GLYCOSYLATED MODIFIED FLAVIN ADENINE DINUCLEOTIDE-DEPENDENT GLUCOSE DEHYDROGENASES, COMPOSITIONS THEREOF AS WELL AS METHODS OF MAKING AND USING THE SAME

摘要

Compositions, devices, kits and methods are disclosed for assaying glucose with a glycosylated, modified flavin adenine dinucleotide-dependent glucose dehydrogenase (FAD-GDH), variant thereof or an active fragment thereof, where at least one asparagine residue at positions N2, N168 and N346 of mature, wild-type A. oryzae FAD-GDH according to SEQ ID NO:2 is substituted by one or more amino acids not suitable for glycosylation, thereby eliminating or inactivating, respectively, a potential glycosylation site at this position.

主权项

1. A glycosylated, modified flavin adenine dinucleotide-dependent glucose dehydrogenase selected from the group consisting of:
(a). a glycosylated, modified flavin adenine dinucleotide-dependent glucose dehydrogenase having at least one asparagine residue selected from the group consisting of N2, N168 and N346 of mature, wild-type Aspergillus oryzae flavin adenine dinucleotide-dependent glucose dehydrogenase according to SEQ ID NO:2 substituted by one or more amino acids not suitable for glycosylation, thereby eliminating or inactivating a potential glycosylation site at the at least one asparagine residue; (b). a glycosylated, modified flavin adenine dinucleotide-dependent glucose dehydrogenase having around 80% amino acid sequence identity or more to the glycosylated, modified flavin adenine dinucleotide-dependent glucose dehydrogenase according to (a); and (c). an active fragment of the glycosylated, modified flavin adenine dinucleotide-dependent glucose dehydrogenase according to (a) or (b), wherein in the glycosylated, modified flavin adenine dinucleotide-dependent glucose dehydrogenase according to (b) or the fragment according to (c), the at least one asparagine substitution eliminating or inactivating the potential glycosylation site(s) is preserved when compared to the glycosylated, modified flavin adenine dinucleotide-dependent glucose dehydrogenase according to (a), and wherein the glycosylated, modified flavin adenine dinucleotide-dependent glucose dehydrogenase according to (b) or the fragment according to (c) exhibits at least about 80% of an enzyme activity of the glycosylated, modified flavin adenine dinucleotide-dependent glucose dehydrogenase according to (a) and exhibits at least about 80% of a temperature stability under dry conditions of the glycosylated, modified flavin adenine dinucleotide-dependent glucose dehydrogenase according to (a).