Thymidine kinase-2 (TK2) is a deoxyribonucleoside kinase that phosphorylates thymidine, deoxycytidine, and deoxyuridine, and also phosphorylates antiviral and anticancer nucleoside analogs. Both recombinant and native forms of the enzyme have broad substrate specificity and complex kinetics, suggesting that it may play a role in the activation of chemotherapeutic nucleoside analogs. The highest levels of expression are observed in testis and ovary. The main supply of deoxyribonucleotides (dNTPs) for mitochondrial DNA synthesis comes from the salvage pathway initiated by deoxyguanosine kinase (DGK) and TK2. The association of mitochondrial DNA depletion with mutations in the genes encoding these 2 kinases suggests that the salvage pathway enzymes are involved in the maintenance of balanced mitochondrial dNTP pools.