Characterization of anti-Z-RNA polyclonal antibodies: epitope properties and recognition of Z-DNA.
Chemically brominated poly[r(C-G)] [Br-poly[r(C-G)]] containing 32% br8G and 26% br5C was recently shown to contain a 1:1 mixture of A- and Z-form unmodified nucleotides under physiological conditions of temperature, pH, and ionic strength [Hardin, C. C., Zarling, D. A., Puglisi, J. D., Trulson, M. O., Davis, P. W., & Tinoco, I., Jr. (1987) Biochemistry 26, 5191-5199]. Proton NMR results show that more extensive bromination of poly[r(C-G)] (49% br8G, 43% br5C) produces polynucleotides containing greater than 80% unmodified Z-form nucleotides. Using these polynucleotides as antigens, polyclonal antibodies were elicited in rabbits and mice specific for the Z-form of RNA. IgG fractions were purified from rabbit anti-Br-poly[r(C-G)] sera and characterized by immunoprecipitation, nitrocellulose filter binding, and ELISA. Two different anti-Z-RNA IgG specificities were observed. Decreased levels of brominated nucleotides in the immunogen correlated with an increased extent of specific cross-reactivity with Z-DNA. Inoculation of rabbits with polynucleotide immunogens containing 49% br8G and 43% of br5C produced specific anti-Z-RNA IgGs that do not recognize Z-DNA determinants. This suggests that the 2'-OH group is part of the anti-Z-RNA IgG determinant. In contrast, Br-poly[r(C-G)] immunogens containing 32% br8G and 26% br5C produced IgGs that specifically recognize both Z-RNA and Z-DNA. These results show that the bromine atoms are not required for recognition of the Z conformation by the antibodies. The affinity of these anti-Z-RNA IgGs for Z-RNA is about 10-fold higher than for Z-DNA.(ABSTRACT TRUNCATED AT 250 WORDS)
Hardin
C C
CC
Department of Chemistry, University of California, Berkeley 94720.
Zarling
D A
DA
Wolk
S K
SK
Ross
W S
WS
Tinoco
I
I
D000818:Animals; D000906:Antibodies / Q000032:analysis*; D055598:Chemical Phenomena; D002621:Chemistry; D002942:Circular Dichroism; D004247:DNA / Q000032:analysis*; D000939:Epitopes / Q000032:analysis*; D007120:Immunochemistry; D007074:Immunoglobulin G / Q000032:analysis; D009682:Magnetic Resonance Spectroscopy; D011119:Polynucleotides / Q000032:analysis; D012313:RNA / Q000276:immunology*; D011817:Rabbits
D000906:Antibodies; D000939:Epitopes; D007074:Immunoglobulin G; D011119:Polynucleotides; D012313:RNA; D004247:DNA