Phenylketonuria (PKU) is an inborn error of metabolism caused by mutations in the phenylalanine hydroxylase ( PAH) gene or by defects in the tetrahydrobiopterin (BH4) synthesis pathway. Here, by positional cloning, we report that the 6-pyruvoyl-tetrahydropterin synthase ( PTPS) gene, encoding a key enzyme of BH4 biosynthesis, is responsible for the al c (albino C) mutation that displays pale body color, head shaking, and eventually lethality after the first molting in silkworm. Compared to wild type, the al c mutant produced more substrates (phenylalanine (Phe) and tyrosine (Tyr)) and generated less DOPA and dopamine. Application of 2,4-diamino-6-hydroxypyrimidine (DAHP) to block BH4 synthesis in the wild type effectively produced the al c -like phenotype, while BH4 supplementation rescued the defective body color and lethal phenotype in both al c and DAHP-treated individuals. The detection of gene expressions and metabolic substances after drugs treatments in al c and normal individuals imply that silkworms and humans have a high similarity in the drugs metabolic features and the gene pathway related to BH4 and the dopamine biosynthesis. We propose that the al c mutant could be used as an animal model for drug evaluation for BH4-deficient PKU.