Mitochondrial helicase involved in mtDNA replication and repair (PubMed:12975372, PubMed:15167897, PubMed:17324440, PubMed:18039713, PubMed:18971204, PubMed:25824949, PubMed:26887820, PubMed:27226550). Might have a role in mtDNA repair (PubMed:27226550). Has DNA strand separation activity needed to form a processive replication fork for leading strand synthesis which is catalyzed by the formation of a replisome complex with POLG and mtSDB (PubMed:12975372, PubMed:15167897, PubMed:18039713, PubMed:22383523, PubMed:26887820, PubMed:27226550). Preferentially unwinds DNA substrates with pre-existing 5'-and 3'- single-stranded tails but is also active on a 5'- flap substrate (PubMed:12975372, PubMed:15167897, PubMed:18039713, PubMed:22383523, PubMed:26887820, PubMed:27226550). Can dissociate the invading strand of immobile or mobile D-loop DNA structures irrespective of the single strand polarity of the third strand (PubMed:27226550). In addition to its DNA strand separation activity, also has DNA strand annealing, DNA strand-exchange and DNA branch migration activities (PubMed:22383523, PubMed:26887820, PubMed:27226550). Lack DNA unwinding and ATP hydrolysis activities (PubMed:18039713). Does not bind single-stranded or double-stranded DNA (PubMed:18039713). ATP + H2O = ADP + H(+) + phosphate Strand annealing activity is inhibited by 150 mM NaCl (in vitro). Interacts with LONP1. Homohexamer (via C-terminus) which assembled in a ring-like structure (PubMed:18039713, PubMed:17324440, PubMed:22383523, PubMed:25824949, PubMed:30496414, PubMed:18971204). Homoheptamer which assembled in a ring-like structure (PubMed:25824949, PubMed:30496414). Oligomerization is Mg(2+), nucleotide and DNA-independent, however, Mg(2+) and nucleotide stabilize the homohexameric form (PubMed:18039713). Interacts with POLG in vitro (PubMed:15167897). Monomer (PubMed:18039713). Does not form oligomers (PubMed:18039713). Colocalizes with mtDNA in mitochondrial nucleoids, a nucleoproteins complex consisting of a number of copies of proteins associated with mtDNA, probably involved in mtDNA maintenance and expression. High relative levels in skeletal muscle, testis and pancreas. Lower levels of expression in the heart, brain, placenta, lung, liver, kidney, spleen, thymus, prostate, ovary, small intestine, colon and leukocytes. Expression is coregulated with MRPL43. N-terminus enhances protein stability and hexamer formation, which is important for DNA binding, and is required for DNA helicase activity and, ultimately, for mtDNA replisome processivity. The disease is caused by variants affecting the gene represented in this entry. The disease is caused by variants affecting the gene represented in this entry. The disease is caused by variants affecting the gene represented in this entry. The N-terminus contains a putative primase-like domain; however the absence of the zinc binding domain and other motifs important for catalysis suggests that TWNK lacks primase activity. In vitro, can catalyze the hydrolysis of different nucleotide triphosphate (NTP) substrates with different efficiency.