A Snapshot of Enzyme Catalysis Using Electrospray Ionization Mass Spectrometry
journal contributionposted on 29.07.2003, 00:00 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.