A combined bioinformatics and experimental approach towards the molecular phenotype of disease-causing single point mutations.

Towards unravelling the enigmatic sequence-structure-function relationship, this study presents a computational-experimental strategy to characterise the effects of disease mutations on protein structure stability and misfolding phenotypes. Firstly, suitable candidates from normally secreted human p...

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Main Author: Lim, Radiance Hui Ting.
Other Authors: School of Biological Sciences
Format: Final Year Project
Language:English
Published: 2010
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Online Access:http://hdl.handle.net/10356/40118
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Institution: Nanyang Technological University
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spelling sg-ntu-dr.10356-401182023-02-28T18:06:20Z A combined bioinformatics and experimental approach towards the molecular phenotype of disease-causing single point mutations. Lim, Radiance Hui Ting. School of Biological Sciences A*STAR Bioinformatics Institute Sebastian Maurer-Stroh DRNTU::Science::Biological sciences::Biochemistry Towards unravelling the enigmatic sequence-structure-function relationship, this study presents a computational-experimental strategy to characterise the effects of disease mutations on protein structure stability and misfolding phenotypes. Firstly, suitable candidates from normally secreted human proteins with a disease-associated non-synonymous single nucleotide polymorphism within a domain of known structure were shortlisted through a newly developed bioinformatics workflow. To predict changes in stability upon mutation, a ΔΔG was calculated for each candidate using the FoldX force field. The two selected candidates are defective lysosomal exoglycohydrolases implicated in Fabry disease: α-galactosidase A C202W and S297F. The native and mutated proteins were overexpressed in model cell lines and changes in subcellular distribution and secretion observed. Intriguingly, the C202W mutant was secreted although at a lower level than the native protein while the S297F mutant was fully retained in the cell. Lysosomal targeting and catalytic function were most efficient in the native protein and significantly impaired in the S297F mutant. As additional control, an I91V mutant with minimal ΔΔG was examined and indeed its phenotype and function were close to native. Therefore, this work illustrates the power of a combined bioinformatics and experimental approach towards understanding the molecular mechanisms underpinning protein folding and disease pathogenesis. Bachelor of Science in Biological Sciences 2010-06-10T07:41:46Z 2010-06-10T07:41:46Z 2010 2010 Final Year Project (FYP) http://hdl.handle.net/10356/40118 en Nanyang Technological University 35 p. application/pdf
institution Nanyang Technological University
building NTU Library
continent Asia
country Singapore
Singapore
content_provider NTU Library
collection DR-NTU
language English
topic DRNTU::Science::Biological sciences::Biochemistry
spellingShingle DRNTU::Science::Biological sciences::Biochemistry
Lim, Radiance Hui Ting.
A combined bioinformatics and experimental approach towards the molecular phenotype of disease-causing single point mutations.
description Towards unravelling the enigmatic sequence-structure-function relationship, this study presents a computational-experimental strategy to characterise the effects of disease mutations on protein structure stability and misfolding phenotypes. Firstly, suitable candidates from normally secreted human proteins with a disease-associated non-synonymous single nucleotide polymorphism within a domain of known structure were shortlisted through a newly developed bioinformatics workflow. To predict changes in stability upon mutation, a ΔΔG was calculated for each candidate using the FoldX force field. The two selected candidates are defective lysosomal exoglycohydrolases implicated in Fabry disease: α-galactosidase A C202W and S297F. The native and mutated proteins were overexpressed in model cell lines and changes in subcellular distribution and secretion observed. Intriguingly, the C202W mutant was secreted although at a lower level than the native protein while the S297F mutant was fully retained in the cell. Lysosomal targeting and catalytic function were most efficient in the native protein and significantly impaired in the S297F mutant. As additional control, an I91V mutant with minimal ΔΔG was examined and indeed its phenotype and function were close to native. Therefore, this work illustrates the power of a combined bioinformatics and experimental approach towards understanding the molecular mechanisms underpinning protein folding and disease pathogenesis.
author2 School of Biological Sciences
author_facet School of Biological Sciences
Lim, Radiance Hui Ting.
format Final Year Project
author Lim, Radiance Hui Ting.
author_sort Lim, Radiance Hui Ting.
title A combined bioinformatics and experimental approach towards the molecular phenotype of disease-causing single point mutations.
title_short A combined bioinformatics and experimental approach towards the molecular phenotype of disease-causing single point mutations.
title_full A combined bioinformatics and experimental approach towards the molecular phenotype of disease-causing single point mutations.
title_fullStr A combined bioinformatics and experimental approach towards the molecular phenotype of disease-causing single point mutations.
title_full_unstemmed A combined bioinformatics and experimental approach towards the molecular phenotype of disease-causing single point mutations.
title_sort combined bioinformatics and experimental approach towards the molecular phenotype of disease-causing single point mutations.
publishDate 2010
url http://hdl.handle.net/10356/40118
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