Tandem mass spectrometry of biomolecules: applications and new methodsPublic Deposited
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MLAEnyenihi, Atim Atte. Tandem Mass Spectrometry of Biomolecules: Applications and New Methods. Chapel Hill, NC: University of North Carolina at Chapel Hill, 2010. https://doi.org/10.17615/qzr4-pv73
APAEnyenihi, A. (2010). Tandem mass spectrometry of biomolecules: applications and new methods. Chapel Hill, NC: University of North Carolina at Chapel Hill. https://doi.org/10.17615/qzr4-pv73
ChicagoEnyenihi, Atim Atte. 2010. Tandem Mass Spectrometry of Biomolecules: Applications and New Methods. Chapel Hill, NC: University of North Carolina at Chapel Hill. https://doi.org/10.17615/qzr4-pv73
- Last Modified
- March 21, 2019
Enyenihi, Atim Atte
- Affiliation: College of Arts and Sciences, Department of Chemistry
- In the work presented here, new methods are developed for quadrupole ion trap (QIT) tandem mass spectrometry (MS/MS) of proteins and peptides and existing MS/MS methods are applied to real world problems. MS/MS is an indispensible tool for structural characterization of molecules. In the most common MS/MS experiment, an ion of interest, the parent ion is isolated. The mass selected ion is then activated so that the ion dissociates to form product ions which are subsequently mass analyzed. Although a variety of ion activation and dissociation methods exist, most often provide limited sequence information for proteins and peptides. A new method called electron ionization dissociation (EID) has been developed. It involves reaction of singly and multiply charged analytes with high energy electrons. EID provides increased chemical information as compared to the existing methods. In proteomics, the large scale study of proteins and peptides, quantification of expressed proteins is an important factor. Stable isotope labeling is a common approach to quantitative MS proteomics. Isobaric tag for relative and absolute quantification (iTRAQ) is one of the most common stable isotope methods and is based on the covalent linkage of primary amines with tags of varying masses. Quantitative information using iTRAQ is obtained from the abundance of low mass reporter ions, m/z 114-117 for a 4-plex reagent, observed in MS/MS spectra. CID is the most commonly used ion activation method in MS/MS. However, conventional CID in a quadrupole ion trap mass spectrometer (QITMS) cannot trap product ions below a low-mass cut-off (LMCO) of ~ 25% of the parent ion mass-to-charge, thus iTRAQ reporter ions are not detected. This represents a significant limitation because the QITMS is the most commonly used instrument for proteomics research. Part of the work presented here is to develop methods for iTRAQ quantification using a QITMS. A part of the work presented here involves application of existing MS/MS methods to real world problems. Recombinant protein technology allows industries to produce large quantities of proteins with specific characteristics for a variety of applications. Edman degradation, the primary method used to validate the integrity of recombinantly expressed proteins, is time consuming. Here, electron capture dissociation (ECD) MS/MS was explored as an alternative approach to Edman degradation for high throughput characterization of recombinant proteins. Finally, collision induced dissociation (CID) MS/MS was used to identify class I major histocompatibility complex (MHC) presented peptides on leukemia for use as anti-cancer immune targets.
- Date of publication
- August 2010
- Resource type
- Rights statement
- In Copyright
- "... in partial fulfillment of the requirements for the degree of Doctor of Philosophy in the Department Chemistry."
- Glish, Gary
- Degree granting institution
- University of North Carolina at Chapel Hill
- Place of publication
- Chapel Hill, NC
- Access right
- Open access
- Date uploaded
- March 18, 2013
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