Proteomics of Alzheimer’s Disease Steven Bhutra Biochem 118 Dr. BrutlagAlzheimer’s disease (AD) is the most common form of dementia in the elderly. This incurable neurological degenerative disease is typically diagnosed in people over 65. Alzheimer’s can be incredibly difficult to diagnose because many of the symptoms can be mistakenly attributed to age-related concerns: the most common symptom is memory loss. As the disease manifests, symptoms include confusion, irritability, mood swings, and language breakdown. Eventually, control of the body is lost, leading to death. Since the disease can be undiagnosed for years, latency has not been determined. When AD is suspected, the diagnosis can be confirmed with brain scans or behavioral evaluations. Since this disease is degenerative, management is essential but it comes at a huge cost, both to the checkbook and to the family. This epidemic of the elderly still has no treatment to delay the progression of the disease. With an increasingly aging population, this disease will only become even more prevalent. Doctors and researchers, hope to end this disease using a new development called proteomics. (4) Proteomics seeks to identify protein structure, modulations and protein-protein interactions in addition to protein expression levels. This procedure attempts to study biological processes comprehensively through the systematic analysis of the proteins expressed in a cell and is often thought of as the next step after genomics. Genomics is much simpler, as it focuses on the genome, which is consistent throughout its cells. However, proteomes change from cell to cell depending on their function. Previous assays of proteins, based on sequencing of the genome, were largely inaccurate because these studies were based on mRNA hypothetical translation. Many enzymes and miRNA inhibit translation altering the expected protein from the original strand. Moreover, the single mRNA strand may not translate continuously giving rise to multiple types of proteins through the process of alternate splicing. Any protein can undergo a wide range ofposttranslational modifications, including attaching to another protein or RNA molecule. The old way of detecting proteins wasn’t easy as a hodgepodge of proteins within the same cells can make any study quite complicated, but with this complication comes a plethora of opportunity to investigate the protein spectrum of a cell and its biological functions; consequently to detect novel drug targets and diagnostic markers. (5, 7) Previous protein analysis techniques examined proteins one by one; in proteomics this is done on a large scale. Despite a variety of new approaches, proteomics (in the study of Alzheimer’s) largely relies on the classic two-dimensional (2-D) electrophoreses and mass spectrometry (MS) to identify proteins. 2-D electrophoreses is used to separate proteins based on two criteria: difference in net charge via pH gradient and molecular masses or molility. MS is an identification technique that identifies the chemical composition of a protein based on the mass to charge ratio. First a gel separates the proteins by specific proteases in the 2-D electrophoreses and then the mass spectrum is obtained via peptide mass fingerprint (PMF). In this process, the protein activation site is catalyzed with a specific enzyme, the resulting peptides are weighed and cross-referenced with computer generated theoretical PMFs, identifying the protein. In this way both 2-D gel and mass spectrum can be compared against databases to identify proteins. (1) Recently, however, a process called protein microarray has revolutionized proteomics. In this process a piece of glass is fixed with different proteins at different locations forming an array. These proteins are bound to similar proteins revealing fluorescent protein-protein interactions. (7) Many recent studies involving proteomics attribute oxidative stress as the major cause of the disease. Oxidative stress can cause irreversible modulations to susceptible proteins leading to structural and functional modifications. Protein modulations such as carbonylation, nitration and protein-protein cross linking are generally associated with loss of function and may lead todegradation of the damaged protein, or accumulation which can have even more detrimental effects. Identifying proteins has given rise to understanding the mechanism of the disease. The oxidatively modified proteins closely catalogue known developments in the disease: accumulation of damaged proteins, shortened dendrite lengths, neurotoxicity, excess ubiquination, and dysfunction of energy metabolism. (2, 3) Alzheimer’s disease is defined as a neurodegenerative disorder characterized by neurofibrillary tangles composed of hyperphosphorylated tau proteins connected with paired helical filaments and senile amyloid plaques that accumulate in inferior parietal lobule and hippocampus. Tau is a microtubule protein that is involved in assembly and stabilization. Neurofibrillary degeneration involves the hyperphosphorylation of tau, which critically impairs its binding capacity to a microtubule disrupting axonal cytoskeleton. Peptide-proyl cis-trans isomerase or Pin1 has been confirmed to regulate phosphorylation of tau by proteomic methods. In Alzheimer’s disease however, Pin1 is oxidized which impairs its regulatory function. (See diagram below) In effect, the oxidation of Pin1 plays a role in the accumulation of phosporylated tau linking oxidative damage to tangle formation causing the disease. Through the use of proteomics, oxidation of certain proteins such as Pin1 have been linked in to the development of the disease. Pin1 could be used as a target for drug designers. (2, 3) Diagram adapted by Rachel Eastwood from original courtesy of Kun Ping LuThe disease can be further characterized by neuronal death and loss of synaptic connections within the already stated brain regions. Dihydropyrimidinase protein (DRP) is also an identified protein in the development of AD. DRP is involved in axonal expansion and guiding through the transmission and adjustment of extracellular signals. Already mentioned, one of the traditional symptoms of AD is memory loss, which is related to a decrease in neuronal connections and a shortened length of the dendrite. In fact, through the use of protein identification, DRP has been found in the neurofibrillary tangles of Alzheimer’s
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