American Chemical Society
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A Snapshot of Enzyme Catalysis Using Electrospray Ionization Mass Spectrometry

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journal contribution
posted on 2003-07-29, 00:00 authored by Zhili Li, Apurba K Sau, Shida Shen, Craig Whitehouse, Timor Baasov, Karen S. Anderson
Insights into the early molecular events involving protein−ligand/substrate interactions such as protein signaling and enzyme catalysis can be obtained by examining these processes on a very short, millisecond time scale. We have used time-resolved electrospray mass spectrometry to delineate the catalytic mechanism of a key enzyme in bacterial lipopolysaccharide biosynthesis, 3-deoxy-d-manno-2-octulosonate-8-phosphate synthase (KDO8PS). Direct real-time monitoring of the catalytic reaction under single enzyme turnover conditions reveals a novel hemiketal phosphate intermediate bound to the enzyme in a noncovalent complex that establishes the reaction pathway. This study illustrates the successful application of mass spectrometry to reveal transient biochemical processes and opens a new time domain that can provide detailed structural information of short-lived protein−ligand complexes.