The University of Southampton
University of Southampton Institutional Repository

A modular approach for a controlled immobilization of enzymes

A modular approach for a controlled immobilization of enzymes
A modular approach for a controlled immobilization of enzymes
Stable, site-specific immobilization of redox proteins and enzymes is of great 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. For such applications, it would be desirable to immobilise redox proteins and enzymes in a specific orientation on the electrode in order to improve direct electron transfer.
In this work, we describe such an approach using site directed mutagenesis to introduce cysteine residues at specific locations on the enzyme surface, and the reaction between the free thiol of the cysteine and maleimide groups attached on the electrode surface to immobilise the mutated enzymes. Using cellobiose dehydrogenase (CDH) as a model system, different types of electrodes (carbon and gold-based, flat and nanostructured) were firstly modified with maleimide groups using a modular approach based on electrografting and solid-phase synthesis. Therefore, the electrodes were used to covalently immobilise CDH variants bearing the free cysteine in different locations at their surface. The key point of this method is that the main elements of the modification can be independently varied to tune the architecture of the electrode surface as required, by simply changing the “bricks” of the structure.
The CDH-modified electrodes were tested for direct and mediated electron transfer, showing excellent long-term storage stability as well as good catalytic responses. The mechanisms of the direct and mediated electron transfer were fully investigated, as well as the kinetics of the CDH electrodic reactions, also employing computer simulations.
University of Southampton
Meneghello, Marta
0978ae7b-821d-49d9-9dac-a2a8e2306d5b
Meneghello, Marta
0978ae7b-821d-49d9-9dac-a2a8e2306d5b
Bartlett, Philip
d99446db-a59d-4f89-96eb-f64b5d8bb075

Meneghello, Marta (2018) A modular approach for a controlled immobilization of enzymes. University of Southampton, Doctoral Thesis, 273pp.

Record type: Thesis (Doctoral)

Abstract

Stable, site-specific immobilization of redox proteins and enzymes is of great 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. For such applications, it would be desirable to immobilise redox proteins and enzymes in a specific orientation on the electrode in order to improve direct electron transfer.
In this work, we describe such an approach using site directed mutagenesis to introduce cysteine residues at specific locations on the enzyme surface, and the reaction between the free thiol of the cysteine and maleimide groups attached on the electrode surface to immobilise the mutated enzymes. Using cellobiose dehydrogenase (CDH) as a model system, different types of electrodes (carbon and gold-based, flat and nanostructured) were firstly modified with maleimide groups using a modular approach based on electrografting and solid-phase synthesis. Therefore, the electrodes were used to covalently immobilise CDH variants bearing the free cysteine in different locations at their surface. The key point of this method is that the main elements of the modification can be independently varied to tune the architecture of the electrode surface as required, by simply changing the “bricks” of the structure.
The CDH-modified electrodes were tested for direct and mediated electron transfer, showing excellent long-term storage stability as well as good catalytic responses. The mechanisms of the direct and mediated electron transfer were fully investigated, as well as the kinetics of the CDH electrodic reactions, also employing computer simulations.

Text
Thesis Marta Meneghello - Version of Record
Restricted to Repository staff only until 28 June 2021.
Available under License University of Southampton Thesis Licence.

More information

Published date: April 2018

Identifiers

Local EPrints ID: 422233
URI: http://eprints.soton.ac.uk/id/eprint/422233
PURE UUID: 766b5d44-2ebc-41c9-ab68-ba9961b12743
ORCID for Marta Meneghello: ORCID iD orcid.org/0000-0003-0099-1783
ORCID for Philip Bartlett: ORCID iD orcid.org/0000-0002-7300-6900

Catalogue record

Date deposited: 19 Jul 2018 16:30
Last modified: 14 Mar 2019 01:53

Export record

Download statistics

Downloads from ePrints over the past year. Other digital versions may also be available to download e.g. from the publisher's website.

View more statistics

Atom RSS 1.0 RSS 2.0

Contact ePrints Soton: eprints@soton.ac.uk

ePrints Soton supports OAI 2.0 with a base URL of http://eprints.soton.ac.uk/cgi/oai2

This repository has been built using EPrints software, developed at the University of Southampton, but available to everyone to use.

We use cookies to ensure that we give you the best experience on our website. If you continue without changing your settings, we will assume that you are happy to receive cookies on the University of Southampton website.

×