Functionalization of Oxidases with Peroxidase Activity Creates Oxiperoxidases: A New Breed of Hybrid Enzyme Capable of Cascade Chemistry | Zendy

Winter Remko T. | Zendy; van den Berg Tomas E. | Zendy; Colpa Dana I. | Zendy; van Bloois Edwin | Zendy; Fraaije Marco W. | Zendy

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Functionalization of Oxidases with Peroxidase Activity Creates Oxiperoxidases: A New Breed of Hybrid Enzyme Capable of Cascade Chemistry

Author(s) -

Winter Remko T.,

van den Berg Tomas E.,

Colpa Dana I.,

van Bloois Edwin,

Fraaije Marco W.

Publication year - 2012

Publication title -

chembiochem

Language(s) - English

Resource type - Journals

SCImago Journal Rank - 1.05

H-Index - 126

eISSN - 1439-7633

pISSN - 1439-4227

DOI - 10.1002/cbic.201100639

Subject(s) - peroxidase , chemistry , biochemistry , phosphofructokinase 2 , protein engineering , flavoprotein , directed evolution , enzyme , heme , thermostability , oxidase test , covalent bond , streptomyces coelicolor , combinatorial chemistry , organic chemistry , gene , mutant

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.

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