Indiana University-Purdue University Indianapolis (IUPUI) / Thousands of missense mutations have been found to be associated with human
diseases, ~60% of which have been predicted to affect protein stability and/or protein-protein
interactions (PPIs). Current proteomic methods for studying the effects of
mutations on the cell focus on measures of protein abundance or post-translational
modifications (PTMs), which cannot directly be used for PPI analysis. High-throughput
methodology to evaluate how mutations in a single protein affect PPI networks would
help streamline the characterization of global effects caused by mutant proteins and aid in
the prediction of phenotypic outcomes resulting from genomic mutations. Temperature
sensitive Mutant Proteome Profiling (TeMPP) is a novel application of a mass
spectrometry (MS) based thermal proteome profiling (TPP) approach that measures
changes in missense mutant containing proteomes without the requirement for large
amounts of starting material, specific antibodies against proteins of interest, and/or
genetic manipulation of the biological system. This study measures the impact of
temperature sensitivity-inducing missense mutations of proteins in the ubiquitin
proteasome system and the transcription termination machinery on the thermal stability
of the proteome at large. Results reveal distinct mechanistic details that were not obtained
using only steady-state transcriptome and proteome analyses. Furthermore, my data
suggests that TeMPP is highly specific to proteins functionally related to the mutated
protein of interest and capable of differentiating effects between two proteins in the same
complex. Overall, TeMPP provides unique mechanistic insights into missense mutation
dysfunction and connection of genotype to phenotype in a rapid, non-biased fashion. Use
of this method along with other complementary -omics approaches will help to
characterize how missense mutations affect cellular protein homeostasis and thus enable
deeper insight into disease phenotypes. / 2022-08-10
| Identifer | oai:union.ndltd.org:IUPUI/oai:scholarworks.iupui.edu:1805/23678 |
| Date | 07 1900 |
| Creators | Justice, Sarah Ann |
| Contributors | Mosley, Amber L., Harrington, Maureen A., Goebl, Mark G., Bidwell, Joseph P. |
| Source Sets | Indiana University-Purdue University Indianapolis |
| Language | en_US |
| Detected Language | English |
| Type | Dissertation |
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