LAUSR

In this report, the use of immobilized nicotine hydroxylase from Pseudomonas sp. ZZ-5 (HSPHZZ) for the production of 2,5-dihydroxypyridine (2,5-DHP) from 6-hydroxy-3-succinoylpyridine (HSP) in the presence of nicotinamide adenine dinucleotide (NADH) and flavin adenine dinucleotide (FAD) is described. HSPHZZ was covalently immobilized on Immobead 150 (ImmHSPHZZ). ImmHSPHZZ (obtained with 5–30 mg of protein per gram of support) catalyzed the hydrolysis of HSP to 2,5-DHP. At a protein loading of 15 mg g−1, ImmHSPHZZ converted 93.6% of HSP to 2,5-DHP in 6 h. The activity of ImmHSPHZZ was compared with that of free HSPHZZ under various conditions, including pH, temperature, enzyme concentration, substrate concentration and stability over time, and kinetic parameters were measured. The results showed that ImmHSPHZZ performed better over wider ranges of pH and temperature when compared with that of HSPHZZ. The optimal concentrations of ImmHSPHZZ and substrate were 30 mg L−1 and 0.75 mM, respectively. Under optimal conditions, 94.5 mg L−1 of 2,5-DHP was produced after 30 min with 85.4% conversion. After 8 reaction cycles and 6 days of storage, 51.3% and 75.0% of the initial enzyme activity remained, respectively. The results provide a framework for development of commercially suitable immobilized enzymes that produce 2,5-DHP.