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Protein mis-folding and human disease

Pal, Mohinder (2010) Protein mis-folding and human disease University of Southampton, School of Biological Sciences, Doctoral Thesis , 279pp.

Record type: Thesis (Doctoral)

Abstract

Serum Amyloid P Component (SAP), a putative molecular chaperone, is a homopentameric
plasma protein of 25kDa subunits. It binds to the amyloid fibrils of
misfolded proteins, which cause amyloidosis in humans. SAP not only stabilizes
amyloid fibrils but also protects them from proteolytic and cell mediated
degradation. SAP has been co-crystallized with three different aminoalkyl
phosphonates that bind at the amyloid recognition site of SAP, and the X-ray
crystal structures were determined at atomic resolution.

A secondary aim of this work was to understand the enhanced amyloidogenic
potential of L55P and V30M transthyretin (TTR) protein. TTR misfolding has been
implicated in number of human diseases such as senile systemic amyloidosis,
familial amyloid polyneuropathy and familial amyloid cardiopathy. TTR protein is a
thyroxine binding protein (14kDa) existing as a tetramer in vivo. L55P and V30M
mutant TTR are the most aggressive and most common mutants, respectively, in
causing FAP. L55P and V30M mutant TTR protein were expressed in E.coli and
purified using anion-exchange chromatography and gel filtration. L55P and V30M
TTR were co-crystallised with MDS84, a compound that has been demonstrated to
stabilize the tetramer in vitro. The X-ray structures of L55P and V30M, TTR mutant
proteins have been determined at 1.5Å and 2.1Å resolution.

In addition, research was carried out on the bacterial protein Burkholderia invasion
protein D (BipD). BipD (33kDa) belongs to type III secretion system of Burkholderia
pseudomallei. It creates a pore in the host cell membrane to help the B.
pseudomallei invasion. This bacterial infection causes melioidosis disease in
humans. To determine the ligand recognition site of BipD, its high-resolution crystal
structure has been determined at 1.5Å. This high resolution BipD structure is more
complete than its previously solved structures and is in the new space group C2.
The BipD structure presented in this research may help to design potential
chemical inhibitors of BipD to prevent bacterial invasion into human body.

The molecular chaperones play an important role in the protein refolding and
assembly. ATJ11 is of 14kDa protein belongs to DnaJ co-chaperone family. The
protein has been expressed in E.coli and purified by affinity and cation exchange
chromatography. The biophysical study of ATJ11 (CD spectrum) has been
conducted showing its predominant ?-helical structure and attempts were made to
crystallise it but no protein crystals have been obtained.

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More information

Published date: July 2010
Organisations: University of Southampton

Identifiers

Local EPrints ID: 167063
URI: http://eprints.soton.ac.uk/id/eprint/167063
PURE UUID: 69455abe-1a4e-4fa7-8fe6-8316b93b0331

Catalogue record

Date deposited: 10 Nov 2010 15:58
Last modified: 18 Jul 2017 12:24

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Contributors

Author: Mohinder Pal
Thesis advisor: Steve P. Wood
Thesis advisor: A.R. Coker

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