Determination by Site-directed Mutagenesis of Sites in Sepiapterin Reductase Phosphorylated by Ca2+/Calmodulin-dependent Protein Kinase II

Abstract

Phosphorylation sites of sepiapterin reductase (SPR) phosphorylated by Ca2+-dependent protein kinase II (CaM KII) were studied. By immunoreaction against phosphorylated amino acids, we found that Ser residues of SPR were phosphorylated. We constructed several point mutants of SPR by site-directed mutagenesis and expressed then in E. coli. In assays with anti-phospho Ser antibody, we determined that each of the three Ser residues, S46, S 196, and S214, of SPR was phosphorylated by CaM KIl. Each of these serine residues in SPR was found in a CaM KII phosphorylation site sequelce (Arg-X-X-Ser/Thr).

Keywords

• sepiapterin reductase
• phosphorylation site
• ca2+-dependent protein kinase ii
• point mutant