Features and regulation of non-enzymatic post-translational modifications

Abstract

Non-enzymatic post-translational modifications of proteins can occur when a nucleophilic or redox-sensitive amino acid side chain encounters a reactive metabolite. In many cases, the biological function of these modifications is limited by their irreversibility, and consequently these non-enzymatic modifications are often considered as indicators of stress and disease. Certain non-enzymatic post-translational modifications, however, can be reversed, which provides an additional layer of regulation and renders these modifications suitable for controlling a diverse set of cellular processes ranging from signaling to metabolism. Here we summarize recent examples of irreversible and reversible non-enzymatic modifications, with an emphasis on the latter category. We use two examples, lysine glutarylation and pyrophosphorylation, to highlight principles of the regulation of reversible non-enzymatic post-translational modifications in more detail. Overall, a picture emerges that goes well beyond nonspecific chemical reactions and cellular damage, and instead portrays multifaceted functions of non-enzymatic post-translational modifications.

A chemoproteomic portrait of the oncometabolite fumarate

“…Building on our analysis of fumarate's unique labeling profile, is tempting to speculate that the polar labeling environment preferred by fumarate-sensitive cysteines may predispose the high stoichiometry display of S-succination towards solvent-exposed surfaces capable of disrupting biomolecular interactions. While relatively few examples of non-enzymatic reactive metabolite-dependent PPIs exist, 51 the role of cysteine oxidation in regulating such interactions is well-precedented. 55,[71][72] Our studies suggest further investigation of fumarate-dependent protein-protein and protein-nucleic acid interactions is warranted.…”

“…Covalent modifications driven by metabolite reactivity are largely expected to exert deleterious effects on protein function. 51 However, as a final experiment we wondered whether chemoproteomic analyses may also be capable of identifying pathways positively influenced by FH mutation. To explore this idea we re-analyzed our chemoproteomic data from FH-/-and FH+/+ rescue HLRCC cell lines, particularly focusing on FH-regulated cysteines with R values <1 (blue region, Fig.…”

Chemoproteomic Profiling of Phosphoaspartate Modifications in Prokaryotes

“…Zuschriften profiling provides unequivocal identification of modified residues.T his not only enables quantitation of pD signalling dynamics at specific sites but also allows for exploratory mapping in unannotated proteomes and organisms.I nl ine with this attribute,t he dataset herein contained many highconfidence,r eproducible DBHA-modified residues that are not previously annotated as phosphoaspartate sites.T hese sites may represent heretofore unrecognized phosphoaspartate sites that are introduced through enzymatic or nonenzymatic means,ashas been observed for other intrinsically reactive metabolic intermediates like acetylphosphate, [30][31][32] which is used in vitro to chemically phosphorylate Asp sites in response regulators. [33] Beyond phosphoaspartate,o ther modifications could contribute to the profile observed, including methylesterification, ADP-ribosylation, and potentially other less-characterized PTMs that occur at aspartic acids in prokaryotes.D espite this potential for overlap,w e view the labelling of additional electrophilic modifications as af eature not af law of DBHA-profiling because the use quantitative LC-MS/MS provides site-specific resolution necessary for follow up analysis at as ite-by-site level.…”

“…In line with this attribute, the dataset herein contained many high confidence, reproducible DBHA-modified residues that are not previously annotated as phosphoaspartate sites. These sites may represent heretofore unrecognized phosphoaspartate sites that are introduced through enzymatic or nonenzymatic means, as has been observed for other intrinsically reactive metabolic intermediates like acetylphosphate, [30–32] which is used in vitro to chemically phosphorylate Asp sites in response regulators. [33] Beyond phosphoaspartate, other modifications could contribute to the profile observed, including methylesterification, ADP-ribosylation, and potentially other less-characterized PTMs that occur at aspartic acids in prokaryotes.…”

Strong anion exchange‐mediated phosphoproteomics reveals extensive human non‐canonical phosphorylation

“…Until recently, phosphorylation of the hydroxyl‐containing amino acids serine (Ser), threonine (Thr) and tyrosine (Tyr) was thought to be the primary mode of phosphorylation‐mediated signalling in non‐plant eukaryotes. However, a growing body of evidence (Besant & Attwood, ; Fan et al , ; Fraczyk et al , ; Fuhs et al , ; Shen et al , ; Wieland & Attwood, ; Panda et al , ; Srivastava et al , ; Xu et al , ,b; Fuhs & Hunter, ) indicates that phosphorylation of other amino acids, termed here “non‐canonical” phosphorylation, including His, Asp, Glu, Lys, Arg and Cys [in addition to pyrophosphorylation of Ser and Thr to form ppSer and ppThr (Chanduri et al , ; Harmel & Fiedler, ), and Lys polyphosphorylation (Bentley‐DeSousa & Downey, )], may also regulate protein signalling functions. Of note, the generation of both site‐specific and generic antibodies against phosphohistidine (pHis) (Kee et al , , ; Fuhs et al , ; Lilley et al , ) has recently allowed mammalian cell‐type independent roles for this PTM to be elucidated (Fuhs et al , ) in processes as diverse as ion channel regulation (Srivastava et al , ) and T‐cell signalling (Panda et al , ), and during cell proliferation, differentiation and migration.…”