Rat GTP cyclohydrolase I is a homodecameric protein complex containing high-affinity calcium-binding sites

Steinmetz, M. O. and Pluss, C. and Christen, U. and Wolpensinger, B. and Lustig, A. and Werner, E. R. and Wachter, H. and Engel, A. and Aebi, U. and Pfeilschifter, J. and Kammerer, R. A.. (1998) Rat GTP cyclohydrolase I is a homodecameric protein complex containing high-affinity calcium-binding sites. Journal of molecular biology, Vol. 279, H. 1. pp. 189-199.

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Official URL: http://edoc.unibas.ch/dok/A5257692

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Recombinant rat liver GTP cyclohydrolase I has been prepared by heterologous gene expression in Escherichia coli and characterized by biochemical and biophysical methods. Correlation averaged electron micrograph images of preferentially oriented enzyme particles revealed a fivefold rotational symmetry of the doughnut-shaped views with an average particle diameter of 10 nm. Analytical ultracentrifugation and quantitative scanning transmission electron microscopy yielded average molecular masses of 270 kDa and 275 kDa, respectively. Like the Escherichia coli homolog, these findings suggest that the active enzyme forms a homodecameric protein complex consisting of two fivefold symmetric pentameric rings associated face-to-face. Examination of the amino acid sequence combined with calcium-binding experiments and mutational analysis revealed a high-affinity, EF-hand-like calcium-binding loop motif in eukaryotic enzyme species, which is absent in bacteria. Intrinsic fluorescence measurements yielded an approximate dissociation constant of 10 nM for calcium and no significant binding of magnesium. Interestingly, a loss of calcium-binding capacity observed for two rationally designed mutations within the presumed calcium-binding loop of the rat GTP cyclohydrolase I yielded a 45% decrease in enzyme activity. This finding suggests that failure of calcium binding may be the consequence of a mutation recently identified in the causative GTP cyclohydrolase I gene of patients suffering from dopa responsive dystonia.
Faculties and Departments:05 Faculty of Science > Departement Biozentrum > Former Organization Units Biozentrum > Structural Biology (Engel)
UniBasel Contributors:Engel, Andreas H
Item Type:Article, refereed
Article Subtype:Research Article
Note:Publication type according to Uni Basel Research Database: Journal article
Last Modified:22 Mar 2012 14:20
Deposited On:22 Mar 2012 13:18

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