expressed on the cell surface. To address the role of the α helices of the ARS in determining T cell crossreactivity, alloreactive cytotoxic T lymphocytes (CTL) were used to analyze L cells expressing Kb/(α)L(d)/(β). CTL raised against Kb or L(d) as alloantigens showed little, if any, ability to lyse L cells expressing Kb/(α)L(d)/(β). Thus, alloreactive CTL did not recognize structures determined by the α helices alone or by the β sheets of the ARS alone. However, bulk and cloned alloreactive CTL that were generated against the mutant Kb glycoprotein Kbm8 reacted strongly with Kb/(α)L(d)/(β). In addition to the Kb α helices, the Kbm8 ARS shares a single polymorphic amino acid at position 24 with Kb/(α)L(d)/(β). Amino acid 24 is located on the β2 strand that forms part of the floor of the ARS and has been identified as a component of pocket B in the HLA class I ARS. The substitution of Glu to Ser at this position was shown previously to be the central determinant of the Kbm8 mutant alloantigenicity. The functional significance of this position in determining crossreactivity between bm8 and Kb/(α)L(d)/(β) identifies pocket B as a strong anchor for allogenic self- peptides. These findings demonstrate that determinants recognized by CTL on class I alloantigens are formed by interactions involving both the α helices and β sheets of the ARS. These interactions are best explained by the influence of the α helices and β sheets on the peptide-binding properties of these antigen-presenting molecules.",
author = "Hunt, {H. D.} and Munitz, {T. I.} and Pease, {Larry R}",
year = "1992",
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pages = "821--829",
journal = "Journal of Experimental Medicine",
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T1 - Alloreactive cytotoxic T lymphocytes recognize epitopes determined by both the α helices and β sheets of the class I peptide binding site

AU - Hunt, H. D.

AU - Munitz, T. I.

AU - Pease, Larry R

PY - 1992

Y1 - 1992

N2 - A chimeric class I glycoprotein was created to investigate the functional contribution of the α helices and the β-pleated sheets in forming the antigen recognition site (ARS) of antigen-presenting molecules. This novel molecule was generated by replacing the DNA sequences encoding the α helices of the L(d) gene with the corresponding sequences from the Kb gene. Serologic analysis of transfected L cells that expressed the chimeric molecule (Kb/(α)L(d)/(β)) revealed that the engineered class I glycoprotein retains two conformational epitopes associated with the α helices of Kb, as defined by monoclonal antibodies K10.56 and 28-13-3. These results demonstrate that the α helices of Kb can associate with the β- pleated sheets of L(d) to form a stable structure, which is expressed on the cell surface. To address the role of the α helices of the ARS in determining T cell crossreactivity, alloreactive cytotoxic T lymphocytes (CTL) were used to analyze L cells expressing Kb/(α)L(d)/(β). CTL raised against Kb or L(d) as alloantigens showed little, if any, ability to lyse L cells expressing Kb/(α)L(d)/(β). Thus, alloreactive CTL did not recognize structures determined by the α helices alone or by the β sheets of the ARS alone. However, bulk and cloned alloreactive CTL that were generated against the mutant Kb glycoprotein Kbm8 reacted strongly with Kb/(α)L(d)/(β). In addition to the Kb α helices, the Kbm8 ARS shares a single polymorphic amino acid at position 24 with Kb/(α)L(d)/(β). Amino acid 24 is located on the β2 strand that forms part of the floor of the ARS and has been identified as a component of pocket B in the HLA class I ARS. The substitution of Glu to Ser at this position was shown previously to be the central determinant of the Kbm8 mutant alloantigenicity. The functional significance of this position in determining crossreactivity between bm8 and Kb/(α)L(d)/(β) identifies pocket B as a strong anchor for allogenic self- peptides. These findings demonstrate that determinants recognized by CTL on class I alloantigens are formed by interactions involving both the α helices and β sheets of the ARS. These interactions are best explained by the influence of the α helices and β sheets on the peptide-binding properties of these antigen-presenting molecules.

AB - A chimeric class I glycoprotein was created to investigate the functional contribution of the α helices and the β-pleated sheets in forming the antigen recognition site (ARS) of antigen-presenting molecules. This novel molecule was generated by replacing the DNA sequences encoding the α helices of the L(d) gene with the corresponding sequences from the Kb gene. Serologic analysis of transfected L cells that expressed the chimeric molecule (Kb/(α)L(d)/(β)) revealed that the engineered class I glycoprotein retains two conformational epitopes associated with the α helices of Kb, as defined by monoclonal antibodies K10.56 and 28-13-3. These results demonstrate that the α helices of Kb can associate with the β- pleated sheets of L(d) to form a stable structure, which is expressed on the cell surface. To address the role of the α helices of the ARS in determining T cell crossreactivity, alloreactive cytotoxic T lymphocytes (CTL) were used to analyze L cells expressing Kb/(α)L(d)/(β). CTL raised against Kb or L(d) as alloantigens showed little, if any, ability to lyse L cells expressing Kb/(α)L(d)/(β). Thus, alloreactive CTL did not recognize structures determined by the α helices alone or by the β sheets of the ARS alone. However, bulk and cloned alloreactive CTL that were generated against the mutant Kb glycoprotein Kbm8 reacted strongly with Kb/(α)L(d)/(β). In addition to the Kb α helices, the Kbm8 ARS shares a single polymorphic amino acid at position 24 with Kb/(α)L(d)/(β). Amino acid 24 is located on the β2 strand that forms part of the floor of the ARS and has been identified as a component of pocket B in the HLA class I ARS. The substitution of Glu to Ser at this position was shown previously to be the central determinant of the Kbm8 mutant alloantigenicity. The functional significance of this position in determining crossreactivity between bm8 and Kb/(α)L(d)/(β) identifies pocket B as a strong anchor for allogenic self- peptides. These findings demonstrate that determinants recognized by CTL on class I alloantigens are formed by interactions involving both the α helices and β sheets of the ARS. These interactions are best explained by the influence of the α helices and β sheets on the peptide-binding properties of these antigen-presenting molecules.

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