PTMs characterize some of the most discussed players amyloid
PTMs characterize some of the most discussed players (amyloid-beta, α-synuclein, tau, etc.) in neurodegenerative disorders, such as Alzheimer's disease (AD) and Parkinson's disease (PD) [1,7,8]. Because of the ways in which PTMs can so drastically alter the functions of proteins, they are strictly controlled biologically. Any disruption in their regulation can result in abnormal pathology. Some of the most common PTMs in neurodegenerative diseases include phosphorylation [, , ], acetylation [, , , ], glycosylation [17,18], ubiquitination [, , , ], and PTMs associated with oxidative stress [, , , , ]. This review will focus on the PTMs associated with important proteins that have been linked to AD pathology. By no means is this an exhaustive list of all PTMs connected to the disease state. There are many thorough reviews [7,8,18,, , , ] referenced herein that outline the known involvement of PTMs in AD, however, very few modification-centric studies have utilized mass spectrometry as a primary analytical tool. Here, we highlight the limited mass spectrometry-based modification work completed by ourselves and a few other groups that have taken advantage of the technique to elucidate structural and quantitative information as it relates to AD pathology.
Protein modifications have classically been investigated in a variety of biochemical/immunological approaches (ELISA, western blots, immunohistochemistry, protein assays, HPLC, mass spectrometry etc.) [2,, , ]. In recent years, mass spectrometry has played a greater role at the forefront of PTM analysis due to its sensitivity, specificity and robust nature [2,33,34]. Probably more significant, is the unbiased analysis mass spectrometry can bring to a field highly dependent on specific probes for identification. While there are still some difficulties associated with PTM analysis by mass spectrometry , which will be discussed herein, great strides have been made in the ability to qualitatively, and even quantitatively, analyze PTMs within various sample mediums (blood, CSF, 2NBDG homogenates, etc.). It may seem odd then that there are so few reports describing this technique as it pertains to AD. We will first give a general overview of the considerations associated with using mass spectrometry to characterize protein modifications and how many of the most prominent modifications, affecting proteins in AD, can be identified. Furthermore, we will examine the individual proteins, discuss briefly their documented roles in the disease state, and introduce how mass spectrometry has been, or can be, utilized, often in conjunction with other techniques, to develop a clearer molecular-based picture of PTM involvement in disease onset and progression.
Mass spectrometry of PTMs According to Parker et al., there are four main aspects to utilizing mass spectrometry for PTM analysis that must always be considered:  These four characteristics will be discussed throughout this section as they pertain to the proteins and modifications. While it is relatively easy to detect and identify a protein via tandem mass spectrometry (MS/MS) by the observation of a few fragments corresponding to highly abundant peptides, the identification of a PTM requires the detection of the modified sequence itself, along with fragments of the peptide also containing the modification  if structural information is to be elucidated. This is highly dependent on the specific peptide and ionization efficiency/proton affinity of the peptide which can be contingent on the modification . Hence, protein purification has become an integral step in the determination of PTMs by mass spectrometry [, , ]. Often times however, particularly with exploratory work or with work focused on the characterization of unknown PTMs, protein purification and/or enrichment can be improbable. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) is a common means of separating components of a sample prior to proteomic analysis, and is most useful for those components, or peptides, in relatively high abundance. It is not uncommon for a single SDS-PAGE band to contain multiple species. Therefore, other separation and purification techniques should be considered when analyzing PTMs such as liquid-chromatography coupled electrospray ionization mass spectrometry (LC-ESI MS).