Novel glucose 6-oxidases
AssigneeCalifornia Institute of Technology
InventorsFrances H. Arnold, Lian-Hong Sun, Ioanna P. Petrounia
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Abstract
Glucose oxidase enzymes are provided, including novel variants of galactose oxidase enzymes. The polynucleotides that encode these novel variants can be expressed in recombinant host cell expression systems. The novel variant oxidase enzymes are capable of oxidizing compounds towards which wild-type galactose oxidase (e.g. D-galactose: oxygen 6-oxidoreductase, GAO; EC 1.1.3.9) has little or no activity. Preferred galactose oxidase variants are those which that have improved capability to oxidize secondary alcohols and/or D-glucose relative to the wild-type enzyme.
Claims35
An isolated enzyme comprising a variant of a parent galactose oxidase,
- the parent having at least 70% sequence identity to SEQ ID NO:2, and
- the variant having a mutation in at least one amino acid residue positioned no more than 15 Å from a metal ion in the catalytic center of the parent, and an improved ability to oxidize D-glucose or a D-glucose derivative as compared to the parent.
An isolated enzyme comprising a variant of a parent galactose oxidase,
- the parent having at least 70% sequence identity to SEQ ID NO:2, and
- the variant having a mutation in at least one amino acid residue positioned no more than 15 Å from a metal ion in the catalytic center of the parent, and an improved ability to oxidize a secondary alcohol-group as compared to the parent.
A method of producing a glucose-6 oxidase comprising the steps of:
- (a) constructing a library of variants of a parent galactose oxidase;
- and
- (b) selecting any variant having improved glucose-6 oxidation activity as compared to the parent galactose oxidase,
- wherein the library comprises variants having different mutations in a first amino acid residue positioned no more than 15 Å from a metal ion in the catalytic center of the parent galactose oxidase.
Dependencies are inferred from the claim text.