A simple two step procedure for purification of the catalytic domain of chicken tryptophan hydroxylase 1 in a form suitable for crystallization

Tryptophan hydroxylase (TPH) [EC 1.14.16.4] catalyzes the conversion of tryptophan to 5-hydroxytryptophan, which is the first and rate-determining step in the biosynthesis of the neurotransmitter serotonin. We have expressed the catalytic domain of chicken (Gallus gallus) TPH isoform 1 in Escherichia coli in high yield. The enzyme was highly purified using only one anion exchange and one gel filtration, with a yield of 11mg/L culture and a specific activity of 0.60μmol/min/mg. The Km values were determined to Km,tryptophan=7.7±0.7μM, Km,BH4=324±10μM and Km,O2=39±2μM. Substrate inhibition by tryptophan was observed at concentrations above 15μM. Furthermore, the purified enzyme has been crystallized without 7,8-dihydro-l-biopterin and a data set to 3Å resolution has been collected.