Spectroscopic Observation of Calcium‐Induced Reorientation of Cellobiose Dehydrogenase Immobilized on Electrodes and its Effect on Electrocatalytic Activity

Kielb, Patrycja; Sezer, Murat; Katz, Stacy; López, Francisco Salinas; Schulz, Christopher; Gorton, Lo; Ludwig, Roland; Wollenberger, Ulla; Zebger, Ingo; Weidinger, Inez M.

doi:10.1002/cphc.201500112

Cited by 36 publications

(25 citation statements)

References 79 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The effect of divalent cations on some CDHs was first reported by C. Schulz, who observed an enhanced catalytic current of CDH in the presence of calcium ions, which points towards a limiting IET rate. Weidinger found a calcium‐induced reorientation of CDH on electrodes employing surface‐enhanced vibrational spectroscopy . The reported DET rate of 4.6 s −1 is 35 times higher than IET rate measured by stopped‐flow spectroscopy (Table ), which confirms that IET is the overall rate‐limiting step in M. thermophilum CDH.…”

Section: Direct Electrochemistry Of Flavocytochromesmentioning

confidence: 99%

Direct Electron Transfer of Enzymes Facilitated by Cytochromes

Ludwig

2018

ChemElectroChem

Self Cite

View full text Add to dashboard Cite

The direct electron transfer (DET) of enzymes has been utilized to develop biosensors and enzymatic biofuel cells on micro‐ and nanostructured electrodes. Whereas some enzymes exhibit direct electron transfer between their active‐site cofactor and an electrode, other oxidoreductases depend on acquired cytochrome domains or cytochrome subunits as built‐in redox mediators. The physiological function of these cytochromes is to transfer electrons between the active‐site cofactor and a redox partner protein. The exchange of the natural electron acceptor/donor by an electrode has been demonstrated for several cytochrome carrying oxidoreductases. These multi‐cofactor enzymes have been applied in third generation biosensors to detect glucose, lactate, and other analytes. This review investigates and classifies oxidoreductases with a cytochrome domain, enzyme complexes with a cytochrome subunit, and covers designed cytochrome fusion enzymes. The structurally and electrochemically best characterized proponents from each enzyme class carrying a cytochrome, that is, flavoenzymes, quinoenzymes, molybdenum‐cofactor enzymes, iron‐sulfur cluster enzymes, and multi‐haem enzymes, are featured, and their biochemical, kinetic, and electrochemical properties are compared. The cytochromes molecular and functional properties as well as their contribution to the interdomain electron transfer (IET, between active‐site and cytochrome) and DET (between cytochrome and electrode) with regard to the achieved current density is discussed. Protein design strategies for cytochrome‐fused enzymes are reviewed and the limiting factors as well as strategies to overcome them are outlined.

show abstract

Section: Direct Electrochemistry Of Flavocytochromesmentioning

confidence: 99%

Direct Electron Transfer of Enzymes Facilitated by Cytochromes

Ludwig

2018

ChemElectroChem

Self Cite

View full text Add to dashboard Cite

show abstract

“…As biological recognition element for lactose we chose CDH, since this enzyme shows a high activity for the oxidation of lactose while it is insensitive to O 2 . Moreover, it can be effectively coupled to electrode surfaces in a direct electron transfer (DET) regime . The top layer of the proposed sensor removes only the interfering analyte glucose whereas lactose can diffuse to the CDH layer.…”

Section: Resultsmentioning

confidence: 63%

“…A monolayer of CDH was immobilized onto graphite electrodes by means of electrostatic grafting of the enzyme using PDADMAC. This approach was used earlier for the fabrication of CDH modified electrodes and showed a good current response in the presence of lactose . The enzyme is in a DET regime and an applied potential of 0 V (vs. Ag/AgCl/3 M KCl) is sufficient for an efficient electron transfer between the heme domain within the enzyme and the electrode surface.…”

Section: Resultsmentioning

confidence: 97%

See 1 more Smart Citation

A Polymer Multilayer Based Amperometric Biosensor for the Detection of Lactose in the Presence of High Concentrations of Glucose

López

Ludwig

et al. 2016

Electroanalysis

Self Cite

View full text Add to dashboard Cite

We report on the development of an amperometric polymer multilayer‐based biosensor for the determination of lactose in the presence of high concentrations of glucose. The sensor platform consists of a two‐layer system with the first layer (sensing layer) comprising cellobiose dehydrogenase (CDH) bound by electrostatic interactions in a favourite orientation for efficient direct electron transfer. The second layer on top of the sensing element consists of a specifically designed hydrophilic polymer which co‐entraps glucose oxidase (GOx) and catalase (CAT). This bi‐enzymatic system is able to remove glucose (up to a concentration of at least 140 mM) in the outer layer of the modified electrode and thus prevents the interfering analyte from reaching the active CDH layer. Moreover, the concomitantly generated H2O2 is efficiently removed by means of CAT. The linear range for the detection of lactose was from 10 to 100 μM. The sensor is able to detect lactose at low concentrations in lactose‐free milk samples. Sample pre‐treatment consist of a simple dilution step. Quantification of the lactose content in lactose‐free dairy products showed that the amount of lactose is below the threshold given by the manufacturer.

show abstract

“…[21,23] Clear evidence that the catalytic current is due to the oxidation of the glucose by the immobilized enzyme is provided by the control experiment using L-glucose, Figure 3, where upon addition of aliquots to increase the concentration of L-glucose in the cell there is no change in the current (compare each pair of dotted and solid curves), whereas upon each addition of an aliquot of D-glucose to increase its concentration in the cell the current increases (compare the solid curves). [21,23] Clear evidence that the catalytic current is due to the oxidation of the glucose by the immobilized enzyme is provided by the control experiment using L-glucose, Figure 3, where upon addition of aliquots to increase the concentration of L-glucose in the cell there is no change in the current (compare each pair of dotted and solid curves), whereas upon each addition of an aliquot of D-glucose to increase its concentration in the cell the current increases (compare the solid curves).…”

Section: Direct Electron Transfer At Mtcdh-modified Electrodesmentioning

confidence: 99%

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

Al‐Lolage

Meneghello

et al. 2017

ChemElectroChem

Self Cite

View full text Add to dashboard Cite

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 dehydrogen-ase (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.

show abstract

Spectroscopic Observation of Calcium‐Induced Reorientation of Cellobiose Dehydrogenase Immobilized on Electrodes and its Effect on Electrocatalytic Activity

Cited by 36 publications

References 79 publications

Direct Electron Transfer of Enzymes Facilitated by Cytochromes

Direct Electron Transfer of Enzymes Facilitated by Cytochromes

A Polymer Multilayer Based Amperometric Biosensor for the Detection of Lactose in the Presence of High Concentrations of Glucose

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

Contact Info

Product

Resources

About