Insilico engineering of transaminase variants for enhanced biocatalytic conversion of an ACE inhibitor precursor

Angiotensin-converting enzyme (ACE) inhibitors are widely prescribed for cardiovascular disorders, yet their conventional chemical synthesis involves multiple steps, high energy consumption, and poor stereoselectivity. In this work, we present a fully insilico framework for the computational design and prediction of transaminase (TA) variants capable of catalyzing the asymmetric amination of ethyl 2-oxo-4-phenylbutanoate (OPBE) to yield L-homophenylalanine ethyl ester (L-HPE), a key chiral intermediate in ACE inhibitor synthesis. A homology model of the Silicibacter transaminase (62.7 % identity to 5KR6) was constructed in its dimeric form. Eleven active-site variants were designed and screened through molecular docking, followed by 100 ns molecular dynamics simulations. The top variants SbTA10, SbTA01, and SbTA11 exhibited reactive distances below 6 Å, binding energies between -17.7 and -20.8 kcal/mol, and substrate RMSD values under 2.0 Å, indicating stable enzyme-substrate complexes. A composite QZ-score integrating productive conformations, binding energy, pocket contacts, and structural stability ranked SbTA10 highest (0.89), followed by SbTA01 (0.88) and SbTA11 (0.85). Free-energy profiles derived from umbrella sampling revealed binding minima of -10.2 kcal/mol (SbTA10) and -8.1 kcal/mol (SbTA11), suggesting distinct substrate retention characteristics that may influence catalytic turnover. Collectively, these results identify plausible transaminase variants with favorable structural and energetic features for the proposed OPBE to L-HPE transformation. This study presents a computational framework for predicting transaminase variants, providing a basis for rational biocatalyst design that warrants future experimental validation to confirm catalytic efficiency and stereoselectivity.