Protein acetylation on 2-isopropylmalate synthase from Thermus thermophilus HB27

Protein lysine N-epsilon-acetylation is one of the important factors regulating cellular metabolism. We performed a proteomic analysis to identify acetylated proteins in the extremely thermophilic bacterium, Thermus thermophilus HB27. A total of 335 unique acetylated lysine residues, including many metabolic enzymes and ribosomal proteins, were identified in 208 proteins. Enzymes involved in amino acid metabolism were the most abundant among acetylated metabolic proteins. 2-Isopropylmalate synthase (IPMS), which catalyzes the first step in leucine biosynthesis, was acetylated at four lysine residues. Acetylation-mimicking mutations at Lys332 markedly decreased IPMS activity in vitro, suggesting that Lys332, which is located in subdomain II, plays a regulatory role in IPMS activity. We also investigated the acetylation-deacetylation mechanism of IPMS and revealed that it was acetylated non-enzymatically by acetyl-CoA and deacetylated enzymatically by TT_C0104. The present results suggest that leucine biosynthesis is regulated by post-translational protein modifications, in addition to feedback inhibition/repression, and that metabolic enzymes are regulated by protein acetylation in T. thermophilus.