In this study, thermostable Thermococcus kodakarensis L-asparaginase (TkA) enzyme, which lacks glutaminase activity, was studied for its structural and dynamic properties. The structural and dynamic properties of TkA, was investigated in its apo state and with the L-asparagine ligand to understand how the active site and general structure of the TkA enzyme changes with ligand binding and what effect this interaction has on the general behavior of the enzyme. T11E, T55E and D86S mutants of TkA were examined by molecular docking and molecular dynamics simulations. Binding results for molecular docking indicate that the structure is largely conserved, with root mean square deviation (RMSD) scores of −5.2 to −5.7 nm for wildtype TkA and mutants. RMSD and root mean square fluctuations (RMSF) data obtained as a result of molecular dynamics studies showed that the mutants had a stability close to that of the WT TkA enzyme, between 0.15 and 0.16 nm. In general, solvent accessible surface area (SASA) and radius of gyration analysis results support this analysis, while the D86S mutant gave more effective results than other mutants with SASA value of 260.38 nm2 /ns and radius of gyration values of 2.61 nm/ns. The total interaction energy of the ligand and WT TkA was -337.98 kJ/mol, while the interaction energy for D86S mutant was larger, at -363.03 kJ/mol. In conclusion, the study showing how the structure and dynamics of the TkA enzyme are affected by the binding of L-asparagine ligand helps to understand the stability and functional behavior of the enzyme.