CHARACTERIZATION OF HUMAN VARIABLE DOMAIN ANTIBODY FRAGMENTS AGAINST THE U1 RNA-ASSOCIATED A-PROTEIN, SELECTED FROM A SYNTHETIC AND A PATIENT-DERIVED COMBINATORIAL V-GENE LIBRARY

This is the first study describing recombinant human antibody fragment s directed to the U1 RNA-associated A protein (U1A). Three anti-U1A an tibody fragments (Fab) were isolated from a semi-synthetic human Fab l ibrary and one anti-U1A single-chain variable fragment (scFv) was isol ated from a library which was derived from the IgG-positive splenic ly mphocytes of an autoimmune patient. Competition studies with autoantib odies against the U1 small nuclear ribonucleoprotein (snRNP) particle from patients with systemic lupus erythematosus (SLE) and SLE-overlap syndromes revealed that U1A binding of these antibody fragments can be inhibited by about 40% of the patient sera. All antibody fragments re cognized the native U1 snRNP in immunoprecipitation assays. Two of thr ee Fab clones as well as the scFv clone derived from the repertoire of an autoimmune patient use the same heavy chain germ-line gene DP-65. Epitope mapping revealed that these three clones appear to recognize a n identical epitope domain present on the C-terminal RNP motif of the U1A protein. The DP-65 heavy chain gene is used in less than 1% of the B cells in healthy individuals, while three out of four anti-U1A anti body fragments use this gene. This points to a restricted V-H gene usa ge in the case of U1A, suggesting that the DP-65 heavy chain has a nat ural shape complementarity to the U1A protein.