Recombinant anti-P protein autoantibodies isolated from a human autoimmune library: reactivity, specificity and epitope recognition.
The ribosomal P proteins are specific and important autoantigens in patients affected by systemic lupus erythematosus. In this study, we describe for the first time the selection and characterization of recombinant human monoclonal anti-P protein (auto)-antibody fragments from an autoimmune patient-derived phage display antibody library. The selected recombinant anti-P antibodies specifically recognize the P proteins in immunofluorescence assays on HEp-2 cells and in immunoblotting assays, and they immunoprecipitate the P proteins under native conditions. Using both anti-P-positive patient sera and the selected recombinant anti-P antibodies, the immunodominant epitope was determined and shown to be located at the C-terminal end of the P proteins (amino acids 111-115). Inhibition of in vitro protein translation demonstrated that interaction of the monoclonal patient-derived anti-P antibodies with their native epitope functionally inhibits the activity of the P proteins on the ribosome, confirming the notion that patient autoantibodies are often directed to the functional centre of their autoantigenic target.
161 Department of Biochemistry, University of Nijmegen, P.O. Box 9101, 6500 HB Nijmegen, The Netherlands.
S
S
Zampieri
M
M
Mahler
M
M
Blüthner
Z
Z
Qiu
K
K
Malmegrim
A
A
Ghirardello
A
A
Doria
W J
WJ
van Venrooij
J M H
JM
Raats
D000918:Antibody Specificity; D001323:Autoantibodies / Q000276:immunology*; D015153:Blotting, Western; D018604:Epitope Mapping; D000939:Epitopes; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D008180:Lupus Erythematosus, Systemic / Q000276:immunology; D019151:Peptide Library; D011233:Precipitin Tests; D015800:Protozoan Proteins; D011994:Recombinant Proteins / Q000276:immunology; D012269:Ribosomal Proteins / Q000276:immunology*
D001323:Autoantibodies; D000939:Epitopes; C064601:L12E protein, Trypanosoma cruzi; D019151:Peptide Library; D015800:Protozoan Proteins; D011994:Recombinant Proteins; D012269:Ribosomal Proteins