Cloning,expression and functional identification of a type III polyketide synthase gene from Huperzia serrata

A cDNA encoding novel type Ⅲ polyketide synthase(PKS) was cloned and sequenced from young leaves of Chinese club moss Huperzia serrata(Thunb.) Trev.by RT-PCR using degenerated primers based on the conserved sequences of known CHSs,and named as H.serrata PKS2.The terminal sequences of cDNA were obtained by the 3'-and 5'-RACE method.The full-length cDNA of H.serrata PKS2 contained a 1 212 bp open reading frame encoding a 46.4 kDa protein with 404 amino acids.The deduced amino acid sequence of H.serrata PKS2 showed 50 %-66% identities to those of other chalcone synthase super family enzymes of plant origin.The recombinant H.serrata PKS2 was functionally expressed in Escherichia coli with an additional hexahistidine tag at the N-terminus and showed unusually versatile catalytic potency to produce various aromatic tetraketides,including chalcones,benzophenones,phloroglucinols,and acridones.In particular,the enzyme accepted bulky starter substrates N-methylanthraniloyl-CoA,and carried out three condensations with malonyl-CoA to produce 1,3-dihydroxy-N-methylacridone.Interestingly,H.serrata PKS2 lacks most of the consensus active site sequences with acridone synthase from Ruta graveolens(Rutaceae).