MOLECULAR-CLONING OF A PEPTIDYLGLYCINE ALPHA-HYDROXYLATING MONOOXYGENASE FROM SEA-ANEMONES

Cnidarians are the lowest animal group having a nervous system. The pr imitive nervous systems of cnidarians produce large amounts of a varie ty of neuropeptides, of which many or perhaps all are amidated at thei r C terminus. In vertebrates, peptide amidation is catalyzed by two en zymes acting sequentially, peptidyl-glycine alpha-hydroxylating monoox ygenase (PHM) and peptidyl-alpha-hydroxyglycine alpha-amidating lyase (PAIL). In mammals both enzymatic activities are contained within a bi functional protein that is coded for by a single gene. Using PCR and d egenerated oligonucleotides derived from conserved regions of PHM, we have now cloned a PHM from the sea anemone Calliactis parasitica showi ng 42% amino acid sequence identity with rat PHM. Among the conserved (identical) amino acid residues are five histidine and one methionine residue, which bind two Cu2+ atoms that are essential for PHM activity . No cDNA coding for PAL could be identified, suggesting that sea anem one PAI, is coded for by a gene that is different from the sea anemone PHM gene, a situation similar to the one found in insects. This is th e first report on the molecular cloning of a cnidarian PHM. (C) 1997 A cademic Press.