4. Conclusion
To improve the thermostability of DAADH, several co-evolved interfacial residue pairs were identified. Based on the amino acid residue interaction network and co-evolutionary analysis, those residues were mutated for redesign the interfacial interaction of subunits. Eleven novel DAADHs were obtained. Mutations induced new salt bridges and hydrogen bonds. The average number of salt bridges of DA06 and DA11 are 1.4 and 1.2-fold of that of DAwild, respectively. Thermostability study indicated the half-life of DA11 was 2-fold of DAwild. MDS was applied to uncover the mechanism of enhanced thermostability from molecular level. These findings suggest that the dimer interface motifs are essential for the compactness and stability of DAADHs and may illuminate design based on evolution in other enzyme families.