THE SECONDARY STRUCTURE OF A PYRIMIDINE-GUANINE SEQUENCE-SPECIFIC RIBONUCLEASE POSSESSING CYTOTOXIC ACTIVITY FROM THE OOCYTES OF RANA-CATESBEIANA

RC-RNase is a pyrimidine-guanine sequence-specific ribonuclease and a sialic-acid-binding lectin purified from Rana catesbeiana (bullfrog) o ocytes. This Ill-amino acid protein exhibits cytotoxicity toward sever al tumor cell lines. In this paper we report the assignments of proton NMR resonances and the identification of the secondary structure dedu ced from NOE constraints, chemical shift index, (3)J(NH alpha) and ami de proton exchange rates. The protein was directly isolated from bullf rog oocytes; we were able to assign all but five of the amino acid bac kbone protons of the unlabeled protein by analyzing a large set of two -dimensional proton NMR spectra obtained at several temperatures and p H conditions. Our results indicate that the structure of RC-RNase is d ominated by the presence of two triple-stranded antiparallel beta-shee ts and three alpha-helices, similar to those of the pyrimidine family ribonucleases. Two sets of resonances were observed for 11 amide proto ns and 8 alpha-protons located in the loop-1 region, an alpha 2 helix, and three beta-strands (beta 1, beta 3 and beta 4), suggesting the pr esence of nonlocalized multiple conformations for RC-RNase.