Investigating protein structure by means of mass spectrometry
Scarff, Charlotte A. (2010) Investigating protein structure by means of mass spectrometry. PhD thesis, University of Warwick.
WRAP_THESIS_Scarff_2010.pdf - Requires a PDF viewer such as GSview, Xpdf or Adobe Acrobat Reader
Official URL: http://webcat.warwick.ac.uk/record=b2485174~S15
The three-dimensional conformation of a protein is central to its biological function.
Mass spectrometry (MS) has become an important tool for the study of various
aspects of protein structure. This project investigates the use of MS for diagnosis of
hemoglobinopathies, through primary structure identification, and for threedimensional
protein structure analysis, through comparison to established methods
and application to protein systems.
Travelling-wave ion mobility mass spectrometry (TWIM-MS) was used to
investigate the biological significance of gas-phase protein structure. Protein
standards were analysed by TWIM-MS. Cross-sections were estimated for proteins
studied, for charge states most indicative of native structure, and were found to be in
good agreement with those calculated from published X-ray crystallography and
nuclear magnetic resonance structures. These results illustrated that the TWIM-MS
approach can provide biologically-relevant data on three-dimensional protein
TWIM-MS was then used to study the structural properties of the hemoglobin
tetramer and its components. Results showed that globin monomers exist in similar
conformations whether in apo- or holo- forms and that a heme-deficient dimer is
unlikely to be a prerequisite for hemoglobin tetramer assembly. TWIM-MS was used
to successfully differentiate between normal and sickle hemoglobin tetramers.
The conformational changes occurring in VanS, a histidine kinase, upon
autophosphorylation were investigated by TWIM-MS. Results provided insights into
the mechanism of autophosphorylation. MS was used to follow the rate of the
autophosphorylation and results obtained compared well with those from an
established method. This demonstrated that MS offers a simple, reproducible
alternative to conventional methods for the study of phosphorylation rates.
MS was used to provide positive identification of a range of hemoglobinopathies
caused by single point mutations. A high-throughput method was used to screen for
hemoglobinopathies in South Asians with and without cardiovascular disease.
Results showed a positive correlation between patients with hemoglobinopathies and
those with cardiovascular disease.
|Item Type:||Thesis or Dissertation (PhD)|
|Subjects:||Q Science > QD Chemistry
Q Science > QP Physiology
|Library of Congress Subject Headings (LCSH):||Proteins -- Structure, Mass spectrometry, Hemoglobinopathy -- Diagnosis|
|Official Date:||July 2010|
|Institution:||University of Warwick|
|Theses Department:||Department of Biological Sciences|
|Supervisor(s)/Advisor:||Scrivens, James H.|
|Sponsors:||Biotechnology and Biological Sciences Research Council (Great Britain) (BBSRC) ; Waters Corporation|
|Extent:||xx, 193 leaves : ill., charts|
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