A unique insertion in the CP1 domain of Giardia lamblia leucyl-tRNA synthetase.
Leucyl-tRNA synthetase (LeuRS) catalyzes the esterification of the tRNA(Leu) isoacceptor with leucine. It contains a large insertion domain, connective peptide 1 (CP1), for amino acid editing. Here, we cloned the gene encoding LeuRS from Giardia lamblia (GlLeuRS), one of the most ancient eukaryotes. GlLeuRS was purified from an Escherichia coli overproduction strain, and its properties were investigated. The isolated CP1 domain of GlLeuRS (GlLeuRS-CP1) was an active protein for editing mischarged G. lamblia tRNA(Leu)(AAG) (GltRNA(Leu)). Insertion of 49 amino acid residues within the CP1 domain (the so-called 49-amino acid motif) was important for the optimal aminoacylation activity of GlLeuRS and was crucial for the editing capacity of GlLeuRS-CP1. Additionally, the motif can confer editing activity on the editing-defective isolated CP1 domain from E. coli LeuRS (EcLeuRS-CP1). We also found that GlLeuRS could not rescue a Saccharomyces cerevisiae leuS null strain, suggesting different recognition modes for these two LeuRSs with respect to tRNA(Leu).
Zhou
Xiao-Long
XL
State Key Laboratory of Molecular Biology, Graduate School of the Chinese Academy of Sciences, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, The Chinese Academy of Sciences, People's Republic of China.
Yao
Peng
P
Ruan
Liang-Liang
LL
Zhu
Bin
B
Luo
Jun
J
Qu
Liang-Hu
LH
Wang
En-Duo
ED
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000201:enzymology; D017125:Genes, Protozoan; D016829:Giardia lamblia / Q000201:enzymology* / Q000235:genetics; D006868:Hydrolysis; D007700:Kinetics; D007935:Leucine-tRNA Ligase / Q000737:chemistry* / Q000235:genetics*; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D017434:Protein Structure, Tertiary; D017393:RNA Editing; D012346:RNA, Transfer, Amino Acyl / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D016415:Sequence Alignment; D017384:Sequence Deletion; D046249:Transfer RNA Aminoacylation
D012346:RNA, Transfer, Amino Acyl; D007935:Leucine-tRNA Ligase