eprintid: 24223
rev_number: 15
eprint_status: archive
userid: 3674
dir: disk0/00/02/42/23
datestamp: 2018-03-27 09:16:23
lastmod: 2019-04-24 08:52:23
status_changed: 2018-03-27 09:16:23
type: doctoralThesis
metadata_visibility: show
creators_name: Zedan, Mostafa
title: Analysis of the role of N-terminal acetylation of newly synthesized proteins in Saccharomyces cerevisiae
subjects: ddc-500
divisions: i-140001
adv_faculty: af-14
cterms_swd: Saccharomyces cerevisiae
cterms_swd: Acetylation
cterms_swd: protein synthesis
abstract: N-terminal acetylation is a conserved co-translational protein modification that is highly abundant among eukaryotes. In Saccharomyces cerevisiae, at least five enzymes with distinct substrate specificities (N-terminal acetyl transferase Nat A to E) act to acetylate 50–70% of the yeast proteins. Despite being one of the most common protein modifications, its biological significance remains largely ambiguous. I set out to study the role of N-terminal acetylation in yeast cells by employing quantitative proteomics and ribosome profiling for analysis of the consequences of failure of N-terminal acetylation in strains lacking specific N-terminal acetyl transferases. My results revealed a multi-faceted stress response in natB deletion mutant that modulates protein quality control machinery, protein biogenesis capacity, and energy regeneration pathways in order to establish protein homeostasis. Systematic analysis of proteome stability in the natB deletion mutant suggests no global effect of the loss of N-terminal acetylation on the turnover of NatB substrates, but an increase in the level of global protein aggregation. SILAC-based mass spectrometry analysis of aggregated proteins isolated from the natB deletion mutant shows no significant enrichment of NatB substrates, indicating that protein aggregation in the natB deletion mutant cannot be solely explained as a direct consequence of the loss of N-terminal acetylation. In contrast, these protein aggregates show strong enrichment for components of specific biological pathways, in particular of the translation apparatus, suggesting an underlying selective sequestration mechanism. Consistently, quantitative proteomics revealed that, on average, approximately 40% of each of the quantified ribosomal proteins is sequestered into protein aggregates in the natB deletion mutant. Moreover, the aggregated proteins showed significantly higher interaction between each other and overlapped with aggregated proteins generated upon environmental stress, suggesting a common mode of sequestration of proteins into aggregates. Interestingly, the aggregated proteins in the natB deletion mutant strongly overlap with those identified upon deletion of the genes encoding the ribosome-associated Hsp70 chaperone Ssb. In addition, deletion of SSB in the natB deletion mutant leads to synthetic growth defects. Moreover, isolation of radiolabeled protein aggregates after 5 min 35S pulse labeling showed that a fraction of the newly synthesized proteins is readily sequestered into aggregates. These findings together suggest a new link between N-terminal acetylation by NatB and co-translational protein folding activity by Ssb. Parallel analysis of natA deletion mutant revealed similar protein aggregation patterns, suggesting a general role of N-terminal acetylation in the maintenance of proteome integrity.
date: 2019
id_scheme: DOI
id_number: 10.11588/heidok.00024223
ppn_swb: 1663486840
own_urn: urn:nbn:de:bsz:16-heidok-242233
date_accepted: 2018-01-25
advisor: HASH(0x55fc36bae130)
language: eng
bibsort: ZEDANMOSTAANALYSISOF2019
full_text_status: public
place_of_pub: Heidelberg
citation:   Zedan, Mostafa  (2019) Analysis of the role of N-terminal acetylation of newly synthesized proteins in Saccharomyces cerevisiae.  [Dissertation]     
document_url: https://archiv.ub.uni-heidelberg.de/volltextserver/24223/1/Zedan_Dissertation_Final.pdf