Tryptophan hydroxylase exists in two isoforms: Isoform 1 catalyses the first and rate-limiting step in the synthesis of serotonin in the peripheral parts of the body while isoform 2 catalyses this step in the brain. The catalytic domains of human tryptophan hydroxylase 1 and 2 have been expressed, purified and the kinetic properties have been studied and are compared. Substrate inhibition by tryptophan is observed for isoform 1 but not for isoform 2. Large differences are observed in the K (m,tetrahydrobiopterin) values for the two isoforms, being >10 times larger for isoform 1 compared to isoform 2.

Tryptophan hydroxylase 2 ( TPH 2) catalyses the initial and rate‐limiting step in the biosynthesis of serotonin, which is associated with a variety of disorders such as depression, obsessive compulsive disorder, and schizophrenia. Full‐length TPH 2 is poorly characterized due to low purification quantities caused by its inherent instability. Three truncated variants of human TPH 2 (rc h TPH 2; regulatory and catalytic domain, NΔ47‐rc h TPH 2; truncation of 47 residues in the N terminus of rc h TPH 2, and c h TPH 2; catalytic domain) were expressed, purified, and examined for changes in transition temperature, inactivation rate, and oligomeric state. c h TPH 2 displayed 14‐ and 11‐fold higher half‐lives compared to rc h TPH 2 and NΔ47‐rc h TPH 2, respectively. Differential scanning calorimetry experiments demonstrated that this is caused by premature unfolding of the less stable regulatory domain. By differential scanning fluorimetry, the unfolding transitions of rc h TPH 2 and NΔ47‐rc h TPH 2 are found to shift from polyphasic to apparent two‐state by the addition of l ‐Trp or l ‐Phe. Analytical gel filtration revealed that rc h TPH 2 and NΔ47‐rc h TPH 2 reside in a monomer–dimer equilibrium which is significantly shifted toward dimer in the presence of l ‐Phe. The dimerizing effect induced by l ‐Phe is accompanied by a stabilizing effect, which resulted in a threefold increase in half‐lives of rc h TPH 2 and NΔ47‐rc h TPH 2. Addition of l ‐Phe to the purification buffer significantly increases the purification yields, which will facilitate characterization of h TPH 2.

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