Mammalian polypeptide chain release factor and tryptophanyl-tRNA synthetase are distinct proteins.
A very high (approximately 90%) structural similarity exists between the bovine, human and murine tryptophanyl-tRNA synthetases (WRS), and quite unexpectedly the rabbit polypeptide chain release factor (eRF). This similarity may point to a very close resemblance or identity between these proteins involved in distinct steps of protein synthesis, or inadvertently to an incorrect assignment of the clone reported to encode eRF, since the structure of clones encoding WRS were confirmed by peptide sequencing. Using high resolution column chromatography and sucrose gradient centrifugation combined with assays for WRS and eRF activities, we show that functionally distinct WRS and eRF proteins can be completely separated from each other. Moreover, a putative anti-eRF monoclonal antibody appears incapable of immunoprecipitating the eRF activity or binding to protein(s) possessing eRF activity. This antibody binds to protein fractions which coincide in various separation procedures with rabbit WRS activity, and to pure bovine WRS. The protein expressed in Escherichia coli from the original cDNA clone initially reported to encode eRF, has WRS activity but not eRF activity. Resequencing of the fragment of the original rabbit cDNA demonstrates the presence of the previously overlooked HXGH motif typical of class I aminoacyl-tRNA synthetases. Consequently, mammalian WRS and eRF are different proteins, and the cDNA clone formerly assigned as encoding eRF encodes rabbit WRS.
Frolova LYu
Institut Jacques Monod, Paris, France.
Dalphin
M E
ME
Justesen
J
J
Powell
R J
RJ
Drugeon
G
G
McCaughan
K K
KK
Kisselev
L L
LL
Tate
W P
WP
Haenni
A L
AL
D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D001483:Base Sequence; D002850:Chromatography, Gel; D003001:Cloning, Molecular; D004247:DNA; D004926:Escherichia coli; D006801:Humans; D015151:Immunoblotting; D008969:Molecular Sequence Data; D010454:Peptide Termination Factors / Q000378:metabolism*; D011817:Rabbits; D017386:Sequence Homology, Amino Acid; D014369:Tryptophan-tRNA Ligase / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D010454:Peptide Termination Factors; C070289:peptide chain release factor, mammalian; D000255:Adenosine Triphosphate; D004247:DNA; D014369:Tryptophan-tRNA Ligase