Functionalization of oxidases with peroxidase activity creates oxiperoxidases: a new breed of hybrid enzyme capable of cascade chemistry.
The covalent flavoprotein alditol oxidase (AldO) from Streptomyces coelicolor A3(2) was endowed with an extra catalytic functionality by fusing it to a microperoxidase. Purification of the construct resulted in the isolation of a synthetic bifunctional enzyme that was both fully covalently flavinylated and heminylated: an oxiperoxidase. Characterization revealed that both oxidase and peroxidase functionalities were active, with the construct functioning as a single-component xylitol biosensor. In an attempt to reduce the size of the oxidase-peroxidase fusion, we replaced portions of the native AldO sequence with the bacterial cytochrome c CXXCH heme-binding motif. By mutating only three residues of the AldO protein we were able to create a functional oxidase-peroxidase hybrid.
Laboratory of Biochemistry, University of Groningen, Nijenborgh 4, 9747 AG Groningen, The Netherlands.
Remko T
RT
Winter
Tomas E
TE
van den Berg
Dana I
DI
Colpa
Edwin
E
van Bloois
Marco W
MW
Fraaije
D004591:Electrophoresis, Polyacrylamide Gel; D005182:Flavin-Adenine Dinucleotide / Q000737:chemistry / Q000378:metabolism; D008958:Models, Molecular; D010088:Oxidoreductases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D010544:Peroxidases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D048372:Streptomyces coelicolor / Q000201:enzymology / Q000235:genetics
D011993:Recombinant Fusion Proteins; D005182:Flavin-Adenine Dinucleotide; D010088:Oxidoreductases; D010544:Peroxidases