Isolation and characterization of rabbit anti-m₃^2,2,7G antibodies

Antibodies specific for intact 2,2,7-trimethylguanosine ($m32,2,7G$) were induced by immunization of rabbits with a nucleoside-human serum albumen (HAS) conjugate. Competition radio-immunoassay showed that the antibody distinguishes well between intact $m32,2,7G$ and its alkali-hydrolysed form ($m32,2,7G*$). Antibody specificity is largely dependent on the presence of all three methyl groups in $m32,2,7G$: none of the less extensively methylated nucleosides M⁷G, M²G and $m32,2G$ is able to compete efficiently with the homologous hapten. Little or no competition was observed with m¹G, m¹A, m⁶A, m⁵U and each of the four unmodified ribonucleosides. Binding studies with nucleoplasmic RNAs from Ehrlich ascites cells suggest that the antibody reacts specifically with the $m32,2,7G$-containing cap structure of the small nuclear U-RNAs (U-snRNAs). Thus the antibody should be a valuable tool for studying the role of the 5'-terminal regions of the U-snRNAs of eukaryotic cells.