In vitro mutagenesis of HLA-B27. Substitution of an unpaired cysteine residue in the α1 domain causes loss of antibody-defined epitopes

J. D. Taurog, F. A K El-Zaatari

Research output: Contribution to journalArticle

29 Citations (Scopus)

Abstract

The HLA class I molecules identified serologically as HLA-B27 are highly associated with ankylosing spondylitis and related human disorders. All known HLA-B27 amino acid sequences contain a cysteine residue at position 67; no other published HLA class I sequence contains a cysteine within the hypervariable region of the α1 domain, which extends from amino acid residues 63-84. To investigate the role of this cysteine residue in the antigenic structure of HLA-B27, we isolated a genomic clone encoding a molecule of the HLA-B27.1 subtype and performed oligonucleotide-directed mutagenesis to convert the cysteine at position 67 to a tyrosine. When transfected into mouse L cells, both the wild-type and Cys67 → Tyr67 mutant B27 genes directed the synthesis and surface expression of molecules reactive with the monomorphic anti-HLA class I antibody W6/32. However, only the L cells transfected with the wild-type B27 gene reacted with the anti-B27 antibody ME1; L cells transfected with the mutant B27 were completely unreactive with this antibody. Experiments with hybrid exons created from the HLA-B27 and HLA-A2 genes yielded results consistent with the mapping of the ME1 epitope to the B27 α1 domain. A second anti-B27 antibody, GS145.2, also showed markedly reduced binding to the Cys67 → Tyr67 mutant. These studies document the importance of the unique Cys67 residue in the antigenic structure of HLA-B27.

Original languageEnglish (US)
Pages (from-to)987-992
Number of pages6
JournalJournal of Clinical Investigation
Volume82
Issue number3
StatePublished - 1988

Fingerprint

HLA-B27 Antigen
Mutagenesis
Cysteine
Epitopes
Antibodies
Anti-Idiotypic Antibodies
Genes
Epitope Mapping
HLA-A2 Antigen
Immunoglobulin Isotypes
Ankylosing Spondylitis
Site-Directed Mutagenesis
Tyrosine
Amino Acid Sequence
Exons
Clone Cells
In Vitro Techniques
Amino Acids

ASJC Scopus subject areas

  • Medicine(all)

Cite this

@article{502ece3f5259467194a5f19b4e386cf7,
title = "In vitro mutagenesis of HLA-B27. Substitution of an unpaired cysteine residue in the α1 domain causes loss of antibody-defined epitopes",
abstract = "The HLA class I molecules identified serologically as HLA-B27 are highly associated with ankylosing spondylitis and related human disorders. All known HLA-B27 amino acid sequences contain a cysteine residue at position 67; no other published HLA class I sequence contains a cysteine within the hypervariable region of the α1 domain, which extends from amino acid residues 63-84. To investigate the role of this cysteine residue in the antigenic structure of HLA-B27, we isolated a genomic clone encoding a molecule of the HLA-B27.1 subtype and performed oligonucleotide-directed mutagenesis to convert the cysteine at position 67 to a tyrosine. When transfected into mouse L cells, both the wild-type and Cys67 → Tyr67 mutant B27 genes directed the synthesis and surface expression of molecules reactive with the monomorphic anti-HLA class I antibody W6/32. However, only the L cells transfected with the wild-type B27 gene reacted with the anti-B27 antibody ME1; L cells transfected with the mutant B27 were completely unreactive with this antibody. Experiments with hybrid exons created from the HLA-B27 and HLA-A2 genes yielded results consistent with the mapping of the ME1 epitope to the B27 α1 domain. A second anti-B27 antibody, GS145.2, also showed markedly reduced binding to the Cys67 → Tyr67 mutant. These studies document the importance of the unique Cys67 residue in the antigenic structure of HLA-B27.",
author = "Taurog, {J. D.} and El-Zaatari, {F. A K}",
year = "1988",
language = "English (US)",
volume = "82",
pages = "987--992",
journal = "Journal of Clinical Investigation",
issn = "0021-9738",
publisher = "The American Society for Clinical Investigation",
number = "3",

}

TY - JOUR

T1 - In vitro mutagenesis of HLA-B27. Substitution of an unpaired cysteine residue in the α1 domain causes loss of antibody-defined epitopes

AU - Taurog, J. D.

AU - El-Zaatari, F. A K

PY - 1988

Y1 - 1988

N2 - The HLA class I molecules identified serologically as HLA-B27 are highly associated with ankylosing spondylitis and related human disorders. All known HLA-B27 amino acid sequences contain a cysteine residue at position 67; no other published HLA class I sequence contains a cysteine within the hypervariable region of the α1 domain, which extends from amino acid residues 63-84. To investigate the role of this cysteine residue in the antigenic structure of HLA-B27, we isolated a genomic clone encoding a molecule of the HLA-B27.1 subtype and performed oligonucleotide-directed mutagenesis to convert the cysteine at position 67 to a tyrosine. When transfected into mouse L cells, both the wild-type and Cys67 → Tyr67 mutant B27 genes directed the synthesis and surface expression of molecules reactive with the monomorphic anti-HLA class I antibody W6/32. However, only the L cells transfected with the wild-type B27 gene reacted with the anti-B27 antibody ME1; L cells transfected with the mutant B27 were completely unreactive with this antibody. Experiments with hybrid exons created from the HLA-B27 and HLA-A2 genes yielded results consistent with the mapping of the ME1 epitope to the B27 α1 domain. A second anti-B27 antibody, GS145.2, also showed markedly reduced binding to the Cys67 → Tyr67 mutant. These studies document the importance of the unique Cys67 residue in the antigenic structure of HLA-B27.

AB - The HLA class I molecules identified serologically as HLA-B27 are highly associated with ankylosing spondylitis and related human disorders. All known HLA-B27 amino acid sequences contain a cysteine residue at position 67; no other published HLA class I sequence contains a cysteine within the hypervariable region of the α1 domain, which extends from amino acid residues 63-84. To investigate the role of this cysteine residue in the antigenic structure of HLA-B27, we isolated a genomic clone encoding a molecule of the HLA-B27.1 subtype and performed oligonucleotide-directed mutagenesis to convert the cysteine at position 67 to a tyrosine. When transfected into mouse L cells, both the wild-type and Cys67 → Tyr67 mutant B27 genes directed the synthesis and surface expression of molecules reactive with the monomorphic anti-HLA class I antibody W6/32. However, only the L cells transfected with the wild-type B27 gene reacted with the anti-B27 antibody ME1; L cells transfected with the mutant B27 were completely unreactive with this antibody. Experiments with hybrid exons created from the HLA-B27 and HLA-A2 genes yielded results consistent with the mapping of the ME1 epitope to the B27 α1 domain. A second anti-B27 antibody, GS145.2, also showed markedly reduced binding to the Cys67 → Tyr67 mutant. These studies document the importance of the unique Cys67 residue in the antigenic structure of HLA-B27.

UR - http://www.scopus.com/inward/record.url?scp=0023680060&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0023680060&partnerID=8YFLogxK

M3 - Article

VL - 82

SP - 987

EP - 992

JO - Journal of Clinical Investigation

JF - Journal of Clinical Investigation

SN - 0021-9738

IS - 3

ER -