Lysine acetylation is a highly abundant and evolutionarily conserved modification in Escherichia coli - PubMed (original) (raw)

Lysine acetylation is a highly abundant and evolutionarily conserved modification in Escherichia coli

Junmei Zhang et al. Mol Cell Proteomics. 2009 Feb.

Abstract

Lysine acetylation and its regulatory enzymes are known to have pivotal roles in mammalian cellular physiology. However, the extent and function of this modification in prokaryotic cells remain largely unexplored, thereby presenting a hurdle to further functional study of this modification in prokaryotic systems. Here we report the first global screening of lysine acetylation, identifying 138 modification sites in 91 proteins from Escherichia coli. None of the proteins has been previously associated with this modification. Among the identified proteins are transcriptional regulators, as well as others with diverse functions. Interestingly, more than 70% of the acetylated proteins are metabolic enzymes and translation regulators, suggesting an intimate link of this modification to energy metabolism. The new dataset suggests that lysine acetylation could be abundant in prokaryotic cells. In addition, these results also imply that functions of lysine acetylation beyond regulation of gene expression are evolutionarily conserved from bacteria to mammals. Furthermore, we demonstrate that bacterial lysine acetylation is regulated in response to stress stimuli.

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Figures

F<sc>ig</sc>. 1.

Fig. 1.

Synopsis of lysine acetylation proteomics. A, schematic representation of the sequential steps used for global profiling of lysine acetylation in E. coli. B, Western blotting analysis of protein lysate from E. coli with anti-acetyllysine antibody from ImmuneChem Inc. Lane 1, 30 μg of whole cell lysate from E. coli probed with anti-acetyllysine antibody; lane 2, competition with acetylated BSA (300 μg/ml). C, an example of MS and MS/MS analysis of a lysine-acetylated peptide for peptide identification and mapping of acetyllysine site in E. coli. Left panel, full MS spectrum at a retention time of 63.69 min. Right panel, MS/MS spectrum of m/z 1261.2, which identifies acetylated peptide YYQGTPSPVK*HPELTDMVIFR in isocitrate dehydrogenase, a metabolic protein in E. coli.

F<sc>ig</sc>. 2.

Fig. 2.

A, pie chart of functionally annotated protein groups that are lysine-acetylated. B, density map of lysine-acetylated peptides. The frequency of occurrence of amino acid residues surrounding sites of lysine acetylation was calculated, relative to the frequency of the residue within the entire E. coli genome, and schematically represented by a density map using a method described previously (11). Prevalence of specific amino acids at positions surrounding lysine acetylation sites is shown.

F<sc>ig</sc>. 3.

Fig. 3.

Lysine-acetylated proteins involved in glucose degradation and TCA cycle. Proteins identified as lysine-acetylated in the E. coli screen are underlined. Those identified in a screen of mammalian cells (11) are marked with an asterisk.

F<sc>ig</sc>. 4.

Fig. 4.

Western blotting analysis of protein lysate with an anti-acetyllysine antibody from ImmuneChem Inc. The 40-μg protein lysate was resolved in 4–20% SDS-PAGE. The Western blotting analysis was carried out as described previously (11). Each of the gel bands that have significant differences between the two E. coli strains or among the four oxygen conditions is labeled with an arrow on the left. The molecular weight markers are labeled on the right. A, lysine acetylation profiles of two E. coli strains (Lanes 1 and 3, MG1655; Lanes 2 and 4, JW1106). Lanes 1 and 2, probed with anti-acetyllysine antibody and non-acetylated BSA (300 μg/2 ml); Lanes 3 and 4, probed with anti-acetyllysine antibody and acetylated BSA (300 μg/2 ml). B, impact of nutritional conditions (E. coli MG1655). Lane 5, control and Lane 6, starvation. C, influences of oxygen conditions (E. coli MG1655). Lane 7, control (100% air); Lane 8, 25% air; Lane 9, 5% air; and Lane 10, 100% nitrogen.

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