Development of a Microfluidic Quench-Flow Interface Compatible with ESI-FT-ICR-MS for the Characterisation of Enzyme Mechanisms

Research output: Chapter in Book/Report/Conference proceedingConference contribution

Abstract

Pre-steady-state kinetics analysis characterizes the first moments after the initiation of an enzymatic reaction. Within this timeframe, typically milliseconds, enzyme-substrate intermediates are formed and consumed. Detection and characterization of these intermediates offers vital evidence to enzymologists- allowing the calculation of rate constants for individual reaction steps, and the direct inference of mechanistic pathways.

In order to achieve millisecond time resolution pre-steady-state analysis requires rapid mixing techniques. In quench flow systems enzyme and substrate and rapidly mixed, allowed to react for a specified time, before the reaction is stopped by the addition of a quench solvent. We have developed a quench flow interface for ESI-mass spectrometry and demonstrate its utility with FT-ICR-MS for direct analysis of enzyme bound intermediates.
Original languageEnglish
Title of host publication57th ASMS Conference on Mass Spectrometry
Publication statusPublished - 2009

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