Phenylalanine hydroxylase (PAH) and tyrosine hydroxylase (TH) are consecutive enzymes in the metabolic pathway leading to the production of catecholamine neurotransmitters. A comparison of recently available sequence data of these enzymes in the rat indicates about 70% homology in the 3' coding regions. We have localized TH by in situ hybridization to human chromosome region 11p15. Consideration of this assignment and that of PAH to chromosome 12, together with the known distribution of other pairs of related genes on these two chromosomes, provides convincing evidence of their ancestral relationship and suggests a role for gene duplication in the diversification of metabolic pathways in the vertebrate ancestors of mammals.