Abstract

Cysteine is a crucial component for all organisms and plays a critical role in the structure, stability, and catalytic functions of many proteins. Tetrahymena has reverse transsulfuration and de novo pathways for cysteine biosynthesis. Cysteine synthase is involved in the de novo cysteine biosynthesis and catalyzes the production of cysteine from O -acetylserine. The novel cysteine synthase TtCSA2 was identified from Tetrahymena thermophila . The TtCSA2 showed high expression levels at the log-phase and the sexual development stage. The TtCsa2 was localized on the outer mitochondrial membrane throughout different developmental stages. However, the truncated N -terminal signal peptide mutant TtCsa2-ΔN23 was localized into the mitochondria. His-TtCsa2 was expressed in Escherichia coli and purified using affinity chromatography. The His-TtCsa2 showed O -acetylserine sulfhydrylase and serine sulfhydrylase activities. Cysteine and glutathione contents decreased in the csa2 KD mutant. Furthermore, mutant cells were sensitive to cadmium and copper stresses. This study indicated that the TtCSA2 was involved in the cysteine synthesis in mitochondria and related to heavy metal stresses resistance in Tetrahymena .