Expression and characterization of the catalytic core of tryptophan hydroxylase

Graham R. Moran, S. Colette Daubner, Paul F. Fitzpatrick

Research output: Contribution to journalArticle

55 Scopus citations

Abstract

Wild type rabbit tryptophan hydroxylase (TRH) and two truncated mutant proteins have been expressed in Escherichia coli. The wild type protein was only expressed at low levels, whereas the mutant protein lacking the 101 amino-terminal regulatory domain was predominantly found in inclusion bodies. The protein that also lacked the carboxyl-terminal 28 amino acids, TRH102-416, was expressed as 30% of total cell protein. Analytical ultracentrifugation showed that TRH102-416 was predominantly a monomer in solution. The enzyme exhibited an absolute requirement for iron (ferrous or ferric) for activity and did not turn over in the presence of cobalt or copper. With either phenylalanine or tryptophan as substrate, stoichiometric formation of the 4α-hydroxypterin was found. Steady state kinetic parameters were determined with both of these amino acids using both tetrahydrobiopterin and 6-methyltetrahydropterin.

Original languageEnglish (US)
Pages (from-to)12259-12266
Number of pages8
JournalJournal of Biological Chemistry
Volume273
Issue number20
DOIs
StatePublished - May 15 1998

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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