Catalytic peptide of human glutaminyl-tRNA synthetase is essential for its assembly to the aminoacyl-tRNA synthetase complex.
Human glutaminyl-tRNA synthetase (QRS) is one of several mammalian aminoacyl-tRNA synthetases (ARSs) that form a macromolecular protein complex. To understand the mechanism of QRS targeting to the multi-ARS complex, we analyzed both exogenous and endogenous QRSs by immunoprecipitation after overexpression of various Myc-tagged QRS mutants in human embryonic kidney 293 cells. Whereas a deletion mutant containing only the catalytic domain (QRS-C) was targeted to the multi-ARS complex, a mutant QRS containing only the N-terminal appended domain (QRS-N) was not. Deletion mapping showed that the ATP-binding Rossman fold was necessary for targeting of QRS to the multi-ARS complex. Furthermore, exogenous Myc-tagged QRS-C was co-immunoprecipitated with endogenous QRS. Since glutaminylation of tRNA was dramatically increased in cells transfected with the full-length QRS, but not with either QRS-C or QRS-N, both the QRS catalytic domain and the N-terminal appended domain were required for full aminoacylation activity. When QRS-C was overexpressed, arginyl-tRNA synthetase and p43 were released from the multi-ARS complex along with endogenous QRS, suggesting that the N-terminal appendix of QRS is required to keep arginyl-tRNA synthetase and p43 within the complex. Thus, the eukaryote-specific N-terminal appendix of QRS appears to stabilize the association of other components in the multi-ARS complex, whereas the C-terminal catalytic domain is necessary for QRS association with the multi-ARS complex.
Kim
T
T
National Creative Research Initiatives Center for ARS Network, Sung Kyun Kwan University, Suwon, Kyunggido 440-746, Korea.
Park
S G
SG
Kim
J E
JE
Seol
W
W
Ko
Y G
YG
Kim
S
S
D000255:Adenosine Triphosphate / Q000378:metabolism; D000604:Amino Acyl-tRNA Synthetases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D020134:Catalytic Domain; D002460:Cell Line; D006801:Humans; D009097:Multienzyme Complexes / Q000737:chemistry / Q000378:metabolism*; D016296:Mutagenesis; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017384:Sequence Deletion; D014162:Transfection
D009097:Multienzyme Complexes; D011994:Recombinant Proteins; D000255:Adenosine Triphosphate; D000604:Amino Acyl-tRNA Synthetases; C020659:glutaminyl-tRNA synthetase