The hyperthermophilic sulfate-reducing archaeon Archaeoglobus fulgidus strain 7324 has been shown to degrade starch via glucose using a modified Embden-Meyerhof pathway. The first enzyme of this pathway, ADP-dependent glucokinase, was purified 600-fold to homogeneity. The enzyme is a monomeric protein with an apparent molecular mass of 50 kDa. It had a temperature optimum at 83 °C and showed a significant thermostability up to 100 °C. The enzyme was highly specific for ADP and glucose as substrates; it did not use ATP, CDP, UDP, or GDP as phosphoryl donors, or mannose, fructose and fructose 6-phosphate as phosphoryl acceptors (at 80 °C). Only glucosamine was phosphorylated at significant rates. The apparent Km values for ADP and glucose (at 50 °C) were 0.07 mM and 0.78 mM, respectively; the apparent Vmax value was about 50 U/mg at 50 °C and 350 U/mg at 80 °C. Divalent cations were required for maximal activity; Mn2+, Mg2+ and Ca2+, which were most effective, could be replaced partially by Cu2+, Ni2+, Co2+ and Zn2+. The N-terminal amino acid sequence (42 amino acids) of ADP-dependent glucokinase was almost identical to that of ADP-dependent glucokinase from Thermococcus litoralis. In the genome of the closely related Archaeoglobus fulgidus strain VC16 a homologous gene for ADP-dependent glucokinase could not be identified.
