Abstract
BackgroundAminoacyl-tRNA synthetases (AARSs) catalyze the first step of protein synthesis. Emerging evidence indicates that AARSs may have additional functions, playing a role in signal transduction pathways regulating thrombopoiesis and inflammation. Recombinant human tyrosyl-tRNA synthetase (rhTyrRS) is engineered with a single amino acid substitution that unmasks its cytokine activity. An industrial production method that provides high yield as well as high purity, quality, and potency of this protein is required for preclinical research.ResultsWe expressed codon-optimized rhTyrRS in Escherichia coli under fermentation conditions. Soluble protein was purified by a three-step purification method using cation exchange chromatography, gel filtration chromatography, and anion exchange chromatography. We also established a method to test the biological activity of rhTyrRS by measuring aminoacylation and IL-8 release in rhTyrRS-treated HL-60 cells.ConclusionsThe characterization of purified rhTyrRS indicated that this protein can be used in pharmacodynamic and pharmacokinetic studies.
Highlights
Aminoacyl-tRNA synthetases (AARSs) catalyze the first step of protein synthesis
Because the reactions require the capacity to recognize tRNAs as well as small chemicals such as amino acids and ATP, the structures of AARSs are well equipped for interacting with diverse molecules that may be associated with their functional versatility [1,2]
Human tyrosyl-tRNA synthetase (TyrRS) was the first example of a tRNA synthetase that played a role in cell signaling and thrombopoiesis
Summary
Aminoacyl-tRNA synthetases (AARSs) catalyze the first step of protein synthesis. Emerging evidence indicates that AARSs may have additional functions, playing a role in signal transduction pathways regulating thrombopoiesis and inflammation. Recombinant human tyrosyl-tRNA synthetase (rhTyrRS) is engineered with a single amino acid substitution that unmasks its cytokine activity. Aminoacyl-tRNA synthetases (AARSs) are enzymes that catalyze the essential first step of protein synthesis by covalently attaching each of the 20 standard amino acids to their cognate tRNA. Because the reactions require the capacity to recognize tRNAs as well as small chemicals such as amino acids and ATP, the structures of AARSs are well equipped for interacting with diverse molecules that may be associated with their functional versatility [1,2]. Human tyrosyl-tRNA synthetase (TyrRS) was the first example of a tRNA synthetase that played a role in cell signaling and thrombopoiesis. Mini-TyrRS has been shown to activate thrombocytopoiesis in a chick chorioallantoic membrane thrombopoiesis assay [8]
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