Enantiopure beta-amino acids are essential precursors of various pharmaceuticals, agrochemicals and other industrially important chemicals. In this study, we selected sixteen potential omega-Transaminases (omega-TAs) by BLAST and phylogenetic tree analysis. These omega-TAs were cloned, purified and tested for their reactivity for the synthesis of model beta-amino acid (R)-3-amino-4-(2,4,5-triflurophenyl) butanoic acid [3-ATfBA], a key precursor for sitagliptin. In an enzymatic cascade, lipase converted beta-ketoester substrate to beta-keto acid, which was subsequently aminated by the selected omega-TA to its corresponding beta-amino acid. A potent enzyme from Ilumatobacter coccineus (omega-TAIC) was identified for the production of 3-ATfBA. The pH dependency of the product inhibition suggested that lowering the reaction pH to 7.0 can circumvent the inhibition of omega-TALC by 3-ATfBA and about 92.3% conversion of 100 mM beta-keto ester substrate could be achieved. The applicability of this enzymatic system was further evaluated at the scale of 140 mM, wherein 3-ATfBA was generated with excellent conversion (81.9%) and enantioselectivity (99% ee). Furthermore, omega-TALC was successfully used for the synthesis of various beta-amino acids from their corresponding beta-keto ester substrates.