Hydrogen / Deuterium Exchange and Fragmentation of Biomolecules to Probe Gas Phase Structure and Energetics

Persistent Link:
http://hdl.handle.net/10150/196052
Title:
Hydrogen / Deuterium Exchange and Fragmentation of Biomolecules to Probe Gas Phase Structure and Energetics
Author:
Herrmann, Kristin Ann
Issue Date:
2005
Publisher:
The University of Arizona.
Rights:
Copyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.
Abstract:
Presented in this dissertation are FT-ICR H/D exchange and fragmentation studies of protonated peptides for the purpose of better understanding gas phase conformation and protonation motifs, and their affect on fragmentation patterns. In addition, a new ion activation method is developed to enhance the abundance of higher energy fragmentation pathways, thereby providing additional structural and/or mechanistic information.Studies to probe the relay mechanism of H/D exchange in small, model peptides found that residue position, proline configuration, and availability of the termini have a measurable effect on the H/D exchange behavior reflective of different conformation / protonation motifs.Charge remote cleavage C-terminal to aspartic acid was studied with the fixed charge derivative tris(2,4,6-trimethoxyphenyl) phosphonium (tTMP-P+). Ab initio calculations demonstrate that tTMP-P+ cannot activate the proposed aspartic acid nucleophile, the acidic side chain hydrogen initiates cleavage. Despite the absence of an ionizing proton, the derivatized peptide P+LDIFSDF (where P+ = tTMP-P+) exchanges three acidic hydrogens for deuterium. This supports the proposed aspartic acid cleavage mechanism that occurs without the direct involvement of an ionizing proton because acidic protons are able to participate in H/D exchange in the absence of an added proton. In addition, H/D exchange of P+LDIFSDF analogues provides insight into the sites and mechanisms of H/D exchange.H/D exchange to separate distinct peptide populations with differing rates and their subsequent fragmentation demonstrate that conformation / protonation motif affects the fragmentation spectrum observed. Studies of [P+LDIFSDF + H]2+, [RPPGFSPFR + 2H]2+ (bradykinin), and [RVYIFPF + 2H]2+ show that at least two distinct structures exist with different rates of H/D exchange and different fragmentation patterns. The overall MS/MS spectrum is a linear combination of all conformations and protonation motifs. In addition, fragmentation of labeled populations suggests that complementary bn+ / ym-n+ ion pairs are being formed from doubly-charged precursor by the same mechanism.SORI-RE CID is a new FT-ICR ion activation method based on the combination of sustained off-resonance irradiation (SORI) and on-resonant excitation (RE) with the purpose of enhancing higher energy fragmentation while maintaining low energy processes. The experiments presented serve to illustrate the usefulness of SORI-RE in diverse cases.
Type:
text; Electronic Dissertation
Keywords:
peptide fragmentation; mass spectrometry; mechanisms; Hydrogen / Deuterium Exchange; FT-ICR; SORI-RE
Degree Name:
Ph.D.
Degree Level:
doctoral
Degree Program:
Chemistry; Graduate College
Degree Grantor:
University of Arizona
Committee Chair:
Wysocki, Vicki H.

Full metadata record

DC FieldValue Language
dc.language.isoenen_US
dc.titleHydrogen / Deuterium Exchange and Fragmentation of Biomolecules to Probe Gas Phase Structure and Energeticsen_US
dc.creatorHerrmann, Kristin Annen_US
dc.contributor.authorHerrmann, Kristin Annen_US
dc.date.issued2005en_US
dc.publisherThe University of Arizona.en_US
dc.rightsCopyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.en_US
dc.description.abstractPresented in this dissertation are FT-ICR H/D exchange and fragmentation studies of protonated peptides for the purpose of better understanding gas phase conformation and protonation motifs, and their affect on fragmentation patterns. In addition, a new ion activation method is developed to enhance the abundance of higher energy fragmentation pathways, thereby providing additional structural and/or mechanistic information.Studies to probe the relay mechanism of H/D exchange in small, model peptides found that residue position, proline configuration, and availability of the termini have a measurable effect on the H/D exchange behavior reflective of different conformation / protonation motifs.Charge remote cleavage C-terminal to aspartic acid was studied with the fixed charge derivative tris(2,4,6-trimethoxyphenyl) phosphonium (tTMP-P+). Ab initio calculations demonstrate that tTMP-P+ cannot activate the proposed aspartic acid nucleophile, the acidic side chain hydrogen initiates cleavage. Despite the absence of an ionizing proton, the derivatized peptide P+LDIFSDF (where P+ = tTMP-P+) exchanges three acidic hydrogens for deuterium. This supports the proposed aspartic acid cleavage mechanism that occurs without the direct involvement of an ionizing proton because acidic protons are able to participate in H/D exchange in the absence of an added proton. In addition, H/D exchange of P+LDIFSDF analogues provides insight into the sites and mechanisms of H/D exchange.H/D exchange to separate distinct peptide populations with differing rates and their subsequent fragmentation demonstrate that conformation / protonation motif affects the fragmentation spectrum observed. Studies of [P+LDIFSDF + H]2+, [RPPGFSPFR + 2H]2+ (bradykinin), and [RVYIFPF + 2H]2+ show that at least two distinct structures exist with different rates of H/D exchange and different fragmentation patterns. The overall MS/MS spectrum is a linear combination of all conformations and protonation motifs. In addition, fragmentation of labeled populations suggests that complementary bn+ / ym-n+ ion pairs are being formed from doubly-charged precursor by the same mechanism.SORI-RE CID is a new FT-ICR ion activation method based on the combination of sustained off-resonance irradiation (SORI) and on-resonant excitation (RE) with the purpose of enhancing higher energy fragmentation while maintaining low energy processes. The experiments presented serve to illustrate the usefulness of SORI-RE in diverse cases.en_US
dc.typetexten_US
dc.typeElectronic Dissertationen_US
dc.subjectpeptide fragmentationen_US
dc.subjectmass spectrometryen_US
dc.subjectmechanismsen_US
dc.subjectHydrogen / Deuterium Exchangeen_US
dc.subjectFT-ICRen_US
dc.subjectSORI-REen_US
thesis.degree.namePh.D.en_US
thesis.degree.leveldoctoralen_US
thesis.degree.disciplineChemistryen_US
thesis.degree.disciplineGraduate Collegeen_US
thesis.degree.grantorUniversity of Arizonaen_US
dc.contributor.chairWysocki, Vicki H.en_US
dc.contributor.committeememberWysocki, Vicki H.en_US
dc.contributor.committeememberAspinwall, Craig A.en_US
dc.contributor.committeememberGhosh, Indraneelen_US
dc.contributor.committeememberMcGrath, Dominicen_US
dc.contributor.committeememberSaavedra, Scotten_US
dc.identifier.proquest1397en_US
dc.identifier.oclc137355436en_US
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