Post-translational Protein Acetylation: An Elegant Mechanism for Bacteria to Dynamically Regulate Metabolic Functions

Abstract

Post-translational modifications (PTM) decorate proteins to provide functional heterogeneity to an existing proteome. The large number of known PTMs highlights the many ways that cells can modify their proteins to respond to diverse stimuli. Recently, PTMs have begun to receive increased interest because new sensitive proteomics workflows and structural methodologies now allow researchers to obtain large-scale, in-depth and unbiased information concerning PTM type and site localization. However, few PTMs have been extensively assessed for functional consequences, leaving a large knowledge gap concerning the inner workings of the cell. Here, we review understanding of N-𝜀-lysine acetylation in bacteria, a PTM that was largely ignored in bacteria until a decade ago. Acetylation is a modification that can dramatically change the function of a protein through alteration of its properties, including hydrophobicity, solubility, and surface properties, all of which may influence protein conformation and interactions with substrates, cofactors and other macromolecules. Most bacteria carry genes predicted to encode the lysine acetyltransferases and lysine deacetylases that add and remove acetylations, respectively. Many bacteria also exhibit acetylation activities that do not depend on an enzyme, but instead on direct transfer of acetyl groups from the central metabolites acetyl coenzyme A or acetyl phosphate. Regardless of mechanism, most central metabolic enzymes possess lysines that are acetylated in a regulated fashion and many of these regulated sites are conserved across the spectrum of bacterial phylogeny. The interconnectedness of acetylation and central metabolism suggests that acetylation may be a response to nutrient availability or the energy status of the cell. However, this and other hypotheses related to acetylation remain untested.

Keywords: acetylation; bacteria; lysine acetyltransferase; mass spectrometry; proteomics.

FIGURE 1

Mechanisms of Acetylation. Acetylation can be catalyzed (1) by a lysine acetyltransferase (KAT) using acetyl-CoA as the acetyl donor or (2) non-enzymatically by acetyl phosphate or acetyl-CoA. Some, but not all, acetylations can be reversed by a lysine deacetylase (KDAC).

FIGURE 2

The Pta-AckA acetate fermentation pathway generates AcP as an intermediate. Phosphotransacetylase (Pta) converts AcCoA to AcP by substituting CoA for inorganic phosphate (P_i). Acetate kinase (AckA) converts AcP to acetate, generating an ATP in the process. This pathway is reversible.

FIGURE 3

Catalytic mechanisms of enzymatic and non-enzymatic acetylation. (Top) In the enzymatic mechanism, a lysine to be acetylated binds at the acceptor site of a lysine acetyltransferase (KAT) and AcCoA binds at the donor site of the KAT called the P-loop (red) with consensus motif Gln/Arg-x-x-Gly-x-Gly/Ala (Salah Ud-Din et al., 2016). A catalytic glutamate deprotonates the epsilon amino group of the target lysine. The lysine performs a nucleophilic attack on the carbonyl carbon of AcCoA, resulting in acetylation of the lysine. The CoA group becomes protonated by a tyrosine, which regenerates the KAT and facilitates the release of the free CoA and the target protein. (Bottom) In this example of non-enzymatic acetylation, AcP is bound through its negatively charged phosphoryl group to a neighboring patch (yellow) that contains positively charged residues and/or residues that can form hydrogen bonds. In this example, the lysine is deprotonated by a glutamate on the same protein. The lysine performs a nucleophilic attack on the carbonyl carbon of AcP resulting in an acetyllysine. Inorganic phosphate is released as a byproduct of the reaction. A mechanism similar to this can be considered for non-enzymatic AcCoA-dependent acetylation.

FIGURE 4

AcP is made as a consequence of overflow metabolism. (Left) When bacteria like E. coli are growing on low concentrations of carbon, much of the carbon is put into the TCA cycle, lipid metabolism, and metabolite biosynthesis. Acetate production is very low. (Right) When the concentration of carbon increases, the flux of carbon through glycolysis depletes the limiting pool of free CoA. To continue to consume carbon, E. coli can regenerate free CoA by fermenting acetate. This results in the production of AcP.