Endo-β-N-acetylglucosaminidases (ENGases) are the key enzymes showing dual specificity with an ability to catalyze hydrolysis and transglycosylation reactions.These enzymes have became the focal point for researchers because of their potentiality for synthesis of glycopeptides. Recently, few important ENGase mutants of Artherobacter protophormiae were screened that affects the catalytic activity of ENGase. Out of various mutants, the N171A mutant is having the property to abolish the whole enzymatic activity, both hydrolytic and transglycosylation activity (Jie Yin.,et al, 2009). In this study, we compared mutant N171A with native protein(PDB 3FHQ) to understand the structural changes, especially the hydrogen bonding pattern at the active site region which is causing the loss of catalytic activity in the mutant. The mutation position at 171 was mapped in the 3FHQ the native protein structure and a mutant with replacement of amino acid residue from Aspargine to Alanine was modeled by using homology modeling and simulation up to 10ns was done by Scrodinger Molecular modeling software. The results obtained from this computational analysis are correlated to experimental data and it Provides the potential insights to understand the key residues for catalytic activity of Atherobacter ENGase of both native and mutant protein to evaluate the stability, conformational differences .