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Characterization of Wild‐Type and Mutants of Recombinant Human GTP Cyclohydrolase I
Author(s) -
Suzuki Takahiro,
Ohye Tamae,
Inagaki Hidehito,
Nagatsu Toshiharu,
Ichinose Hiroshi
Publication year - 1999
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1046/j.1471-4159.1999.0732510.x
Subject(s) - gtp cyclohydrolase i , mutant , missense mutation , recombinant dna , microbiology and biotechnology , frameshift mutation , wild type , mutant protein , biology , gtp' , mutation , western blot , enzyme , biochemistry , tetrahydrobiopterin , gene , cofactor
: To explore the molecular etiology of two disorders caused by a defect in GTP cyclohydrolase I—hereditary progressive dystonia with marked diurnal flucturation (HPD), also known as dopa‐responsive dystonia (DRD), and autosomal recessive GTP cyclohydrolase I deficiency—we purified and analyzed recombinant human wild‐type and mutant GTP cyclohydrolase I proteins expressed in Escherichia coli . Mutant proteins showed very low enzyme activities, and some mutants were eluted at a delayed volume on gel filtration compared with the recombinant wild‐type. Next, we examined the GTP cyclohydrolase I protein amount by western blot analysis in phytohemagglutinin‐stimulated mononuclear blood cells from HPD/DRD patients. We found a great reduction in the amount of the enzyme protein not only in one patient who had a frameshift mutation, but also in an HPD/DRD patient who had a missense mutation. These results suggest that a dominant‐negative effect of chimeric protein composed of wild‐type and mutant subunits is unlikely as a cause of the reduced enzyme activity in HPD/DRD patients. We suggest that reduction of the amount of the enzyme protein, which is independent of the mutation type, could be a reason for the dominant inheritance in HPD/DRD.