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Tandem mass spectrometry of non-enzymatically glycated peptides and proteins
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Lopez-Clavijo, Andrea F. (2013) Tandem mass spectrometry of non-enzymatically glycated peptides and proteins. PhD thesis, University of Warwick.
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WRAP_THESIS_Lopez-Clavijo_2013.pdf - Submitted Version Download (20Mb) | Preview |
Official URL: http://webcat.warwick.ac.uk/record=b2698508~S1
Abstract
The thesis presents the study of the reaction of glyoxal (ethanedial) with polypeptides. This
reaction is important in the food industry as well as during ageing and diabetes mellitus. To
study this reaction a Fourier transform ion cyclotron resonance mass spectrometer coupled with
electron capture dissociation and collisionally activated dissociation was used. Initially this
reaction was carried out in the neuropeptide Substance P to set up the reaction conditions,
sample preparation, as well as the instrumental parameters in the mass spectrometer.
The results in Substance P revealed two compounds, with mass additions assigned as C2O and
C2H2O2 from glyoxal, were formed. MS/MS results showed that the modification site for both
species could be located at either the arginine residue or at the N-terminus. Thus, in order to
distinguish N-terminus from arginine modification the position of the arginine was varied in
four model peptides. The results indicated that both mass additions C2O, C2H2O2 were located
at the arginine residue. Interestingly, two of those model peptides showed an unusual mass
addition of 21.9843 Da, which was assigned as a new type of glyoxal modification at the arginine
residue showing the addition of two carbon atoms from glyoxal and the loss of two hydrogen
atoms from the peptide (C2-H2), herein referred to as 2-imino-imidazole.
In order to assess the involvement of other residues in the reaction with glyoxal a new set of
experiments in acetylated and non-acetylated undecapeptides were carried out. Unexpectedly,
these experiments revealed that two species with the same mass (16.01092 Da) were being
formed in the non-acetylated peptide. One of the species corresponded to diglycation, where the
results suggest that the glyoxal binding at the lysine residue is crosslinked with the N-terminus.
The second species showing the addition of 116.01092 Da was formed at the arginine residue
forming a species, here called a glyoxal dimer, at the arginine residue. The formation of the
glyoxal dimer species was also observed in the acetylated peptide. Although is clear that
crosslinking between the lysine residue and the N-terminus is not possible in the acetylated
peptide, the results seem to indicate that crosslinking between the amino group of the lysine
and the amide group of glutamine could occur. However, a systematic study varying the position
of the lysine relative to the glutamine residue and also relative to the N-terminus needs to be
addressed in the future in order to determine the extent of the involvement of the N-terminus
and amide group in the glyoxal glycation reaction.
Item Type: | Thesis (PhD) |
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Subjects: | Q Science > QD Chemistry Q Science > QP Physiology |
Library of Congress Subject Headings (LCSH): | Peptides, Tandem mass spectrometry, Proteins, Polypeptides, Glyoxal, Glycosylation, Amino acids |
Official Date: | October 2013 |
Institution: | University of Warwick |
Theses Department: | Department of Chemistry |
Thesis Type: | PhD |
Publication Status: | Unpublished |
Supervisor(s)/Advisor: | O'Connor, Peter B. |
Sponsors: | Engineering and Physical Sciences Research Council (EPSRC) (EP/F03421/1) |
Extent: | xxi, 361 leaves : illustrations. |
Language: | eng |
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