Review of fructosyl amino acid oxidase engineering research: a glimpse into the future of hemoglobin A1c biosensing

Abstract

Glycated proteins, particularly glycated hemoglobin A1c, are important markers for assessing the effectiveness of diabetes treatment. Convenient and reproducible assay systems based on the enzyme fructosyl amino acid oxidase (FAOD) have become attractive alternatives to conventional detection methods. We review the available FAOD-based assays for measurement of glycated proteins as well as the recent advances and future direction of FAOD research. Future research is expected to lead to the next generation of convenient, simple, and economical sensors for glycated protein, ideally suited for point-of-care treatment and self-monitoring applications.

Figure 1.

The chemical reactions behind the synthesis and FAOD-based sensing of A1C. (A) The nonenzymatic glycation of the N-terminal valine residue of the hemoglobin β subunit, resulting in A1C. (B) Fructosyl-amino-acid-oxidase-catalyzed oxidation of f-^αVal that is released by the proteolytic digestion of A1C.

Figure 2.

Principle behind various experimental f-^αVal sensors using the fructosyl amino acid oxidase from Pichia N1-1 FAOD. The sensor systems are based on (A) hydrogen peroxide detection, (B) mPMS mediator-type electrode, (C) FAOD-peroxidase-ferrocene electrode system, and (D) Prussian-blue-based FAOD electrode. For each sensor, the units of FAOD used, the reported linear correlation and sensitivity, as well as the electrode composition and the applied potential are listed. The reduced and oxidized states of molecules are indicated with “red” and “ox,” respectively.

Figure 3.

Docking models of (A) f-^αVal and (B) f-εLys at the active site of the Pichia N1-1 FAOD. Hydrophobic residues are depicted in black, the FAD cofactor in thin lines, nitrogen atoms in blue, and oxygen atoms in red.