Acetyl coenzyme A synthetase is acetylated on multiple lysine residues by a protein acetyltransferase with a single Gcn5-type N-acetyltransferase (GNAT) domain in Saccharopolyspora erythraea

Reversible lysine acetylation (RLA) is used by cells of all domains of life to modulate protein function. To date, bacterial acetylation/deacetylation systems have been studied in a few bacteria (e.g., Salmonella enterica , Bacillus subtilis , Escherichia coli , Erwinia amylovora , Mycobacterium tuberculosis , and Geobacillus kaustophilus ), but little is known about RLA in antibiotic-producing actinomycetes. Here, we identify the Gcn5-like protein acetyltransferase AcuA of Saccharopolyspora erythraea ( Sac AcuA, SACE_5148) as the enzyme responsible for the acetylation of the AMP-forming acetyl coenzyme A synthetase ( Sac AcsA, SACE_2375). Acetylated Sac AcsA was deacetylated by a sirtuin-type NAD + -dependent consuming deacetylase ( Sac SrtN, SACE_3798). In vitro acetylation/deacetylation of Sac AcsA enzyme was studied by Western blotting, and acetylation of lysine residues Lys 237 , Lys 380 , Lys 611 , and Lys 628 was confirmed by mass spectrometry. In a strain devoid of Sac AcuA, none of the above-mentioned Lys residues of Sac AcsA was acetylated. To our knowledge, the ability of Sac AcuA to acetylate multiple Lys residues is unique among AcuA-type acetyltransferases. Results from site-specific mutagenesis experiments showed that the activity of Sac AcsA was controlled by lysine acetylation. Lastly, immunoprecipitation data showed that in vivo acetylation of Sac AcsA was influenced by glucose and acetate availability. These results suggested that reversible acetylation may also be a conserved regulatory posttranslational modification strategy in antibiotic-producing actinomycetes.