Abstract
Acetylation is a protein post-translational modification (PTM) that can affect a variety of cellular processes. In bacteria, two PTM Nε-acetylation mechanisms have been identified: non-enzymatic/chemical acetylation via acetyl phosphate or acetyl coenzyme A and enzymatic acetylation via protein acetyltransferases. Prior studies have shown that extensive acetylation of Nε-lysine residues of numerous proteins from a variety of bacteria occurs via non-enzymatic acetylation. In Escherichia coli, new Nε-lysine acetyltransferases (KATs) that enzymatically acetylate other proteins have been identified, thus expanding the repertoire of protein substrates that are potentially regulated by acetylation. Therefore, we designed a study to leverage the wealth of structural data in the Protein Data Bank (PDB) to determine: (1) the 3D location of lysine residues on substrate proteins that are acetylated by E. coli KATs, and (2) investigate whether these residues are conserved on 3D structures of their homologs. Five E. coli KAT substrate proteins that were previously identified as being acetylated by YiaC and had 3D structures in the PDB were selected for further analysis: adenylate kinase (Adk), isocitrate dehydrogenase (Icd), catalase HPII (KatE), methionyl-tRNA formyltransferase (Fmt), and a peroxide stress resistance protein (YaaA). We methodically compared over 350 protein structures of these E. coli enzymes and their homologs; to accurately determine lysine residue conservation requires a strategy that incorporates both flexible structural alignments and visual inspection. Moreover, our results revealed discrepancies in conclusions about lysine residue conservation in homologs when examining linear amino acid sequences compared to 3D structures.
Highlights
Post-translational modifications (PTMs) are chemical changes that occur on proteins and include, but are not limited to, the covalent attachment of various functional groups on proteins
Lysine (K) acetylation sites are shown with sticks and their labels are colored based on type of acetylation that occurs: K-12 lysine acetyltransferase (KAT) sites are in blue, acetyl phosphate (AcP) sites are in red, and sites acetylated by both a KAT and AcP are in purple. (Adk) Two conformations of the adenylate kinase (Adk) monomer are shown: the closed (PDB ID: 1ake) and open (PDB ID: 6f7u) conformations are at the top and middle of the panel, respectively; an overlay of the two conformations is shown at the bottom of the panel
Overview of Escherichia. coli Lysine Acetyltransferase Substrate Protein Functions, 3D Crystal Structures and Location of Acetylated Lysine Residues Adenylate Kinase Adenylate kinase (Adk) is a phosphotransferase that catalyzes the reversible conversion of ATP and AMP to two molecules of ADP in the presence of Mg2+
Summary
Post-translational modifications (PTMs) are chemical changes that occur on proteins and include, but are not limited to, the covalent attachment of various functional groups on proteins. Often, these modifications can affect protein function by altering DNA/RNA binding (Matsuzaki et al, 2005; Xu et al, 2019), protein-protein interactions (Su et al, 2017), and protein localization (Ishfaq et al, 2012). While hundreds of bacterial proteins have been identified as Nε-acetylated in E. coli, these reports have led to significant debate in the field regarding the relevance of this modification in prokaryotes. If Nε-acetylation is a physiologically important mode of bacterial protein regulation, these sites of Nε-acetylation theoretically should be conserved across homologs
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