DOI Seite / Zitierlink:
https://doi.org/10.11588/diglit.48120#0118
100 Molecular and Cell Biology of Autoantibodies and Autoimmunity
Anti-Jo-1 Antibodies are Directed at an Evolutionarily-Conserved,
Conformational Site on Human Histidyi-tRNA Synthetase
K. Waite, EW. Miller, and P. H. Plotz, N. I.H.
Arthritis and Rheumatism Branch, Clinical Center 9N244, National Institutes of Health,
Bethesda, MD 20892, USA
Anti-Jo-1 antibodies (AJoA) directed against the cellular enzyme histidyl-
tRNA synthetase (HRS) are found in approximately 25% of patients with
idiopathic myositis. They occur in a distinct subset of patients whose inflam-
matory muscle disease is often accompanied by interstitial lung disease, arthritis,
and Raynaud’s phenomenon. Less commonly, patients in this clinical group have
an autoantibody to a different aminoacyl-tRNA synthetase. The autoantibodies
target proteins which are functionally but not antigenically related.
In previous studies, we have shown that all antisera which bind to histidyl-
tRNA synthetase by western blotting or ELISA inhibit its enzymatic activity, and
that all antisera appear to recognize the same proteolytic fragments of the
polypeptide.
In order to identify any linear epitopes recognized by AJoA, all possible
overlapping hexapeptides corresponding to the 508 amino acid sequence of
human HRS, as determined by Tsui and Siminovich, were synthesized. These
hexapeptides were tested in an ELISA assay with IgG isolated from the sera of
myositis patients positive for AJoA, myositis patients negative for AJoA, and
normal volunteers. In addition, antibodies raised in rabbits against two decapep-
tides of human HRS which were predicted to be antigenic were assessed for reac-
tivity against the appropriate solid phase peptides.
As expected, the rabbit anti-peptide Abs reacted strongly with the correct hex-
apeptides corresponding to the immunogen. The Abs raised against peptide A
(residues 18-27) bound to hexapeptides starting with residues 18 through 24,
whereas the pre-immune sera from the same rabbit showed no reactivity to these
peptides. Thus, a minimum of four amino acid residues could be recognized by
Abs in this system. Similarly, Abs against peptide B (residues 436-445) bound
to hexapeptides starting with residue 435 through 441. Additional reactivity was
observed, however. The hexapeptides showing this additional reactivity posses no
sequence homology to the immunizing decamer and may, therefore, have been
recognized by other antibodies raised in response to the KLH and Freund’s ad-
juvant used for immunization.
Each of four AJoA positive sera tested showed a similar pattern of reactivity
across the entire sequence of HRS. The same pattern of reactivity was observed,
however, when the peptides were tested with the IgG from the sera of normal con-
trols (N-5) and AJoA-negative myositis patients (N-2), and from the AJoA-nega-
Anti-Jo-1 Antibodies are Directed at an Evolutionarily-Conserved,
Conformational Site on Human Histidyi-tRNA Synthetase
K. Waite, EW. Miller, and P. H. Plotz, N. I.H.
Arthritis and Rheumatism Branch, Clinical Center 9N244, National Institutes of Health,
Bethesda, MD 20892, USA
Anti-Jo-1 antibodies (AJoA) directed against the cellular enzyme histidyl-
tRNA synthetase (HRS) are found in approximately 25% of patients with
idiopathic myositis. They occur in a distinct subset of patients whose inflam-
matory muscle disease is often accompanied by interstitial lung disease, arthritis,
and Raynaud’s phenomenon. Less commonly, patients in this clinical group have
an autoantibody to a different aminoacyl-tRNA synthetase. The autoantibodies
target proteins which are functionally but not antigenically related.
In previous studies, we have shown that all antisera which bind to histidyl-
tRNA synthetase by western blotting or ELISA inhibit its enzymatic activity, and
that all antisera appear to recognize the same proteolytic fragments of the
polypeptide.
In order to identify any linear epitopes recognized by AJoA, all possible
overlapping hexapeptides corresponding to the 508 amino acid sequence of
human HRS, as determined by Tsui and Siminovich, were synthesized. These
hexapeptides were tested in an ELISA assay with IgG isolated from the sera of
myositis patients positive for AJoA, myositis patients negative for AJoA, and
normal volunteers. In addition, antibodies raised in rabbits against two decapep-
tides of human HRS which were predicted to be antigenic were assessed for reac-
tivity against the appropriate solid phase peptides.
As expected, the rabbit anti-peptide Abs reacted strongly with the correct hex-
apeptides corresponding to the immunogen. The Abs raised against peptide A
(residues 18-27) bound to hexapeptides starting with residues 18 through 24,
whereas the pre-immune sera from the same rabbit showed no reactivity to these
peptides. Thus, a minimum of four amino acid residues could be recognized by
Abs in this system. Similarly, Abs against peptide B (residues 436-445) bound
to hexapeptides starting with residue 435 through 441. Additional reactivity was
observed, however. The hexapeptides showing this additional reactivity posses no
sequence homology to the immunizing decamer and may, therefore, have been
recognized by other antibodies raised in response to the KLH and Freund’s ad-
juvant used for immunization.
Each of four AJoA positive sera tested showed a similar pattern of reactivity
across the entire sequence of HRS. The same pattern of reactivity was observed,
however, when the peptides were tested with the IgG from the sera of normal con-
trols (N-5) and AJoA-negative myositis patients (N-2), and from the AJoA-nega-