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Abstract

Tryptophan hydroxylase (TPH) is the rate-limiting enzyme in serotonin biosynthesis. A novel gene, termed TPH2, has recently been described. This gene is preferentially expressed in the central nervous system, while the original TPH1 is the peripheral gene. We have expressed human tryptophan hydroxylase 2 (hTPH2) and two deletion mutants (N{Delta}150 and N{Delta}150/C{Delta}24) using isopropyl beta-D-thiogalactopyranoside-free autoinduction in Escherichia coli. This expression system produced active wild type TPH2 with relatively low solubility. The solubility was increased for mutants lacking the NH2-terminal regulatory domain. The solubility of hTPH2, N{Delta}150, and N{Delta}150/C{Delta}24 are 6.9, 62, and 97.5%, respectively. Removal of the regulatory domain also produced a more than 6-fold increase in enzyme stability (t1/2 at 37 °C). The wild type hTPH2, like other members of the aromatic amino acid hydroxylase superfamily, exists as a homotetramer (236 kDa on size exclusion chromatography). Similarly, N{Delta}150 also migrates as a tetramer (168 kDa). In contrast, removal of the NH2-terminal domain and the COOH-terminal, putative leucine zipper tetramerization domain produces monomeric enzyme (39 kDa). Interestingly, removal of the NH2-terminal regulatory domain did not affect the Michaelis constants for either substrate but did increase Vmax values. These data identify the NH2-terminal regulatory domain as the source of hTPH2 instability and reduced solubility.