Proteomics

Dataset Information

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Accurate quantification of site-specific acetylation stoichiometry reveals the impact of sirtuin deacetylase CobB on the E. coli acetylome

ABSTRACT: We applied SD-SILAC to determine absolute acetylation stoichiometry in exponentially-growing and stationary-phase wild type and sirtuin deacetylase CobB-deficient E. coli. We further assayed stoichiometry in phophotransacetylase (pta) and acetate kinase (ackA) mutant strains in the presence and absence of the sirtuin inhibitor nicotinamide. Comparison to relative quantification under the same conditions validated our stoichiometry estimates at hundreds of sites, demonstrating the accuracy of our method. We measured acetylation stoichiometry at 3,669 unique sites and found that the vast majority of acetylation occurs at very low stoichiometry (median 0.04%). Manipulations that cause increased nonenzymatic acetylation by acetyl-phosphate (AcP), such stationary-phase arrest and deletion of ackA, resulted in globally increased acetylation stoichiometry. Similar to sirtuin deacetylase 3 (SIRT3) in mitochondria, CobB suppressed acetylation to lower than median stoichiometry at hundreds of sites in WT, pta, and ackA cells. These results suggest an evolutionarily conserved function of sirtuin deacetylases in suppressing low stoichiometry acetylation.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Escherichia Coli

SUBMITTER: Brian Weinert  

LAB HEAD: Brian T. Weinert

PROVIDER: PXD005757 | Pride | 2017-03-07

REPOSITORIES: Pride

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Dataset's files

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20160809_QE2_nLC0_SS_SA_Ecoli_CCA_L_PCA_H_ratio1_10_AcKIP_pH4.raw Raw
20160809_QE2_nLC0_SS_SA_Ecoli_CCA_L_PCA_H_ratio1_10_AcKIP_pH4_5.raw Raw
20160809_QE2_nLC0_SS_SA_Ecoli_CCA_L_PCA_H_ratio1_10_AcKIP_pH5_5.raw Raw
20160809_QE2_nLC0_SS_SA_Ecoli_CCA_L_PCA_H_ratio1_10_AcKIP_pH9.raw Raw
20160809_QE2_nLC0_SS_SA_Ecoli_CCA_L_PCA_H_ratio1_250_AcKIP_pH4.raw Raw
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Publications

Accurate Quantification of Site-specific Acetylation Stoichiometry Reveals the Impact of Sirtuin Deacetylase CobB on the <i>E. coli</i> Acetylome.

Weinert Brian Tate BT   Satpathy Shankha S   Hansen Bogi Karbech BK   Lyon David D   Jensen Lars Juhl LJ   Choudhary Chunaram C  

Molecular & cellular proteomics : MCP 20170302 5

Lysine acetylation is a protein posttranslational modification (PTM) that occurs on thousands of lysine residues in diverse organisms from bacteria to humans. Accurate measurement of acetylation stoichiometry on a proteome-wide scale remains challenging. Most methods employ a comparison of chemically acetylated peptides to native acetylated peptides, however, the potentially large differences in abundance between these peptides presents a challenge for accurate quantification. Stable isotope lab  ...[more]