Biomolecular recognition of proteins and nucleic acids is mainly mediated by their structural features and the molecular dynamics simulations approach has been used to explore this recognition processes at the atomic level. L1-Endonuclease, an enzyme involved in L1 retrotransposition, cleaves the TA junction DNA (5'-TTTT/AA-3') and expresses high specificity for target site recognition. The present study highlights the structural features of L1-endonuclease as well as DNA responsible for such specific recognition. Especially, the importance of βB6-B5 hairpin loop in DNA recognition has been elucidated by analyzing the dynamics of Thr192 mutated L1-endonuclease. In addition, simulations of the endonuclease complexed with DNA substrates (sequences having TA and CG junctions) revealed the specificity of L1 endonuclease towards TA junction. Molecular dynamics simulations revealed that the βB6-B5 hairpin loop protrudes well into the minor groove of DNA having TA junction and induces DNA bending such that the width of minor groove is increased. Such endonuclease induced bending of TA junction DNA sequence positions the scissile phosphodiester bond of DNA for cleavage. The innate property of minor groove widening in TA junction than in CG junction is utilized by the βB6-βB5 hairpin loop of endonuclease while recognizing the DNA sequences. The present study also highlights the role of Mg cation in catalysis and attempts to explore the possible target site DNA cleavage mechanism.