D-Amino acids are important building blocks for various compounds, such as pharmaceuticals and agrochemicals. A more cost-effective enzymatic method for D-amino acid production is needed in the industry. We improved a one-pot enzymatic method for D-amino acid production by the dynamic kinetic resolution of N-succinyl amino acids using two enzymes: D-succinylase (DSA) from Cupriavidus sp. P4-10-C, which hydrolyzes N-succinyl-D-amino acids enantio-selectively to their corresponding D-amino acid, and N-succinyl amino acid racemase (NSAR, EC.4.2.1.113) from Geo-bacillus stearothermophilus NCA1503. In this study, DSA and NSAR were purified and their properties were investigated. The optimum temperature of USA was 50 degrees C and it was stable up to 55 degrees C. The optimum pH of DSA and NSAR was around 7.5. In D-phenylalanine production, the optical purity of product was improved to 91.6% ee from the examination about enzyme concentration. Moreover, 100 mM N-succinyl-DL-tryptophan was converted to D-tryptophan at 81.8% yield with 94.7% ee. This enzymatic method could be useful for the industrial production of various n-amino acids. (C) 2017, The Society for Biotechnology, Japan. All rights reserved.