New insights into copper monooxygenases and peptide amidation: structure, mechanism and function.
Many bioactive peptides must be amidated at their carboxy terminus to exhibit full activity. Surprisingly, the amides are not generated by a transamidation reaction. Instead, the hormones are synthesized from glycine-extended intermediates that are transformed into active amidated hormones by oxidative cleavage of the glycine N-C alpha bond. In higher organisms, this reaction is catalyzed by a single bifunctional enzyme, peptidylglycine alpha-amidating monooxygenase (PAM). The PAM gene encodes one polypeptide with two enzymes that catalyze the two sequential reactions required for amidation. Peptidylglycine alpha-hydroxylating monooxygenase (PHM; EC 1.14.17.3) catalyzes the stereospecific hydroxylation of the glycine alpha-carbon of all the peptidylglycine substrates. The second enzyme, peptidyl-alpha-hydroxyglycine alpha-amidating lyase (PAL; EC 4.3.2.5), generates alpha-amidated peptide product and glyoxylate. PHM contains two redox-active copper atoms that, after reduction by ascorbate, catalyze the reduction of molecular oxygen for the hydroxylation of glycine-extended substrates. The structure of the catalytic core of rat PHM at atomic resolution provides a framework for understanding the broad substrate specificity of PHM, identifying residues critical for PHM activity, and proposing mechanisms for the chemical and electron-transfer steps in catalysis. Since PHM is homologous in sequence and mechanism to dopamine beta-monooxygenase (DBM; EC 1.14.17.1), the enzyme that converts dopamine to norepinephrine during catecholamine biosynthesis, these structural and mechanistic insights are extended to DBM.
Prigge
S T
ST
Department of Biophysics and Biophysical Chemistry, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
Mains
R E
RE
Eipper
B A
BA
Amzel
L M
LM
D019760:Amidine-Lyases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D003300:Copper / Q000378:metabolism*; D004299:Dopamine beta-Hydroxylase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D006899:Mixed Function Oxygenases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes; D011487:Protein Conformation; D051381:Rats; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D009097:Multienzyme Complexes; D003300:Copper; D006899:Mixed Function Oxygenases; D004299:Dopamine beta-Hydroxylase; C035189:peptidylglycine monooxygenase; D019760:Amidine-Lyases; C068521:peptidylamidoglycolate lyase