# Acetyl‐Phosphate Dependent Protein Acetylation in Neisseria gonorrhoeae

## TL;DR

This study shows that protein acetylation in Neisseria gonorrhoeae affects metabolism and virulence, with acetyl-phosphate playing a key role.

## Contribution

The study reveals the role of non-enzymatic acetylation in N. gonorrhoeae's metabolism and pathogenicity.

## Key findings

- 88% of the N. gonorrhoeae proteome is acetylated, with many proteins involved in central metabolism.
- The ΔackA strain cannot use pyruvate as a carbon source, showing acetylation affects carbon utilization.
- Altered acetyl-phosphate levels change bacterial virulence, with ΔackA being more pathogenic in a wax moth model.

## Abstract

The disease gonorrhoea is caused by the sexually transmitted pathogen Neisseria gonorrhoeae. This bacterium is an obligate human pathogen that can survive intracellularly through the expression of specific pathogenicity determinants. Protein post‐translational modifications have been shown to be involved in the regulation of gene transcription and metabolism. Here, we studied the role of non‐enzymatic acetylation by acetyl‐phosphate in N. gonorrhoeae. This was achieved through the deletion of pta and ackA genes from the phosphotransacetylase‐acetate kinase pathway (PTA‐AKA) that modulate the level of acetyl‐phosphate in the cell. As predicted, more protein acetylation was observed in the ΔackA strain. Using immunoaffinity purification of acetylated peptides and LC‐MS/MS we demonstrated that 88% of the detectable N. gonorrhoeae proteome (1343 proteins) is acetylated. With many of the acetylated proteins involved in central metabolism especially in pyruvate utilisation. Growth studies showed that the ΔackA strain was unable to utilise pyruvate as a carbon source, whereas it could grow on glucose as well as the wild‐type. Furthermore, a deacetylase enzyme was identified and its gene mutated (Δhdac), this allowed the identification of a number of putative targets for HDAC, including phosphotransacetylase. We found that gonococcal pathogenicity was changed by acetyl‐phosphate concentration, with the ΔackA strain killing the wax moth larvae faster than the wild‐type, whereas the Δpta strain was non‐pathogenic in this model. The data obtained suggest that non‐enzymatic protein acetylation in N. gonorrhoeae plays an important role in the central metabolism, carbon source utilisation, and virulence of this bacterium.

The role of acetyl‐phosphate‐dependent non‐enzymatic protein acetylation in Neisseria gonorrhoeae was investigated through the deletion of the phosphotransacetylase (pta) and the acetate kinase (ackA) genes, this led to a decrease and increase in protein acetylation, respectively. This work suggests that non‐enzymatic protein acetylation in N. gonorrhoeae has a crucial role in central metabolism, carbon source utilisation, and virulence.