Issue 13, 2013

An electrospray ms-coupled microfluidic device for sub-second hydrogen/deuterium exchange pulse-labelling reveals allosteric effects in enzyme inhibition

Abstract

In this work, we introduce an integrated, electrospray mass spectrometry-coupled microfluidic chip that supports the complete workflow for ‘bottom up’ hydrogen/deuterium exchange (HDX) pulse labelling experiments. HDX pulse labelling is used to measure structural changes in proteins that occur after the initiation of a reaction, most commonly folding. In the present case, we demonstrate the device on the β-lactamase enzyme TEM-1, identifying active site changes that occur upon acylation by a covalent inhibitor and subtle changes in conformational dynamics that occur away from the active site over a period of several second after the inhibitor is bound. Our results demonstrate the power of microfluidics-enabled sub-second HDX pulse labelling as a tool for studying allostery and show some intriguing correlations with mutagenesis studies.

Graphical abstract: An electrospray ms-coupled microfluidic device for sub-second hydrogen/deuterium exchange pulse-labelling reveals allosteric effects in enzyme inhibition

Supplementary files

Article information

Article type
Paper
Submitted
03 Jan 2013
Accepted
04 Feb 2013
First published
04 Feb 2013

Lab Chip, 2013,13, 2528-2532

An electrospray ms-coupled microfluidic device for sub-second hydrogen/deuterium exchange pulse-labelling reveals allosteric effects in enzyme inhibition

T. Rob, P. K. Gill, D. Golemi-Kotra and D. J. Wilson, Lab Chip, 2013, 13, 2528 DOI: 10.1039/C3LC00007A

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