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Native mass spectrometry protein structural characterization via surface induced dissociation: instrumentation and applications

Yan, Jing
http://rave.ohiolink.edu/etdc/view?acc_num=osu1500570925461292

Abstract Details

2017, Doctor of Philosophy, Ohio State University, Chemistry.
Protein complexes are essential in cellular processes, and protein complex characterization plays a key role in the understanding of biological processes. Native mass spectrometry has become a useful tool for the study of protein complexes. Surface induced dissociation (SID) has been shown to provide folded noncovalent products of protein complexes, reflecting the native structure of protein complexes. This dissertation focuses on characterizing the structure of protein complexes by native mass spectrometry coupled to an SID device, developing methods to solve structural biology questions. In Chapter 2, correlations between SID fragmentation patterns and the subunit interactions of globular homotetramers with known structure is determined. This work uses ion mobility (IM) and SID in a quadrupole time of flight (Q-TOF) instrument. SID provides insight into the interface of homotetramers studied with the smallest interfaces initially cleaved. Different SID fragmentation patterns of protein complex ions with different conformations further show that SID is capable of monitoring conformational changes of protein complexes. Chapter 3 describes the application of SID to three uncharacterized protein complex systems: a protein oligomer (lambda exonuclease), a DNA-protein complex (nucleosome) and an enzyme-substrate complex (methyltransferase substrates), providing structural information to answer different biological questions. Subunit exchange in the mixture of lambda exonuclease wild-type and mutant is confirmed by using SID to provide the stoichiometry information. Subunit information and nucleosome stability information are gained from the SID of wild-type and H2A.Z variant nucleosomes. Both SID and CID of methyltransferase substrates can identify a substrate from the enzyme-substrate complex in a cellular milieu, providing a native MS method to use in broad screening for enzyme-subtract pairs. In the work described in Chapter 4, conformations of globular protein complex ions generated in gas phase ion-ion reactions and solution additive charge reduction are compared. When the conformation of the precursor ions is disrupted by the supercharging or cone activation, the difference in SID fragmentation patterns of ions generated from gas phase ion-ion reactions can be observed, indicating that the conformation of protein complex ions can be preserved in gas phase ion-ion reactions. Chapter 5 describes the study of protein conformational changes during gas phase trapping in a Q-TOF instrument. No significant conformational changes were observed during the trapping time studied, based on similar SID fragmentation patterns that were observed from streptavidin ions trapped for various amount of time in the gas phase. Chapter 6 describes the implementation of an SID device in a Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometer, which provides ultrahigh resolution m/z measurements, to pursue more detailed structural information on protein complexes in SID. SID of several protein complex systems in the FT-ICR provides similar SID fragmentation patterns to those obtained previously from Q-TOF platforms. Also, the ultrahigh resolution and mass accuracy measurements make it possible to assign modifications and metal adduct to specific protein subunits. In summary, SID can not only provide connectivity, stoichiometry and ligand/adduct binding information on various protein complex systems, but also reveals conformational changes of globular protein complexes in the gas phase.
Vicki Wysocki (Advisor)
Abraham Badu-Tawiah (Committee Member)
Jennifer Ottesen (Committee Member)
249 p.
Chemistry
Native mass spectrometry; surface induced dissociation; protein complexes

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Citations

  • Yan, J. (2017). Native mass spectrometry protein structural characterization via surface induced dissociation: instrumentation and applications [Doctoral dissertation, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1500570925461292

    APA Style (7th edition)

  • Yan, Jing. Native mass spectrometry protein structural characterization via surface induced dissociation: instrumentation and applications. 2017. Ohio State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu1500570925461292.

    MLA Style (8th edition)

  • Yan, Jing. "Native mass spectrometry protein structural characterization via surface induced dissociation: instrumentation and applications." Doctoral dissertation, Ohio State University, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=osu1500570925461292

    Chicago Manual of Style (17th edition)

osu1500570925461292
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This open access ETD is published by The Ohio State University and OhioLINK.