| dc.description.abstract | Aminoacyl–tRNA synthetases (aaRSs) belong to a ubiquitous and ancient family of enzymes that play an important role in protein synthesis by attaching a specific amino acid to its cognate tRNA. Since protein synthesis takes place in both cytoplasm and mitochondria in eukaryotes, two distinct sets of aaRSs are required, one functioning in the cytoplasm and the other in mitochondria. In most cases, the cytoplasmic and mitochondrial isozymes of an aaRS are encoded by two different nuclear genes, each recognizing its own tRNA isoacceptor. The only identity element in tRNAAla is G3:U70 in the acceptor stem through all three kingdoms of life. G3:U70 is recognized by two highly conserved residues Asp/Asn in alanyl-tRNA synthetase (AlaRS). Interestingly, mouse mitochondrial tRNAAla contains G1:U72, instead of G3:U70. We were wondering whether G1:U72 actually serves as the identity element for mouse mitochondrial tRNAAla. Pursuant to this objective, we have cloned the gene encoding mouse mitochondrial AlaRS (AlaRSm or AARS2) and sequenced the mature tRNAmAla. Our results showed that mouse AARS2 can charge yeast unfractionated tRNAs to a significant level, suggesting that certain non-cognate tRNAs with G1:U72 might be mischarged by this enzyme. Consistent with the finding, mouse AARS2 was toxic to yeast when expressed from a vector with a strong TEF1 promoter, suggesting that AARS2 might cause mistranslation by attaching alanine to non-cognate tRNAs with G1:U72. In addition, our sequencing data confirmed that the U5:U68 mismatch in the acceptor stem of mouse tRNAmAla is retained during processing. Moreover, purified mouse AARS2 could charge its own tRNAmAla (with G1:U72), but failed to charge H. sapiens tRNAAla or E. coli total tRNAs, suggesting that the anticodon and G3:U70 are not the major identity elements for recognition of tRNAAla by mouse AARS2. Thus, mouse AARS2 is a unique AlaRS that can recognize tRNAAla without the canonical identity element G3:U70. | en_US |