Mitochondrial DNA depletion syndrome (MTDPS) is a group of severe mitochondrial disorders caused by nuclear gene variants that affect mitochondrial DNA (mtDNA) replication and nucleotide synthesis. Deoxyguanosine kinase deficiency is one of the most common subtypes, typically presenting with liver dysfunction in infancy and having a poor prognosis. We report a case of MTDPS presenting with cholestasis and mild hyperlactatemia in the neonatal period, which complicated early diagnosis. Histopathological and genetic analyses established the diagnosis. The patient, a female born at 36 weeks and 1 day of gestation, weighing 2124 g, developed cholestasis, poor feeding, and failure to thrive. Hyperlactatemia was not evident at presentation but gradually increased during the clinical course. Based on suspected mitochondrial disease, mitochondrial cocktail therapy was initiated on day 64. Liver transplantation was not feasible owing to cardiac and neurological complications, and conservative treatment was continued. However, the patient died of multiple organ failure on day 89. Postmortem liver biopsy showed a markedly reduced mtDNA copy number (8.1% of control), and genetic testing revealed a homozygous c.609_610del (p.Tyr204fs) variant in the