Cell signaling, post-translational protein modifications and NMR spectroscopy

Abstract

Post-translationally modified proteins make up the majority of the proteome and establish, to a large part, the impressive level of functional diversity in higher, multi-cellular organisms. Most eukaryotic post-translational protein modifications (PTMs) denote reversible, covalent additions of small chemical entities such as phosphate-, acyl-, alkyl- and glycosyl-groups onto selected subsets of modifiable amino acids. In turn, these modifications induce highly specific changes in the chemical environments of individual protein residues, which are readily detected by high-resolution NMR spectroscopy. In the following, we provide a concise compendium of NMR characteristics of the main types of eukaryotic PTMs: serine, threonine, tyrosine and histidine phosphorylation, lysine acetylation, lysine and arginine methylation, and serine, threonine O-glycosylation. We further delineate the previously uncharacterized NMR properties of lysine propionylation, butyrylation, succinylation, malonylation and crotonylation, which, altogether, define an initial reference frame for comprehensive PTM studies by high-resolution NMR spectroscopy.

Identification of the Tau phosphorylation pattern that drives its aggregation

“…S15). Phosphorylation of this Tau441 AT8 protein by RBE leads to a simpler spectrum than for the phosphorylated Tau441-Ser262A protein, albeit with several resonances for each phospho-site because of incomplete phosphorylation (37) (Fig. 5A).…”

Differentially Isotope‐Labeled Nucleosomes To Study Asymmetric Histone Modification Crosstalk by Time‐Resolved NMR Spectroscopy

“…We analyzed crosstalks that involved the most common PTMs (S/T phosphorylation, K acetylation/methylation), but we have already delineated the NMR characteristics of equally or less abundant PTMs, such as arginine methylation, lysine crotonylation/propionylation, tyrosine phosphorylation. [16] Of outmost importance for the utility of our approach is the ability to prepare specifically and quantitatively modified histones that can be used for asNucs reconstitution. In cases where histone modifications cannot be established enzymatically, alternative schemes for the specific incorporation of modified amino acids can be employed, including non-natural amino acid technologies [17] or the use of cysteine mutant-derived PTM mimetics (histones are devoid of cysteines).…”

Detection of methylation, acetylation and glycosylation of protein residues by monitoring 13C chemical-shift changes

“…Consequently, the change in protonation upon acetylation should be the reason for the above unexpected result. Indeed, the change of protonation for nonmodified Lys, as occurs at a high pH value, leads to a  distribution (see Figure S1c Having presented the above test on lysine, we have a validation of our theoretical approach and have shown that computation of the  values, for a given nucleus, is a useful method with which to detect the methylation states of Lys (Theillet et al, 2012a(Theillet et al, , 2012b, but not acetylation (Theillet et al, 2012a). Consequently, we decided to extend this analysis to discuss methylation of Arg, and glycosylation of Ser, Asn and Thr, with model tripeptides, and the results are discussed below.…”

“…Therefore, perturbation of the pKa upon methylation is not large enough to be used as a probe with which to sense Arg modification. Second, there are other nuclei, than carbons, of the Arg side-chain, such as N  , that show large chemical-shift dispersion upon methylation (Theillet et al, 2012b). However, as noted by Theillet et al (2012b) …”

“…Since the pioneer observation of lysine methylation of a bacterial protein by Ambler & Rees (1959), there has been a rising interest in investigating protein post-translational modifications (PTMs) and their role as modulators of protein activity (Zobel-Tropp et al, 1998;Bienkiewicz & Lumb, 1999;Bannister et al, 2002;Paik et al, 2007;Bedford & Clarke, 2009;Kamieniarz & Schneider, 2009;Kamath et al, 2011;Luo, 2012;Theillet et al, 2012a;Theillet et al, 2012aTheillet et al, 2012bEvich et al, 2015;Rahimi & Costello, 2015;Schubert et al, 2015). As a consequence of their relevance, the development of fast and accurate experimental methods for detection of PTMs has also been an object of active research in the field (Kamath et al, 2011).…”