Using Molecular Dynamics calculations, we find that the mode of DNA strand separation that is energetically favoured is where the helicase enzyme forces open the B1 bond first at an angle of ∼35 • . We also find that the minimum lifetime of the tautomer to be mechanistically feasible for genetic mutation is ∼1.6 ps. This minimum lifetime of the A*-T* base pair is two orders of magnitude shorter than previously suggested by Florian et al. To the best of our knowledge, our theoretical study provides the first strong argument against the widely-held belief that adenine-thymine tautomers are not relevant to spontaneous mutagenesis. By incorporating the dynamical process of DNA strand separation -a key step in the mitotic replication process and much more realistic than a static DNA picture -we show that adenine-thymine tautomerisation is just as relevant in genetic mutation mechanisms as guanine-cytosine tautomerisation.