Functional analyses of a highly thermostable hexokinase from Pyrobaculum calidifontis.
Carbohydr Res
; 523: 108711, 2023 Jan.
Article
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| MEDLINE
| ID: mdl-36395717
ABSTRACT
The gene encoding a repressor open reading frame sugar kinase (ROK) family protein from hyperthermophilic crenarchaeon Pyrobaculum calidifontis, Pcal-HK, was cloned and expressed in Escherichia coli. The recombinant protein was produced in soluble and highly active form. Purified Pcal-HK was highly thermostable and existed in a monomeric form in solution. The enzyme was specific to ATP as phosphoryl donor but showed broad specificity to phosphoryl acceptors. It catalyzed the phosphorylation of a number of hexoses, including glucose, glucosamine, N-acetyl glucosamine, fructose and mannose, at nearly the same rate and similar affinity. The enzyme was metal ion dependent exhibiting highest activity at 90-95 °C and pH 8.5. Mg2+ was most effective metal ion, which could be partially replaced by Mn2+, Ni2+ or Zn2+. Kinetic parameters were determined at 90 °C and the enzyme showed almost similar catalytic efficiency (kcat/Km) towards the above mentioned hexoses. To the best of our knowledge, Pcal-HK is the most active thermostable ROK family hexokinase characterized to date which catalyzes the phosphorylation of various hexoses with nearly similar affinity.
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Base de datos:
MEDLINE
Asunto principal:
Pyrobaculum
/
Hexoquinasa
Idioma:
En
Revista:
Carbohydr Res
Año:
2023
Tipo del documento:
Article