A Flexible Method for the Stable, Covalent Immobilization of Enzymes at Electrode Surfaces

作者：Al Lolage Firas A; Meneghello Marta; Ma Su; Ludwig Roland; Bartlett Philip N^*

来源：ChemElectroChem, 2017, 4(6): 1528-1534.

DOI：10.1002/celc.201700135

• enzyme immobilization
• site-specific immobilization
• maleimide
• solid-phase synthesis
• cellobiose dehydrogenase

Stable, site-specific immobilization of redox proteins and enzymes is of interest for the development of biosensors and biofuel cells, where the long-term stability of enzymatic electrodes as well as the possibility of controlling the orientation of the biomolecules at the electrode surface have a great importance. Ideally, it would be desirable to immobilize redox proteins and enzymes in a specific orientation, but still with some flexibility to optimize reaction kinetics. In this work, we establish such an approach by using site-directed mutagenesis to introduce cysteine residues at specific locations on the protein surface and the reaction between the free thiol group and maleimide groups attached to the electrode surface to immobilize the mutated enzymes. Using cellobiose dehydrogenase (CDH) as a model system, carbon nanotube electrodes were first covalently modified with maleimide groups following a modular approach based on electrografting of primary amines at the carbon surface and solid-phase synthesis methodology to elaborate the surface-modified electrode. The CDH-modified electrodes were tested for direct electron transfer (DET), showing high catalytic currents as well as excellent long-term storage stability. The key advantage of this method is its great flexibility, as the main components of the modification can be independently varied to change the local environment at the electrode surface and a wide range of redox proteins or enzymes can be specifically engineered to present cysteine residues at their surface for oriented immobilization.

• 出版日期2017-6