DAAO mutant sites among different mice strains and their effects on enzyme activity

Previous studies reported that the up-regulation of DAAO expression and enzyme activity levels were closely related to the development of neuropathic pain, cognitive characteristics of schizophrenia and so on. To determine DAAO mutant sites in different strains of mice and their effects on enzyme activity, we successfully constructed a prokaryotic expression system for heterologous expression of DAAO in vitro . There were total five nucleotide mutations distributed in exons 2, 8, 9, 10 of C57 mice. Three mutations located on exons 8 and 9 were synonymous mutations and had no variation on the encoded amino acid. The remaining two mutations in exons 2 (V64A) and 10 (R296H) were non-synonymous mutations, which might affect enzymatic activity and protein structure of mDAAO. Based on the determination of the kinetic constants and IC ₅₀ of mDAAO mutants in vitro , the differences in amino acid levels at these two sites (V64A, R295H) partly impacted the catalytic efficiency of DAAO mutants and their affinity with some inhibitors in vitro .