Cell-Free Synthesis of Dopamine and Serotonin in Two Steps with Purified Enzymes

Abstract

The synthesis of serotonin and dopamine with purified enzymes is described. Both pathways start from an amino acid substrate and synthesize the monoamine neurotransmitter in two enzymatic steps. The enzymes human tryptophan hydroxylase isoform 2, Rattus norvegicus tyrosine hydroxylase, Chlamydia pneumoniae Cpn1046, and aromatic amino acid decarboxylase from Drosophila melanogaster are recombinantly expressed, purified, and shown to be functional in vitro. The hydroxylases efficiently convert L-DOPA (L-dihydroxy-phenylalanine) and 5-HTP (5-hydroxytryptophan) from L-tyrosine and L-tryptophan, respectively. A single aromatic amino acid decarboxylase is capable of converting both hydroxylated intermediates into the final neurotransmitter. The platform described here may facilitate future efforts to generate medically useful artificial cells and nanofactories.

Keywords: cell-free synthetic biology; dopamine; enzymatic synthesis; serotonin.