Strains, plasmids, and media
All strains and plasmids used in this study are listed in Table S1 . The variants were constructed by whole plasmid PCR protocol with the main primers listed in Table S2 . The PCR system (100 µl) was composed of template (100‐150 ng), corresponding primers (20 µM with 2 µl), PrimeSTAR polymerase (1 µl; Takara Biomedical Technology, Dalian, China), 5 × PrimeSTAR Buffer (20 µl), dNTP mix (8 µl), and sterilized water. Next, DpnI was added to the PCR reaction mixture and incubated for 3 h at 37°C to eliminate the template plasmid. The digested product was transformed into Escherichia coli BL21(DE3) cells for the following screening or DNA sequencing (Genewiz, China). Luria-Bertani (LB) medium containing 5 g·L-1 yeast extract, 10 g·L-1 tryptone, and 10 g·L-1 NaCl was used for strain selection and propagation. Terrific Broth (TB) medium containing 4 g·L-1 glycerin, 24 g·L-1 yeast extract, 12 g·L-1tryptone, 2.31 g·L-1KH2PO4, and 16.43 g·L-1 K2HPO4 was used for protein expression. Chloramphenicol (30 μg·mL-1), kanamycin (50 μg·mL-1), and ampicillin (100 μg·mL-1), and IPTG (0.4 mM) were added at the appropriate time.