Stepwise building of a 115-kDa macromolecular edifice monitored by electrospray mass spectrometry - The case of acetohydroxy acid isomeroreductase

The macromolecular complexes formed by the enzyme acetohydroxy acid isomero reductase with NADPH. magnesium ions and the competitive inhibitor N-hydrox y-N-isopropyloxamate (IpOHA) were analysed by electrospray mass spectrometr y. Each ligand was added successively to a protein solution, allowing the s toichiometry of the whole macromolecular edifice (115 583 Da) to be unambig uously determined. The combination of an electrospray ion source with the h igh mass range magnetic instrument used in the present studies proved to be a very powerful tool for characterizing, in a specific manner, the quatern ary structures of proteins by single mass measurements.