Identification and characterization of human mitochondrial tryptophanyl-tRNA synthetase

A full-length cDNA clone encoding the human mitochondrial tryptophanyl-tRNA synthetase (h(mt)TrpRS) has been identified. The deduced amino acid sequen ce shows high homology to both the mitochondrial tryptophanyl-tRNA syntheta se ((mt)TrpRS) from Saccharomyces cerevisiae and to different eubacterial f orms of tryptophanyl-tRNA synthetase (TrpRS). Using the baculovirus express ion system, we have expressed and purified the protein with a carboxyl-term inal histidine tag. The purified His-tagged h(mt)TrpRS catalyzes Trp-depend ent exchange of PPi in the PPi-ATP exchange assay. Expression of h(mt)TrpRS in both human and insect cells leads to high levels of h(mt)TrpRS localizi ng to the mitochondria, and in insect cells the first 18 amino acids consti tute the mitochondrial localization signal sequence, Until now the human cy toplasmic tryptophanyl-tRNA synthetase (hTrpRS) was thought to function as the h(mt)TrpRS, possibly in the form of a splice variant. However, no mitoc hondrial localization signal sequence was ever detected and the present ide ntification of a different (mt)TrpRS almost certainly rules out that possib ility. The h(mt)TrpRS shows kinetic properties similar to human mitochondri al phenylalanyl-tRNA synthetase (h(mt)-PheRS), and h(mt)TrpRS is not induce d by interferon-gamma as is hTrpRS.