Scallop phenylalanine hydroxylase implicates in immune response and can be induced by human TNF-α

Phenylalanine hydroxylase (PAH) is an important metabolic enzyme of aromatic amino acids, which is responsible for the irreversible oxidation of phenylalanine to tyrosine. In the present study, the full-length cDNA encoding PAH from Chlamys farreri (designated CfPAH) was cloned by using rapid amplification of cDNA ends (RACE) approaches and expression sequence tag (EST) analysis. The open reading frame of CfPAH encoded a polypeptide of 460 amino acids, and its sequence shared 64.4–74.2% similarity with those of PAHs from other animals. There were an N-terminal regulatory ACT domain and a C-terminal catalytic Biopterin_H domain in the deduced CfPAH protein. The mRNA transcripts of CfPAH could be detected in all the tested tissues, including adductor muscle, mantle, gill, gonad, haemocytes and hepatopancreas. And its expression level in haemocytes was increased significantly during 3–48 h after bacteria Vibrio anguillarum challenge with the highest level (9.1-fold, P < 0.05) at 24 h. Furthermore, the mRNA expression of CfPAH in haemocytes also increased significantly to 2.6-fold (P < 0.05) at 4 h and 3.3-fold (P < 0.05) at 6 h after the stimulation of 50.0 ng mL−1 human TNF-α. The cDNA fragment encoding the mature peptide of CfPAH was recombined and expressed in the prokaryotic expression system, and 1 mg recombinant CfPAH protein (rCfPAH) could catalyze the conversion of 192.23 ± 32.35 nmol phenylalanine to tyrosine within 1 min (nmol min−1 mg−1 protein) in vitro. These results indicated collectively that CfPAH, as a homologue of phenylalanine hydroxylase in scallop C. farreri, could be induced by cytokine and involved in the immunomodulation of scallops by supplying the starting material tyrosine for the synthesis of melanin and catecholamines.

► The CfPAH is a homologue of phenylalanine hydroxylase in Chlamys farreri.
► Its mRNA expression can be induced by bacteria challenge and human TNF-α stimulation.
► Recombinant CfPAH catalyses the irreversible oxidation of phenylalanine to tyrosine.
► CfPAH is involved in the immunomodulation of scallops.