news Probing radical-induced modifications of proteins lysozyme, only the tryptophans at positions 62 and 123 were oxidized, Because reactive ion radicals are thought to contribute to cancer, carwhich is consistent with the solvent accessibilities determined by NMR. diovascular diseases, and Alzheimer’s, there is considerable interest in On the basis of these preliminary results, the researchers suggest that understanding their activities at the molecular level. It is possible to this technique could be used to study how reactive oxygen species generate oxygen-containing radicals by g-radiolysis, X-ray radiolysis, interact with specific protein domains to cause disease. (Rapid Comand other methods, but Simin D. Maleknia and colleagues at the Albert mun. Mass Spectrom. 1999, 13, 2352–2358). Einstein College of Medicine have a new approach: using a reactive nebulizer gas and a high-voltage discharge to produce these radicals within an electrospray ionization (ESI) source. The researchers say the approach is similar to atmospheric pressure chemical ionization, in which ionic species are generated in the presence of a nitrogen nebulizer gas. However, because the new method uses a reactive nebulizer gas within an ESI source, it retains “soft” ionization characteristics, making it suitable for protein analysis. Maleknia and colleagues also say the method is effective at inducing oxidative modification of proteins. In studies of ubiquitin, >50% of the protein was modified. Similar results were achieved for a larger protein, hen lysozyme (reported elsewhere as having 129 residues). Most important, the ESI modification method combined with MS/MS allowed the researchers to correlate the sites of oxidation with the solvent accessibiliESI-MS/MS spectrum of (a) unoxidized and (b) dioxidized tryptic peptide 117–125 of ties of side chains. For example, in hen lysozyme. (Adapted with permission. Copyright 1999 John Wiley & Sons)
Well-characterized FTICR MS When it comes to exact mass measurements, nothing beats Fourier transform ion cyclotron resonance (FTICR) and magnetic sector MS. But despite the proven track record of FTICR MS for high accuracy and reliability, U.S. Food and Drug Administration regulations require all mass spectrometers to be fully characterized via a standardized protocol, to define the average statistical deviation in error, before they can be used for exact mass measurements by those laboratories that must operate under Good Laboratory Practices (GLP). In an effort to characterize a 7-T FTICR mass spectrometer for use in a GLP-regulated pharmaceutical environment, Richard D. Burton and co-workers at Abbott Laboratories, the Centers for Disease Control and Prevention, and the University of Florida performed multiple exact mass measurements for more than 40 compounds. In their systematic study, the researchers used two different internal calibration methods, as well as external calibration with a single internal mass standard. In one internal calibration method, both electrospray ionization (ESI) and MALDI were used simultaneously—ESI for the introduction of analyte ions and MALDI for the formation of polymethylmethacrylate (PMMA) calibrant ions-for higher throughput. In this case, an average absolute mass determination error of 0.8 ppm (SD = 1.0 ppm) was obtained. In comparison, for internal calibration with a co-dissolved polyethylene glycol standard, an average absolute error of 0.7 ppm (SD = 0.9 ppm) was obtained. Last, when exact mass measurements of MS/MS fragment ions were made using external calibration with PMMA as the internal mass standard, an average absolute error of 0.7 ppm (SD = 1.0 ppm) was obtained. The authors admit the samples used in this study were relatively pure. They warn that lower overall mass accuracy and/or precision may result with less pure samples. (J. Am. Soc. Mass Spectrom. 1999, 10, 1291–1297)
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