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Functional modulation and directed assembly of an enzyme through designed non-natural post-translation modification

Hartley, Andrew M., Zaki, Athraa J., McGarrity, Adam R., Robert-Ansart, Cecile, Moskalenko, Andriy V., Jones, Gareth F., Craciun, Monica F., Russo, Saverio, Elliott, Martin, MacDonald, John Emyr and Jones, Darran Daffydd 2015. Functional modulation and directed assembly of an enzyme through designed non-natural post-translation modification. Chemical Science 6 (7) , pp. 3712-3717. 10.1039/C4SC03900A

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Abstract

Post-translational modification (PTM) modulates and supplements protein functionality. In nature this high precision event requires specific motifs and/or associated modification machinery. To overcome the inherent complexity that hinders PTM's wider use, we have utilized a non-native biocompatible Click chemistry approach to site-specifically modify TEM β-lactamase that adds new functionality. In silico modelling was used to design TEM β-lactamase variants with the non-natural amino acid p-azido-L-phenylalanine (azF) placed at functionally strategic positions permitting residue-specific modification with alkyne adducts by exploiting strain-promoted azide–alkyne cycloaddition. Three designs were implemented so that the modification would: (i) inhibit TEM activity (Y105azF); (ii) restore activity compromised by the initial mutation (P174azF); (iii) facilitate assembly on pristine graphene (W165azF). A dibenzylcyclooctyne (DBCO) with amine functionality was enough to modulate enzymatic activity. Modification of TEMW165azF with a DBCO–pyrene adduct had little effect on activity despite the modification site being close to a key catalytic residue but allowed directed assembly of the enzyme on graphene, potentially facilitating the construction of protein-gated carbon transistor systems

Item Type: Article
Date Type: Publication
Status: Published
Schools: Biosciences
Physics and Astronomy
Subjects: Q Science > QC Physics
Q Science > QH Natural history > QH301 Biology
Additional Information: First published online 31 Mar 2015 This article is licensed under a Creative Commons Attribution 3.0 Unported Licence.
Publisher: Royal Society of Chemistry
ISSN: 2041-6520
Funders: EPSRC, BBSRC
Date of First Compliant Deposit: 30 March 2016
Date of Acceptance: 31 March 2015
Last Modified: 22 Mar 2019 21:41
URI: http://orca.cf.ac.uk/id/eprint/81057

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Cited 8 times in Scopus. View in Scopus. Powered By Scopus® Data

Cited 1 time in Web of Science. View in Web of Science.

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