Hilton, Gillian R. (2009) Mass spectrometry-based studies of synthetic and natural macromolecules. PhD thesis, University of Warwick.

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Abstract

Originally established as an analytical technique in the fields of physics and
chemistry, mass spectrometry has recently become an essential tool in biological
research. Advances in ionisation methods and novel types of instrumentation have
led to the development of mass spectrometry for the analysis of a wide variety of
biological samples. The work presented here describes the use of mass spectrometry
to characterise a variety of synthetic and natural macromolecules.
Transmissible spongiform encephalopathies (TSEs), also known as prion diseases,
are a class of fatal, infectious neurodegenerative diseases that affect both humans
and animals. Prion proteins are unprecedented infectious pathogens that cause a
group of invariably fatal neurodegenerative diseases by means of an entirely novel
mechanism. Ion mobility mass spectrometry (IM-MS) was used to probe the
conformation of a variety of different prion proteins in the gas-phase. It was shown
that IM-MS could distinguish between two recombinant structures representative of
normal cellular prion protein, PrPC and the pathogenic scrapie form (PrPSc). The
structure of the full-length prion protein was probed by means of IM-MS. A
comparison of the estimated cross-sections of truncated prion protein constructs and
full-length constructs suggested that the N-terminal flexible tail was associated with
the core structure. Metal binding to two different prion protein constructs was
investigated. It was observed that copper coordination to the N-terminal fragment
could induce conformational changes in the octarepeat fragment. These changes
were relatively small and could not be measured in the full-length prion protein. The
data suggested that minor structural changes in the N-terminal could stimulate
endocytosis via a minor, undetected, conformational change in the C-terminal
domain.
IM-MS was used as a high resolution separation technique to distinguish between
mixtures of isobaric synthetic polymers. It was observed that the resolving power of
IM-MS/MS was insufficient to resolve the higher molecular weight oligomers. In
comparison, gel permeation chromatography (GPC)-nuclear magnetic resonance
(NMR) spectroscopy (GPC-NMR) analysis of the same isobaric mixture could not
separate the two components. It was observed that IM-MS was better than GPCNMR
at separating isobaric poly(ethylene glycol) mixtures, especially when taking
speed and sensitivity into account.

Item Type:	Thesis (PhD)
Subjects:	Q Science > QH Natural history > QH301 Biology
Library of Congress Subject Headings (LCSH):	Mass spectrometry -- Research, Prion diseases -- Research, Macromolecules -- Research
Official Date:	December 2009
Dates:	Date Event December 2009 Submitted
Institution:	University of Warwick
Theses Department:	Department of Biological Sciences
Thesis Type:	PhD
Publication Status:	Unpublished
Supervisor(s)/Advisor:	Scrivens, James H. ; Jackson, Tony
Sponsors:	Engineering and Physical Sciences Research Council (Great Britain) (EPSRC) ; AzkoNobel ; Intertek Measurement Science Group (IMSG)
Format of File:	pdf
Extent:	298 leaves : ill., charts
Language:	eng

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