Site-specific introduction of an electroactive label into a non-electroactive enzyme (β-lactamase I)

Abstract

A cysteine residue was introduced close to the active site of β-lactamase I by site-directed mutagenesis to replace tyrosine-105 and was subsequently modified with an electroactive SH-specific reagent, N-(2-ferrocene-ethyl)maleimide. The resulting modified enzyme became electroactive, showing good quasi-reversible electrochemistry which was characteristic of the attached ferrocene moiety while retaining its specific enzymatic activity. In the presence of a suicide substrate, 6β-iodopenicillanic acid, the redox potential shifted +20 mV suggesting that the label was sensitive to changes in the active site of the enzyme.