de Castro Nascimento Sousa, A. C.

bioRxiv2024

Beyond Histones: Unveiling the Functional Roles of Protein Acetylation in Prokaryotes and Eukaryotes

Lysine acetylation plays a crucial role in cellular processes and is found across various evolutionary organisms. Recent advancements in proteomic techniques revealed the presence of acetylation in thousands of non-histone proteins. Here, we conducted extensive meta-analysis of 48 acetylomes spanning diverse organisms, including archaea, bacteria, fungi, protozoa, worms, plants, insects, crustacea, fish, and mammals. Our analyzes revealed a predominance of a single acetylation site in a protein detected in all studied organisms, and proteins heavily acetylated, with >5-10 acetylated-sites, were represented by Hsp70, histone or transcription GTP-biding domain. Moreover, using gene enrichment approaches we found that ATP metabolic processes, glycolysis, aminoacyl-tRNA synthetase pathways and oxidative stress response are among the most acetylated cellular processes. Finally, to better explore the regulatory function of acetylation in glycolysis and oxidative stress we used aldolase and superoxide dismutase A (SODA) enzymes as model. For aldolase, we found that K147 acetylation, responsible to regulate human enzyme, conserved in all phylogenic clade, suggesting that this acetylation might play the same role in other species; while for SODA, we identified many lysine residues in different species present in the tunnel region, which was demonstrated for human and Trypanosoma cruzi, as negative regulator, also suggesting a conserved regulatory mechanism. In conclusion, this study provides insights into the conservation and functional significance of lysine acetylation in different organisms emphasizing its roles in cellular processes, metabolic pathways, and molecular regulation, shedding light in the extensive function of non-histone lysine acetylation.

biochemistry↗

bioRxiv2021

Mitochondrial sirtuin TcSir2rp3 affects TcSODA activity and oxidative stress response in Trypanosoma cruzi

Trypanosoma cruzi, the etiological agent of Chagas disease, faces a variety of environmental scenarios during its life cycle in both invertebrate and vertebrate hosts, which include changes in the redox environment that requires a fine regulation of a complex antioxidant arsenal of enzymes. Reversible post-translational modifications, as lysine acetylation, are a fast and economical way for cells to react to environmental conditions. Acetylation neutralizes the lysine positive charge conferring novel properties to the modified proteins, from changes in enzymatic activity to subcellular localization. Recently, we found that the main antioxidant enzymes, including the mitochondrial superoxide dismutase A (TcSODA) are acetylated in T. cruzi, suggesting that protein acetylation could participate in the oxidative stress response in T. cruzi. Therefore, we investigated whether mitochondrial lysine deacetylase sirtuin 3 (TcSir2rp3) was involved in the activity control of TcSODA. We observed an increased resistance to hydrogen peroxide and menadione two oxidant compounds in parasites overexpressing TcSir2rp3. Increased resistance was also found for benznidazole and nifurtimox, the two drugs available for treatment of Chagas disease, known to induce reactive oxidative and nitrosactive species in the parasite. In parallel, TcSir2rp3 overexpressing parasites showed parasites showed a reduction in the ROS levels after treatment with benznidazole and nifurtimox, suggesting a role of TcSir2rp3 in the oxidative stress response. To better understand the way TcSir2rp3 could contributes to oxidative stress response, we analyzed the expression of a key antioxidant enzyme, TcSODA, in the TcSir2rp3 overexpressing parasites and did not detect any increase in protein levels of this enzyme. However, we found that parasites overexpressing TcSir2rp3 presented higher levels of superoxide dismutase activity, and also that TcSir2rp3 and TcSODA interacts in vivo. Knowing that TcSODA is acetylated at lysine residues K44 and K97, and that K97 is located at similar region in the protein structure as K68 in human manganese superoxide dismutase (MnSOD), responsible to regulates MnSOD activity, we generated mutated versions of TcSODA at K44 and K97 and found that replacing K97 by glutamine, which mimics an acetylated lysine, negatively affects the enzyme activity in vitro. By using molecular dynamics approaches we revealed that acetylation of K97 induces specific conformational changes in TcSODA with respect of hydrogen bonding pattern to neighbor residues, specifically D94 and E96, suggesting a key participation of this residue to modulate the affinity to O2- by changing the charge availability on the surface of the enzyme. Taken together, our results showed for the first time the involvement of lysine acetylation in the maintenance of homeostatic redox state in trypanosomatids, contributing to the understanding of mechanisms used by T. cruzi to progress during the infection and opening the opportunity to explore protein acetylation as potential drug target in this parasite.

microbiology↗