Browsing by Subject "Farnesyltransferase"

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    Protein prenylation in the pathogenesis of Alzheimer’s Disease and its therapeutic potential
    (2021-06) Jeong, Angela
    With the unprecedented growth of senior population, Alzheimer’s disease (AD) has risen in prevalence to approximately 50 million people afflicted worldwide. While the molecular mechanisms underlying the AD pathogenesis have yet to be elucidated, emerging evidence suggests that a key posttranslational lipid modification of proteins, called prenylation, may play an important role in the pathogenesis of AD. Isoprenoids, including farnesyl pyrophosphate (FPP) and geranylgeranyl pyrophosphate (GGPP), are synthesized from mevalonate in the cholesterol biosynthesis pathway, and the attachment of these lipid prenyl groups is catalyzed by farnesyltransferase (FT) and geranylgeranyl transferases (GGTs), respectively. Once prenylated, proteins localize to cell membranes, where they interact with downstream effectors, and regulate various cellular processes. Multiple lines of evidence have shown that modulation of protein prenylation affects AD neuropathology. To investigate the contribution of each prenylation pathway to AD pathogenesis, our laboratory previously generated the transgenic AD model mice (APP/PS1) with systemic haplodeficiency of FT or GGT. While haplodeficiency of either FT or GGT reduced Aβ deposition and neuroinflammation, only FT haplodeficiency rescued cognitive deficits, suggesting distinct functions of the two prenylation pathways. To follow up on the previous work, my dissertation research is aimed to: 1) investigate the AD pathology-associated changes in prenylation; 2) elucidate the role of neuronal FT in the AD pathogenesis; 3) identify prenylation substrates involved in AD pathogenesis, which can be potentially targeted for therapeutic development. Studies with postmortem human brain tissue samples from two cohorts found that FT levels and the farnesylation of H-Ras were significantly elevated in the brains with AD. In line with this evidence from human samples, neuronal FT deletion attenuated memory impairment and mitigated the Aβ levels by reducing its production as well as normalizing the heightened mTORC1 activity in APP/PS1 mice. Finally, experiments employing a novel prenylomic profiling approach identified proteins whose prenylations were significantly upregulated in APP/PS1 mice compared to WT controls. Taken together, the findings from my dissertation research indicate that aberrant upregulation of protein farnesylation contributes to the pathogenic process of AD, and targeting protein farnesylation and/or downstream signaling pathways of farnesylated proteins may confer therapeutic benefits against AD.
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    Solid-phase synthesis of C-Terminal peptide Libraries for studying the specificity of protein farnesyltransferase
    (2014-07) Wang, Yen-Chih
    Protein prenylation is a common post-translational modification of specific protein-derived cysteine residues in eukaryotic cells. To study the substrate specificity of these enzymes, the primary strategy employed to date has involved the synthesis, purification and assaying of individual peptides. As an improvement, here we describe the synthesis of peptides containing free C-termini on solid supports. The libraries were screened using an alkyne-containing isoprenoid analogue followed by click chemistry with biotin azide and subsequent visualization with streptavidin-AP. Screening of the CVa2X and CCa2X libraries with R. norvegicus PFTase revealed reaction by many known recognition sequences as well as numerous unknown ones. Screening of the CVa2X library with alkyne-functionalized isoprenoid substrates showed that those prepared from C10 or C15 precursors gave similar results while the analogue synthesized from a C5 unit gave a different pattern of reactivity. Finally the substrate specificities of PFTases from three organisms (R. norvegicus, S. cerevisiae and C. albicans) were compared using CVa2X libraries. R. norvegicus PFTase was found to share more peptide substrates with S. cerevisiae PFTase than with C. albicans PFTase. In general, this method is a highly efficient strategy for rapidly probing the specificity of this important enzyme.