Cyclohexylamine (CHAM) is widely used in various industries, but it is harmful to human beings and the environment. sp. YT-02 can degrade CHAM via cyclohexanone as an intermediate. In this study, the cyclohexylamine oxidase (CHAO) gene from sp. YT-02 was cloned. Amino acid sequence alignment indicated that the cyclohexylamine oxidase (CHAO) was 48% identical to its homolog from IH-35A (CHAO). The enzyme was expressed in BL21 (DE3), and purified to apparent homogeneity by Ni-affinity chromatography. The purified enzyme was proposed to be a dimer of molecular mass of approximately 91 kDa. The enzyme exhibited its maximum activity at 50°C and at pH 7.0. The enzyme was thermolabile as demonstrated by loss of important percentage of its maximal activity after 30 min incubation at 50°C. Metal ions Mg, Co, and K had certain inhibitory effect on the enzyme activity. The kinetic parameters and were 0.25 ± 0.02 mM and 4.3 ± 0.083 μM min, respectively. The biochemical properties, substrate specificities, and three-dimensional structures of CHAO and CHAO were compared. Our results are helpful to elucidate the mechanism of microbial degradation of CHAM in the strain YT-02. In addition, CHAO, as a potential biocatalyst, is promising in controlling CHAM pollution and deracemization of chiral amines.
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| http://dx.doi.org/10.3389/fmicb.2018.02848 | DOI Listing |
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