Kinetics and thermodynamics for aldo/keto conversion of D‐xylose and D‐glucose catalyzed by xylose isomerase from Thermotoga naphthophila RKU ‐10

BACKGROUND Aldo‐keto isomerizations of D‐xylose and D‐glucose catalyzed by xylose isomerase (XI) is commercially carried out to produce bio‐ethanol and high fructose corn syrup. XI is a metalloenzyme. In this paper, kinetics and thermodynamics of the activity and stability of hyperthermophilic recombinant XI from Thermotoga naphthophila RKU‐10 T (TnapXI) were studied in the presence and absence of metal ions. RESULTS Kinetics parameters of recombinant XI from TnapXI were calculated as K m  = 0.99 mmol L –1 , V max  = 500 μmol.mg −1 .min −1 , and k cat  = 68.3 min −1 for the isomerization of D‐xylose (aldo) ⇌D‐xylulose (keto) , whereas for D‐glucose (aldo) ⇌D‐fructose (keto) conversion K m  = 7.72 mmol L –1 , V max  = 90 μmol.mg −1 .min −1 , and k cat  = 12.4 min −1 . Ionization constants were calculated as pK a1  = 6.0 and pK a2  = 7.6, whereas activation constants ( K a ) of TnapXI were 0.3, 0.5, and 4.0 mmol L –1 for Co 2+ , Mn 2+ , and Mg 2+ , respectively. Inhibition constant ( K i ) was 0.119 mmol L –1 for Ca 2+ . Q 10 was 2.8 and E a  = 82.25 kJ.mol −1 . Thermodynamics for isomerization were calculated as ∆ H * = 79.2 kJ.mol −1 , ∆ G * = 83 kJ.mol −1 , and ∆ S * = −10.5 J.mol −1 .K −1 . The z ‐value was 12.6 °C for apo‐TnapXI and 32.6 °C for holo‐TnapXI. Deactivation parameters were ∆ G * (d)  = 100 kJ.mol −1 , ∆ H * (d)  = 206 kJ.mol −1 , ∆ S * (d)  = 0.28 kJ.mol −1 .K −1 for apo‐TnapXI and t 1/2  = 18 min at 95 °C, while ∆ G * (d)  = 104 kJ.mol −1 , ∆ H * (d)  = 767 kJ.mol −1 , ∆ S * (d)  = 1.8 kJ.mol −1 .K −1 for holo‐TnapXI and t 1/2  = 65 min at 95 °C. CONCLUSION These noteworthy features make this enzyme an attractive candidate for the industrial production of fuel ethanol and high fructose corn syrup at high temperatures to maximize the product yield. © 2021 Society of Chemical Industry (SCI).