The long-term goal of the research proposed here is to improve the throughput and accuracy of automated tandem mass spectrometric peptide and protein sequencing. There are three specific aims in the proposal: (1) Continue investigations of enhanced cleavage at acidic amino acidic residues in order to confirm structural features that lead to enhanced cleavage. Use ion mobility (ion chromatography) spectrometry and molecular modeling to confirm whether proposed secondary structure leads to unusual fragmentation. (2) Use the SEQUEST program and known proteins to search for structural features that cause unusual fragmentation patterns. SEQUEST is a program that is generally used to correlate uninterpreted product ion MS/MS spectra with sequences from protein and nucleotide databases. In our study, the SEQUEST program will be used as a tool to identify sequence stretches that give spectra that are not correctly predicted based on the current state of knowledge of peptide dissociation. Perform detailed mechanistic studies to clarify/identify the structural features leading to the unusual fragmentation patterns and develop """"""""rules"""""""" that can be made available for incorporation into automated sequencing protocols. (3) Compare the gas-phase collision- induced dissociation and surface-induced dissociation activation methods to determine whether surface-induced dissociation provides a quantifiable improvement in sequence determination by tandem mass spectrometry. Determine whether sequence identification can be obtained by surface-induced dissociation of small proteins without prior digestion, using chemokines as representative structures. These different, but related, projects are expected to increase our fundamental knowledge of peptide dissociation in the gas phase, improve practical automated sequencing of peptides, and characterize whether the surface-induced dissociation activation method should be pursued as a versatile method for practical sequencing of peptides and proteins.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM051387-06
Application #
6164794
Study Section
Special Emphasis Panel (ZRG1-SSS-6 (01))
Program Officer
Edmonds, Charles G
Project Start
1994-08-01
Project End
2003-02-28
Budget Start
2000-03-01
Budget End
2001-02-28
Support Year
6
Fiscal Year
2000
Total Cost
$220,765
Indirect Cost
Name
University of Arizona
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
City
Tucson
State
AZ
Country
United States
Zip Code
85721
Morrison, L J; Chamot-Rooke, J; Wysocki, V H (2014) IR action spectroscopy shows competitive oxazolone and diketopiperazine formation in peptides depends on peptide length and identity of terminal residue in the departing fragment. Analyst 139:2137-43
Gucinski, Ashley C; Chamot-Rooke, Julia; Steinmetz, Vincent et al. (2013) Influence of N-terminal residue composition on the structure of proline-containing b2+ ions. J Phys Chem A 117:1291-8
Basha, Eman; Jones, Christopher; Blackwell, Anne E et al. (2013) An unusual dimeric small heat shock protein provides insight into the mechanism of this class of chaperones. J Mol Biol 425:1683-96
Fritz, Bradley G; Roberts, Sue A; Ahmed, Aqeel et al. (2013) Molecular model of a soluble guanylyl cyclase fragment determined by small-angle X-ray scattering and chemical cross-linking. Biochemistry 52:1568-82
Bernier, Matthew C; Paizs, Bela; Wysocki, Vicki H (2012) Influence of a Gamma Amino Acid on the Structures and Reactivity of Peptide a(3) Ions. Int J Mass Spectrom 316-318:259-267
Gucinski, Ashley C; Chamot-Rooke, Julia; Nicol, Edith et al. (2012) Structural influences on preferential oxazolone versus diketopiperazine b(2+) ion formation for histidine analogue-containing peptides. J Phys Chem A 116:4296-304
Morrison, Lindsay; Somogyi, Arpad; Wysocki, Vicki H (2012) The influence glutamic acid in protonated b3 ýýý b2 formation from VGEIG and related analogs. Int J Mass Spectrom 325-327:139-149
Li, Wenzhou; Wysocki, Vicki H (2012) ETD fragmentation features improve algorithm. Expert Rev Proteomics 9:241-3
Li, Wenzhou; O'Neill, Heather A; Wysocki, Vicki H (2012) SQID-XLink: implementation of an intensity-incorporated algorithm for cross-linked peptide identification. Bioinformatics 28:2548-50
Li, Wenzhou; Song, Chi; Bailey, Derek J et al. (2011) Statistical analysis of electron transfer dissociation pairwise fragmentation patterns. Anal Chem 83:9540-5

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