Abstract
BackgroundAcetylation of lysine residues is a reversible post-translational modification conserved from bacteria to humans. Several recent studies have revealed hundreds of lysine-acetylated proteins in various bacteria; however, the physiological role of these modifications remains largely unknown. Since lysine acetylation changes the size and charge of proteins and thereby may affect their conformation, we assumed that lysine acetylation can stimulate aggregation of proteins, especially for overproduced recombinant proteins that form inclusion bodies.ResultsTo verify this assumption, we used Escherichia coli strains that overproduce aggregation-prone VP1GFP protein. We found that in ΔackA-pta cells, which display diminished protein acetylation, inclusion bodies were formed with a delay and processed faster than in the wild-type cells. Moreover, in ΔackA-pta cells, inclusion bodies exhibited significantly increased specific GFP fluorescence. In CobB deacetylase-deficient cells, in which protein acetylation was enhanced, the formation of inclusion bodies was increased and their processing was significantly inhibited. Similar results were obtained with regard to endogenous protein aggregates formed during the late stationary phase in ΔackA-pta and ΔcobB cells.ConclusionsOur studies revealed that protein acetylation affected the aggregation of endogenous E. coli proteins and the yield, solubility, and biological activity of a model recombinant protein. In general, decreased lysine acetylation inhibited the formation of protein aggregates, whereas increased lysine acetylation stabilized protein aggregates. These findings should be considered during the designing of efficient strategies for the production of recombinant proteins in E. coli cells.Electronic supplementary materialThe online version of this article (doi:10.1186/s12934-016-0590-8) contains supplementary material, which is available to authorized users.
Highlights
Acetylation of lysine residues is a reversible post-translational modification conserved from bacteria to humans
Overproduction of VP1GFP and formation of inclusion bodies (IBs) are enhanced in ΔcobB cells and inhibited in the ΔackA‐pta strain First, we analyzed the formation and processing of IBs containing the fusion VP1GFP protein, which consists of GFP and foot-and-mouth disease virus VP1 capsid protein, and retains its fluorescence in IBs [22]
We found that the overproduction of VP1GFP was enhanced in the ΔcobB strain and inhibited in ΔackA-pta cells compared to wild-type cells (Fig. 1a)
Summary
Acetylation of lysine residues is a reversible post-translational modification conserved from bacteria to humans. Several recent studies have revealed hundreds of lysine-acetylated proteins in various bacteria; the physiological role of these modifications remains largely unknown. In some cases, insoluble but active proteins sequestered in IBs can be the preferred form of the product [7] Another obstacle in recombinant protein production in E. coli cells is accumulation of acetate during extensive aerobic fermentation in cultures supplemented with glucose. Various strategies have been developed to limit acetate accumulation or reduce its negative effects [10] One of such strategies is reduction of acetate concentration and improvement of protein production achieved by deletion or downregulation of the ackA-pta pathway using antisense-RNA strategy [11, 12]. Nε-lysine acetylation can be reversed by the deacetylase CobB regardless of acetylation mechanism [17]
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