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Issue 7, 2015
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Functional modulation and directed assembly of an enzyme through designed non-natural post-translation modification

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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.

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

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Publication details

The article was received on 16 Dec 2014, accepted on 31 Mar 2015 and first published on 31 Mar 2015


Article type: Edge Article
DOI: 10.1039/C4SC03900A
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Chem. Sci., 2015,6, 3712-3717
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    Functional modulation and directed assembly of an enzyme through designed non-natural post-translation modification

    A. M. Hartley, A. J. Zaki, A. R. McGarrity, C. Robert-Ansart, A. V. Moskalenko, G. F. Jones, M. F. Craciun, S. Russo, M. Elliott, J. E. Macdonald and D. D. Jones, Chem. Sci., 2015, 6, 3712
    DOI: 10.1039/C4SC03900A

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