MAMMALIAN POLYPEPTIDE-CHAIN RELEASE FACTOR AND TRYPTOPHANYL-TRANSFER RNA-SYNTHETASE ARE DISTINCT PROTEINS

A very high (approximately 90%) structural similarity exists between t he bovine, human and murine tryptophanyl-tRNA synthetases (WRS), and q uite unexpectedly the rabbit polypeptide chain release factor (eRF). T his similarity may point to a very close resemblance or identity betwe en these proteins involved in distinct steps of protein synthesis, or inadvertently to an incorrect assignment of the clone reported to enco de eRF, since the structure of clones encoding WRS were confirmed by p eptide sequencing. Using high resolution column chromatography and suc rose gradient centrifugation combined with assays for WRS and eRF acti vities, we show that functionally distinct WRS and eRF proteins can be completely separated from each other. Moreover, a putative anti-eRF m onoclonal antibody appears incapable of immunoprecipitating the eRF ac tivity or binding to protein(s) possessing eRF activity. This antibody binds to protein fractions which coincide in various separation proce dures with rabbit WRS activity, and to pure bovine WRS. The protein ex pressed in Escherichia coli from the original cDNA clone initially rep orted to encode eRF, has WRS activity but not eRF activity. Resequenci ng of the fragment of the original rabbit cDNA demonstrates the presen ce of the previously overlooked HXGH motif typical of class I aminoacy l-tRNA synthetases. Consequently, mammalian WRS and eRF are different proteins, and the cDNA clone formerly assigned as encoding eRF encodes rabbit WRS.