Method for fixing key enzymes and cofactors in formaldehyde decomposition path and application thereof
A formaldehyde decomposition, key enzyme technology, applied in the field of enzyme engineering
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Embodiment 1
[0022] Immobilization of recombinant protein of PADH and recombinant protein of FDH and its cofactor NAD:
[0023] Add the following solutions to the 30ml reaction system: 12ml of 30% acrylamide, 7.8ml of 1.5mol / L (pH7.5) potassium phosphate buffer, 7.9ml of sterile water, 1.5mg of PADH recombinant protein, 2.25mg of FDH recombinant protein, 100mmol / L cofactor NAD + 2ml, stirred slowly at room temperature for 20min to mix the enzyme protein, cofactor and acrylamide carrier matrix evenly, then added 0.3ml of 10% ammonium persulfate (APS), 0.012ml of TEMED, stirred slowly for 2min to completely cross-link the acrylamide monomer into After polyacrylamide, quickly pour the mixture into the gel maker that has been installed, and place it at room temperature for 3-4 hours to make the polyacrylamide completely solidify, carefully peel off the gel, and cut it into a length of 5cm with a clean scalpel , 1cm wide and 0.5cm thick strips, put them into a 6ml empty chromatographic column....
Embodiment 2
[0025] Analysis of the absorption effect of immobilized enzyme on formaldehyde:
[0026] Use a syringe to simultaneously inject 6ml of gaseous formaldehyde into the upper end of the chromatography tube and the control tube (blank tube) equipped with immobilized enzyme strips, and the outlet of the tube is connected to a hand-held formaldehyde analyzer ( figure 1 B), the measured initial concentration of formaldehyde in the tube is 0.47ppm, and the remaining formaldehyde concentration in the tube is measured every 1 hour. Take the remaining formaldehyde concentration in the tube as the ordinate, and take the measurement time as the abscissa to draw the formaldehyde absorption curve as follows: figure 2 shown. The results show that compared with the blank tube, the immobilized enzyme does have an absorption effect on formaldehyde, especially in the first 3 hours, the formaldehyde decline rate is faster, from the initial 0.47ppm to 0.32ppm, and 32% of the injected gas formaldeh...
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