Recombinant production of Aβ1-42 peptide and analysis of interactions between Aβ1-42 and Peptide Inhibitor NanoParticles (PINPs) developed as a potential novel treatment for Alzheimer’s Disease

摘要

This study addressed two main aims relating to the Aβ1-42 peptide – widely demonstrated to play a key role in the pathogenesis of Alzheimer’s Disease (AD). Aβ1-42 is an amyloid peptide that readily self-associates forming neurotoxic aggregates. With the primary risk factor for AD being age, and ageing populations increasing, the need for research into methods of reducing the levels of Aβ1-42 aggregates has never been greater. The first aim of this study was to recombinantly produce the Aβ1-42 peptide using a previously published protocol in order to produce stocks for future experimental use. Producing Aβ1-42 recombinantly results in less peptide variability than occurs via peptide synthesis, and is a much cheaper source of the peptide than commercial procurement. The peptide was expressed by induction of Escherichia coli to express a fusion protein encoding the Aβ1-42 peptide in addition to specific regions necessary for purification. Analysis of the purified Aβ1-42 peptide by transmission electron microscopy demonstrated that the peptide was able to self-associate forming a variety of structures characteristic of Aβ1-42 aggregation as illustrated in the literature. The second aim of the study was to evalauate the effect of Peptide Inhibitor NanoParticles (PINPs) upon Aβ1-42 aggregation. PINPs are second-generation liposomes with the RI-OR2- TAT peptide attached to the surface. RI-OR2-TAT has been found previously to reduce Aβ1-42 aggregation in mouse models of AD. Analysis of the effect of PINPs upon Aβ1-42 aggregation 2 was performed using transmission electron microscopy and fluorescence based assays with PINPs being found to directly bind early and late stage Aβ1-42 aggregates and reduce levels of aggregation. Based on the findings of this study, amendments to the Aβ1-42 production protocol are proposed and it is recommended that PINPs be carried forward into clinical trials as a potential treatment option for AD.