ADP-dependent glucokinase from the hyperthermophilic sulfate-reducing archaeon Archaeoglobus fulgidus strain 7324.
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 degrees C and showed a significant thermostability up to 100 degrees 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 degrees C). Only glucosamine was phosphorylated at significant rates. The apparent K(m) values for ADP and glucose (at 50 degrees C) were 0.07 mM and 0.78 mM, respectively; the apparent V(max) value was about 50 U/mg at 50 degrees C and 350 U/mg at 80 degrees C. Divalent cations were required for maximal activity; Mn(2+), Mg(2+ )and Ca(2+), which were most effective, could be replaced partially by Cu(2+), Ni(2+), Co(2+) and Zn(2+). 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.
Labes
Antje
A
Institut für Allgemeine Mikrobiologie, Christian-Albrechts-Universität Kiel, Am Botanischen Garten 1-9, 24118 Kiel, Germany.
Schönheit
Peter
P
D001105:Archaea; D019611:Archaeoglobus fulgidus / Q000201:enzymology* / Q000254:growth & development / Q000378:metabolism; D019766:Genes, Archaeal; D005941:Glucokinase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D020539:Sequence Analysis, Protein; D013379:Substrate Specificity; D013431:Sulfates / Q000378:metabolism*
D013431:Sulfates; C420842:ADP-dependent glucokinase; D005941:Glucokinase