GTP cyclohydrolase I utilizes metal-free GTP as its substrate.
GTP cyclohydrolase I (GCH) is the rate-limiting enzyme for the synthesis of tetrahydrobiopterin and its activity is important in the regulation of monoamine neurotransmitters such as dopamine, norepinephrine and serotonin. We have studied the action of divalent cations on the enzyme activity of purified recombinant human GCH expressed in Escherichia coli. First, we showed that the enzyme activity is dependent on the concentration of Mg-free GTP. Inhibition of the enzyme activity by Mg2+, as well as by Mn2+, Co2+ or Zn2+, was due to the reduction of the availability of metal-free GTP substrate for the enzyme, when a divalent cation was present at a relatively high concentration with respect to GTP. We next examined the requirement of Zn2+ for enzyme activity by the use of a protein refolding assay, because the recombinant enzyme contained approximately one zinc atom per subunit of the decameric protein. Only when Zn2+ was present was the activity of the denatured enzyme effectively recovered by incubation with a chaperone protein. These are the first data demonstrating that GCH recognizes Mg-free GTP and requires Zn2+ for its catalytic activity. We suggest that the cellular concentration of divalent cations can modulate GCH activity, and thus tetrahydrobiopterin biosynthesis as well.
Suzuki
Takahiro
T
Department of Life Science, Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, Yokohama, Japan.
Kurita
Hideki
H
Ichinose
Hiroshi
H
D001708:Biopterins / Q000031:analogs & derivatives* / Q000378:metabolism*; D002413:Cations, Divalent / Q000494:pharmacology; D003035:Cobalt / Q000494:pharmacology; D004926:Escherichia coli / Q000201:enzymology*; D006136:GTP Cyclohydrolase / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism*; D006801:Humans; D007501:Iron / Q000494:pharmacology*; D008274:Magnesium / Q000494:pharmacology*; D008345:Manganese / Q000494:pharmacology; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D013379:Substrate Specificity
D002413:Cations, Divalent; D011994:Recombinant Proteins; D001708:Biopterins; D003035:Cobalt; D008345:Manganese; D006160:Guanosine Triphosphate; D007501:Iron; D006136:GTP Cyclohydrolase; C003402:sapropterin; D008274:Magnesium