Acetylation dynamics and stoichiometry in Saccharomyces cerevisiae

BT Weinert, V Iesmantavicius, T Moustafa… - Molecular Systems …, 2015 - embopress.org
BT Weinert, V Iesmantavicius, T Moustafa, C Schölz, SA Wagner, C Magnes, R Zechner…
Molecular Systems Biology, 2015embopress.org
We were recently made aware of an error in the equation that we used to estimate
acetylation stoichiometry in the above-mentioned article. While this error does not have a
major impact on our results or their interpretation, some of the numbers are changed slightly
and here we provide updated figures and tables, the equation used, and the revised
stoichiometry estimates for individual sites. To calculate initial stoichiometry (SI), we
previously divided the degree of partial chemical acetylation (C) by the SILAC ratio (R) of …
We were recently made aware of an error in the equation that we used to estimate acetylation stoichiometry in the above-mentioned article. While this error does not have a major impact on our results or their interpretation, some of the numbers are changed slightly and here we provide updated figures and tables, the equation used, and the revised stoichiometry estimates for individual sites. To calculate initial stoichiometry (SI), we previously divided the degree of partial chemical acetylation (C) by the SILAC ratio (R) of increased acetylation after partial chemical acetylation, SI= C/R. However, in the correct equation, the SILAC ratio should be subtracted by (1ÀC), SI= C/(RÀ (1ÀC)). This equation was derived as follows. If the final stoichiometry is (SF)= SI+ C (1ÀSI) and the acetylated peptide SILAC ratio (R)= SF/SI, then solving the above equation for SI in terms of SF/SI yields SI= C/((SF/SI) À (1ÀC))= C/(RÀ (1ÀC)). We determined that C was< 1%, and we estimated stoichiometry using the conservative assumption that C= 1%(0.01); therefore, we previously calculated stoichiometry using the equation S= 0.01/R, while the correct equation is S= 0.01/(RÀ0. 99). We further realized that we could use a previously published equation (Olsen et al, 2010) for calculating stoichiometry and this equation yielded results identical to the revised formula presented above. We now include both equations and the updated stoichiometry estimates in Supplementary Table S9. The new equation had a very small impact on our stoichiometry estimates. Regardless, we have updated Figure 5D and 5H (see below), which were based on our stoichiometry estimates. Most of our other analyses were based on the SILAC ratio after chemical acetylation; therefore, these results were independent of the stoichiometry calculation.
An additional issue is that several of the sites we previously classified as having a high stoichiometry (SILAC ratio L/H< 2) were quantified on doubly acetylated peptides. However, we cannot accurately estimate the degree of partial chemical acetylation at individual sites on doubly acetylated peptides, and for this reason, we could not accurately estimate stoichiometry of acetylation sites occurring on such peptides. In our dataset, doubly acetylated peptides accounted for 20 sites, 14 of which were estimated to have high stoichiometry. These 14 sites included histone H4 (Hhf1) lysines 6, 9, 13, and 17; histone H2AZ (Htz1) lysines 4, 9, and 11; histone H2B (Htb2) lysine 17; Esa1 lysines 6 and 13; Ahc1 lysines 532 and 538; Sgf73 lysine 199; and Snf2 lysine 1494. Sites quantified
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