Evidence of a self-catalytic mechanism of 2,4,5-trihydroxyphenylalanine quinone biogenesis in yeast copper amine oxidase

Abstract

Copper amine oxidases are representative of a new class of redox enzymes that contain a peptide-bound quinone cofactor, generated by posttranslational modification of amino acid side chain(s). We have investigated the mechanism for the biogenesis of 2,4,5-trihydroxyphenylalanine quinone (TPQ) in amine oxidase with two site-specific mutants of the yeast methylamine oxidase. Our results show that the capacity for TPQ formation in vivo is abolished when a putative ligand to copper, His-456, is changed to Asp; this H456D mutant binds copper at a low level (approximately 4.1%), relative to the wild-type protein. In contrast, altering the active site consensus sequence that contains the precursor tyrosine does not affect TPQ production. The data implicate a self-catalysis mechanism for TPQ biogenesis, in which the protein-bound copper plays a key role. We propose that the minimal information required for TPQ biogenesis lies in a structural motif consisting of the copper site and the precursor tyrosine.