Mass Spectrometry Analysis of Lysine Posttranslational Modifications of Tau Protein from Alzheimer’s Disease Brain

Abstract

Recent advances in mass spectrometry (MS)-based proteomics have greatly facilitated the robust identification and quantification of posttranslational modifications (PTMs), including those that are present at substoichiometric site occupancies. The abnormal posttranslational modification and accumulation of the microtubule-associated protein tau has been implicated in the pathogenesis of Alzheimer’s disease (AD), and it is thought that the primary mode of regulation of tau occurs through PTMs. Several studies have been published regarding tau phosphorylation; however, other tau PTMs such as ubiquitylation, acetylation, methylation, oxidation, sumoylation, nitration, and glycosylation have not been analyzed as extensively. The comprehensive detection and delineation of these PTMs is critical for drug target discovery and validation. Lysine-directed PTMs including ubiquitylation, acetylation, and methylation play key regulatory roles with respect to the rates of tau turnover and aggregation. MS-based analytical approaches have been used to gain insight into the tau lysine-directed PTM signature that is most closely associated with neurofibrillary lesion formation. This chapter provides details pertaining to the liquid chromatography tandem mass spectrometry (LC-MS/MS)-based analysis of the lysine-directed posttranslational modification of tau.

Protein SUMOylation modification and its associations with disease

“…The SUMOylation of tau protein is also associated with the development of AD [ 59 , 60 ]. Tau protein can be both SUMOylated and ubiquitylated [ 61 ]. Inhibition of the proteasomal degradation pathway increases the tau ubiquitination and decreases its SUMOylation, suggesting that SUMO and ubiquitin might compete to regulate tau stability [ 53 ].…”

Origin and pathophysiology of protein carbonylation, nitration and chlorination in age-related brain diseases and aging

“…Nitration of ATP synthase α-chain and VDAC-1 is associated with mitochondrial dysfunction and neuronal cell death in the AD hippocampus. Moreover, nitrated proteins are usually tagged for selective destruction in proteasomes, but in AD this pathway may be defective due to oxidation of ubiquitin carboxy-terminal hydrolase L-1 in the inferior parietal lobule and hippocampus [ 122 , 125 , 126 ]. Main brain proteins nitrated in AD are listed in Table 3 .…”

“…MRM-based assays are considered to be the “gold standard” for MS-based targeted protein quantification since they are highly specific, precise, and accurate, and they can be multiplexed (hundreds of peptides can be quantified in a single assay), standardized and readily reproduced. A targeted proteomics method that is similar to MRM is parallel reaction monitoring (PRM) wherein an accurate mass and high-resolution mass spectrometer is used to permit the parallel detection of all target product ions [ 142 ].…”

2017 White Paper on Recent Issues in Bioanalysis: Rise of Hybrid LBA/LCMS Immunogenicity Assays (Part 2: Hybrid LBA/LCMS Biotherapeutics, Biomarkers & Immunogenicity Assays and Regulatory Agencies’ Inputs)

“…However, for precipitation-based workflows, it should be considered that since not all proteins precipitate, conditions need to be optimized, for example, solvents, temperature and salts. Multiplexing in hybrid LBA/LCMS is commonly used to measure multiple peptides from the same protein to verify and understand its biology and various proteoforms [27].…”