Lysine acetylation regulates the activity of <italic>Escherichia coli</italic> S-adenosylmethionine synthase

Abstract

Lysine acetylation is one of the most abundant post-translational modifications. However, physiological roles of this modification in bacteria are largely unknown. Previous protein acetylome analysis showed that Escherichia coli adenosylmethionine synthase (MAT) undergoes acetylation in vivo, but the biological functions of this modification still need to be uncovered. In this study, MAT of E. coli was over-expressed and purified. Subsequent mass spectrometry analysis showed that 12 lysine residues of the protein were acetylated. Site-directed mutagenesis analysis was performed and the results showed that acetylated lysine residues play important roles in the enzymatic activity of MAT. Next, deacetylation assay was performed by using CobB as the deacetylase, and the results showed that CobB could deacetylate MAT in vitro. In addition, the enzymatic activities of acetylated and deacetylated MAT were compared in vitro, and results showed that acetylation led to a decrease in its enzymatic activity, which could be reversed by CobB deacetylation. Altogether, our data suggest that CobB modulates the enzymatic activity of E. coli MAT in vitro.

The Global Acetylome of the Human Pathogen Vibrio cholerae V52 Reveals Lysine Acetylation of Major Transcriptional Regulators

“…The V. cholerae S-adenosylmethionine synthetase (KNH48827) was shown to be acetylated on five residues. Three of these acetylation events (K4, K267, and K285) regulate the activity of the enzyme in E. coli (Sun et al, 2016 ). A homolog of the Mycobacterium smegmatis universal stress protein was identified in V. cholerae (KNH48736) that shared 23% identity with that of the Mycobacterium smegmatis enzyme.…”

Mechanisms, Detection, and Relevance of Protein Acetylation in Prokaryotes

“…However, due to the presence and availability of AcCoA in cells, these nonenzymatic reactions most likely also modify the bacterial proteome (105, 108). Indeed, as shown for mitochondrial proteins, exposure of several purified bacterial proteins to AcCoA alone leads to their acetylation (108–110).…”

Regulation, Function, and Detection of Protein Acetylation in Bacteria

“…Mutations of the Lys residues to Gln (acetyl mimic) or Arg (unacetylated) resulted in partial loss of NhoA enzymatic function in vivo. The enzyme adenosylmethionine synthase (MAT) contains 12 acetylation sites (93). MAT is not a substrate of the acetyltransferase YfiQ but rather becomes autoacetylated in the presence of acetyl-CoA and then deacetylated by CobB in vitro.…”

“…One problem is that acetylases are often promiscuous in vitro and can acetylate nonendogenous sites, making potential downstream biochemical assays difficult to interpret (61). Gln substitution mutants purified from E. coli can be used for such in vitro studies (93,101), but the same caveats apply as discussed above for the in vivo interpretation of results using mutations to mimic acetylated proteins. An alternative approach is to generate fully acetylated proteins by using an E. coli strain that has a coevolved tRNA-tRNA synthetase pair, which inserts acetyllysine when an amber codon is read (138).…”