Abstract
The MH+ ions of matrix assisted laser desorption ionization time-of-flight (MALDI-TOF) spectra for a series of closely related but otherwise indistinguishable proteins were analyzed for singularity using a distribution free statistic, the Kolmogorov-Smirnov non-parametric statistic, K-S. The approach allows spectra which might otherwise be taken as identical, to be distinguished. Such analysis of the spectra may lead to a greater understanding of the chemistry of the proteins under investigation. The analysis requires only standard instrumentation. A standard data analysis protocol was developed and applied to generate a normalized cumulative distribution function (NCDF) for each spectrum. Differences in the NCDF for two different spectra were calculated and the maximum difference, Δmax compared to critical values of K-S. Values of Δmax exceeding the critical value of K-S are taken as the basis for rejecting the statistical null-hypothesis and assigning statistical significance to the differences in the two spectra. We have shown that this approach allows spectra of 1:1 mixtures of closely related recombinant proteins to be distinguished from either protein alone, and that mixtures of a 45 kDa protein and a labeled version of that protein can be distinguished from the pure material and from one another at the level of about 25%. In addition, we are able to use this approach to characterize the extent to which a synthetic glyococonjugation reaction has proceeded under circumstances of differing reaction times.
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Creighton, T. Structures and Molecular Properties. W. H. Freeman and Co.: New York, 1984, pp 31–34.
Clauser, K., et al. Rapid Mass Spectrometric Peptide Sequencing and Mass Matching for Characterization of Human Melanoma Proteins Isolated by Two-Dimensional PAGE. Proc. Natl. Acad. Sci. U.S.A. 1995, 92, 5072–5076.
Yates, J. R., III, et al. Search of Sequence Databases with Uninterpreted High-Energy Collision-Induced Dossociation Spectra of Peptides. J. Am. Soc. Mass Spectrom. 1996, 7(11), 1089–1098.
Eng, J. K.; McCormack, A. L.; Yates, J. R., III. An Approach to Correlate Tandem Mass Spectral Data of Peptides with Amino Acid Sequences in a Protein Database. J. Am. Soc. Mass Spectrom. 1994, 5(11), 976–989.
Patterson, S. D.; Aebersold, R. Mass Spectrometric Approaches for the Identification of Gel-Separated Proteins. Electrophoresis 1995, 16(10), 1791–1814.
Figeys, D.; Aebersold, R. High Sensitivity Identification of Proteins by Electrospray Ionization Tandem Mass Spectrometry: Initial Comparison Between an Ion Trap Mass Spectrometer and a Triple Quadrupole Mass Spectrometer. Electrophoresis 1997, 18(3–4), 360–368.
Yergey, J. General Approach to Calculating Isotopic Distributions for Mass Spectrometry. Int. J. Mass Spectrom. Ion Processes 1983, 52, 337–349.
Feller, W. On the Kolmogorov-Smirnov Limit Theorems for Empirical Distributions. Annals Math. Stat. 1948, 19, 177–189.
Hollander, M.; Wolfe, D. Nonparametric Statistical Methods. John Wiley & Sons: New York, 1999, pp 178–186.
Beyer, W. CRC Standard Probability and Statistics, Tables, and Formulae. CRC Press: Boca Raton, 1991, p 337.
Zhu, H.; Yalcin, T.; Li, L. Analysis of the Accuracy of Determining Average Molecular Weights of Narrow Polydispersity Polymers by Matrix-Assosted Laser Desorption Ionization Time-of-Flight Mass Spectrometry. J. Am. Soc. Mass Spectrom. 1998, 9, 275–281.
Yefimov, S., et al. Recovery of Sodium Dodecyl Sulfate-Proteins from Gel Electrophoretic Bands in a Single Electroelution Step for Mass Spectrometric Analysis. Analytical Chemistry 2000, 284, 288–295.
Yefimov, S., et al. Stacking of Unlabeled Sodium Dodecyl Sulfate-Proteins with Fluorimetrically Detected Moving Boundary, Electroelution and Mass Spectrometric Identification. Electrophoresis 2001, 22, 999–1003.
Zhang, J., et al. Studies Toward Neoglycoconjugates from the Monosaccharide Determinant of Vibrio cholerae O:1, Serotype Ogawa Using the Diethyl Squarate Reagent. Carbohyd. Res. 1998, 313, 15–20.
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Blank, P.S., Sjomeling, C.M., Backlund, P.S. et al. Use of cumulative distribution functions to characterize mass spectra of intact proteins. J Am Soc Mass Spectrom 13, 40–46 (2002). https://doi.org/10.1016/S1044-0305(01)00338-5
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DOI: https://doi.org/10.1016/S1044-0305(01)00338-5