Characterization of metal ligand mutants of tyrosine hydroxylase: Insights into the plasticity of a 2-histidine-1-carboxylate triad

Paul F. Fitzpatrick, Erik C. Ralph, Holly R. Ellis, Opal J. Willmon, S. Colette Daubner

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Abstract

The amino acid ligands to the active site iron in the aromatic amino acid hydroxylase tyrosine hydroxylase are two histidines and a glutamate. This 2-histidine-1-carboxylate motif has been found in a number of other metalloenzymes which catalyze a variety of oxygenase reactions. As a probe of the plasticity of this metal binding site, each of the ligands in TyrH has been mutated to glutamine, glutamate, or histidine. The H336E and H336Q enzymes show dramatic decreases in iron affinity but retain substantial activity for both tyrosine hydroxylation and tetrahydropterin oxidation. The H331E enzyme shows a lesser decrease in iron affinity and is unable to hydroxylate tyrosine. Instead, this enzyme oxidizes tetrahydropterin in the absence of added tyrosine. The E376H enzyme has no significant activity, while the E376Q enzyme hydroxylates tyrosine at about 0.4% the wild-type rate. When dopamine is bound to either the H336Q or H331E enzymes, the position of the long wavelength charge-transfer absorbance band is consistent with the change in the metal ligand. In contrast, the H336E enzyme does not form a stable binary complex with dopamine, while the E376H and E376Q enzymes catalyze dopamine oxidation.

Original languageEnglish (US)
Pages (from-to)2081-2088
Number of pages8
JournalBiochemistry
Volume42
Issue number7
DOIs
StatePublished - Feb 25 2003

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ASJC Scopus subject areas

  • Biochemistry

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