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Application of MS/MS and Ion Mobility to the Characterization of Secondary and Tertiary Protein Structure

Morrison, Lindsay J
http://rave.ohiolink.edu/etdc/view?acc_num=osu1406193833

Abstract Details

2014, Doctor of Philosophy, Ohio State University, Chemistry.
A growing trend is emerging where traditional methods for structural characterization of biological systems, such as circular dichroism, nuclear magnetic resonance, and X-ray crystallography, are increasingly falling short of complete structural characterization and interface characterization in certain classes of biomolecule. In particular, membrane proteins and intrinsically disordered proteins are not amenable to characterization by these approaches. An overrepresentation of crystallizable structures in public databases has been suspected for some time. One concerning facet of this problem lies in the possibility that some proteins may exist in solution in a number of states, but only one or two of these may crystallize, resulting in the obscuration of certain states and the subsequent loss of functional information via the traditional structure-function relationship. Development of alternative characterization strategies is thus appealing. Mass spectrometry is a rapidly evolving analysis method and is increasingly being applied to biological systems. Owing largely to the small sample quantities required for analysis and relative ease of coupling to LC methods, the method has taken hold in metabolomic and proteomic studies. Recently, however, the development of native electrospray, improved ion transfer optics, and analyzers with high m/z limits has greatly facilitated the study of intact proteins and protein complexes. Dissociation of these molecules by surface induced dissociation has been shown to be extremely useful in characterizing the inter-protein architecture of these complexes. Further developments in dissociation strategies and the general understanding of the fundamental chemistry by which macromolecules dissociate is sought in order to extend these applications to other structural problems. This dissertation focuses on the application of collision induced and surface induced dissociation to the characterization of secondary and tertiary protein structure. Although tremendous study has been dedicated to gas-phase peptide fragmentation, little work has examined the fragmentation and structures of the fragments of specific secondary structures by CID. The work described herein investigates the fragmentation of small peptides and a series of small protein complexes. The fragmentation chemistry of residues with carboxylic and amide side chains are described in Chapter 3, and the unique pathways associated with these residues are again addressed in Chapters 4 and 6. In Chapter 4, the fragmentation pathways at the N-terminal region of the peptide are examined to determine if the N-terminal fragmentation products can be used to characterize the N-terminal regions of proteins, an area which is often hard to characterize due to its inherent flexibility. This concept is extended in Chapter 5, which describes the characterization of the cis/trans isomerization of the amide bond of proline in triply charged bradykinin and variant peptides by use of extremely low energy CID coupled to ion mobility. Finally, the fragmentation of alpha-helices is examined in Chapters 6 and 7, with emphasis on the characterizing the fragment ions in order to assess whether the fragment structure is related to the precursor structure. Throughout these studies, dissociation using very low collision energies is repeatedly shown to provide the most structural information and additional development of tertiary and secondary structural studies is expected to follow this approach.
Vicki Wysocki, PhD (Advisor)
Thomas Magliery, PhD (Committee Member)
Marcos Sotomayor, PhD (Committee Member)
262 p.
Biochemistry; Chemistry
mass spectrometry, protein, structural characterization, ion mobility, chemistry

Recommended Citations

Citations

  • Morrison, L. J. (2014). Application of MS/MS and Ion Mobility to the Characterization of Secondary and Tertiary Protein Structure [Doctoral dissertation, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1406193833

    APA Style (7th edition)

  • Morrison, Lindsay. Application of MS/MS and Ion Mobility to the Characterization of Secondary and Tertiary Protein Structure . 2014. Ohio State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu1406193833.

    MLA Style (8th edition)

  • Morrison, Lindsay. "Application of MS/MS and Ion Mobility to the Characterization of Secondary and Tertiary Protein Structure ." Doctoral dissertation, Ohio State University, 2014. http://rave.ohiolink.edu/etdc/view?acc_num=osu1406193833

    Chicago Manual of Style (17th edition)

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