Table 1.

Immobilization technologies for orientation-controlled enzyme on electrode and their advantages and disadvantages

Orientation methods	Platform	Enzyme (cofactor)	Advantages	Disadvantages	References
Electrostatic adsorption	Reduced graphene oxide-gold nanoparticles composites	Glucose oxidase (FAD)	✓Simple adsorption procedure ✓Preservation of enzyme activity	✓Random orientation ✓Low immobilization stability	Das et al. (2014)
	Carbon nanotubes	Glucose oxidase (FAD)			Liu et al. (2018)
Chemical covalent linking	Functionalized gold nanoparticles on porous graphite	Laccase (multi-copper)	✓Strong bonding strength ✓Increased immobilized enzyme density	✓Random attachment points of the enzyme ✓Unfavorable conformational changes ✓Reduced enzymatic activity	Gutierrez-Sanchez et al. (2012)
	Gold nanoparticles-modified carbon nanotubes	Laccase (multi-copper)			Lalaoui et al. (2016)
	Self-assembly monolayer-modified gold disk electrode	Glucose dehydrogenase (FAD)			Lee et al., 2018a, 2018b
	Maleimide-modified gold electrodes	Cellobiose dehydrogenase (FAD)			Ma et al. (2019)
DNA-directed hybridization	Single-walled carbon nanotubes	Bilirubin oxidase (multi-copper)	✓Site-directed immobilization ✓Diverse experiments possible	✓Increased cofactor-Surface spacing due to long length of DNA ✓Required additional enzyme for electron delivery to main enzyme	Chakraborty et al. (2015)
	DNA origami tiles	Glucose oxidase (FAD)/horseradish peroxidase (Heme)			Fu et al. (2012)
Genetic fusion of inorganic binding peptide	Plane gold surface	Glucose dehydrogenase (FAD)	✓Preservation of enzyme activity ✓Enhanced binding capability ✓Highly controllable surface-binding orientation	✓Difficulty of designing fusion constructs	Lee et al. (2018a, 2018b, 2019), this article