Inhibitory Activity of Insulin on Aß Aggregation Is Restricted Due to Binding Selectivity and Specificity to Polymorphic Aß States.

Clinical trials of intranasal insulin treatment for Alzheimer's patients have shown cognitive and memory improvement, but the effect of insulin has shown a limitation. It was suggested that insulin molecule binds to Aß aggregates and impedes Aß aggregation. Yet, the specific interactions between insulin molecule and Aß aggregates at atomic resolution are still elusive. Three main conclusions are observed in this work. First, insulin can interact across the fibril only to "U-shape" Aß fibrils and not to "S-shape" Aß fibrils. Therefore, insulin is not expected to influence the "S-shape" Aß fibrils. Second, insulin disrupts ß-strands along Aß fibril-like oligomers via interaction with chain A, which is not a part of the recognition motif. It is suggested that insulin affects as an inhibitor of Aß fibrillation, but it is limited due to the specificity of the polymorphic Aß fibril-like oligomer. Third, the current work proposes that insulin promotes Aß aggregation, when interacting along the fibril axis of Aß fibril-like oligomer. The coaggregation could be initiated via the recognition motif. The lack of the interactions of insulin in the recognition motif impede the coaggregation of insulin and Aß. The current work reports the specific binding domains between insulin molecule and polymorphic Aß fibril-like oligomers. This research provides insights into the molecular mechanisms of the functional activity of insulin on Aß aggregation that strongly depends on the particular polymorphic Aß aggregates.

Unique Inversion Events of Residues around the Backbone in the Turn Domain of ß-Arches in Amylin Fibrils.

Orientational inversion events of residues along the turn domains of amylin fibrils have been detected. This exceptional phenomenon has been observed in isolated amylin fibrils and in the cross-seeding amylin-Aß and amylin-NAC fibrils. These new findings provide new avenues for detection of side chain flipping and side chain inversion events in turn domains and loops of various proteins.

Mechanistic perspective and functional activity of insulin in amylin aggregation.

Insulin is a key regulatory polypeptide that is secreted from pancreatic ß-cells and has several important effects on the synthesis of lipids, regulation of enzymatic activities, blood glucose levels and the prevention of hyperglycemia. Insulin was demonstrated to self-assemble into ordered amyloid fibrils upon repeated injections, although the possible biological significance of the supramolecular structures is enigmatic. Amylin is also an amyloidogenic polypeptide that is secreted from pancreatic ß-cells and plays an important role in glycemic regulation preventing post-prandial spikes in blood glucose levels. These two amyloidogenic proteins are secreted together from the pancreas and have the ability to interact and produce insulin-amylin aggregates. So far, the molecular architecture of insulin-amylin complexes at the atomic resolution has been unknown. The current work identifies for the first time the specific π-π interactions between Y16 in insulin and F19 in amylin that contribute to the stability of the insulin-amylin complex, by using experimental and molecular modeling techniques. We performed additional experiments that verify the functional activity of insulin in amylin aggregation. Our findings illustrate for the first time the specific interactions between insulin and amylin aggregates at the atomic resolution and provide a new mechanistic perspective on the effect of insulin on amylin aggregation and may pave the way towards pharmacological intervention in this process.

Amylin-Aß oligomers at atomic resolution using molecular dynamics simulations: a link between Type 2 diabetes and Alzheimer's disease.

Clinical studies have identified Type 2 diabetes (T2D) as a risk factor of Alzheimer's disease (AD). One of the potential mechanisms that link T2D and AD is the loss of cells associated with degenerative changes. Amylin1-37 aggregates (the pathological species in T2D) were found to be co-localized with those of Aß1-42 (the pathological species in AD) to form the Amylin1-37-Aß1-42 plaques, promoting aggregation and thus contributing to the etiology of AD. However, the mechanisms by which Amylin1-37 co-aggregates with Aß1-42 are still elusive. This work presents the interactions between Amylin1-37 oligomers and Aß1-42 oligomers at atomic resolution applying extensive molecular dynamics simulations for relatively large ensemble of cross-seeding Amylin1-37-Aß1-42 oligomers. The main conclusions of this study are first, Aß1-42 oligomers prefer to interact with Amylin1-37 oligomers to form single layer conformations (in-register interactions) rather than double layer conformations; and second, in some double layer conformations of the cross-seeding Amylin1-37-Aß1-42 oligomers, the Amylin1-37 oligomers destabilize the Aß1-42 oligomers and thus inhibit Aß1-42 aggregation, while in other double layer conformations, the Amylin1-37 oligomers stabilize Aß1-42 oligomers and thus promote Aß1-42 aggregation.

Bacterial magnetosome biomineralization--a novel platform to study molecular mechanisms of human CDF-related Type-II diabetes.

Cation diffusion facilitators (CDF) are part of a highly conserved protein family that maintains cellular divalent cation homeostasis in all organisms. CDFs were found to be involved in numerous human health conditions, such as Type-II diabetes and neurodegenerative diseases. In this work, we established the magnetite biomineralizing alphaproteobacterium Magnetospirillum gryphiswaldense as an effective model system to study CDF-related Type-II diabetes. Here, we introduced two ZnT-8 Type-II diabetes-related mutations into the M. gryphiswaldense MamM protein, a magnetosome-associated CDF transporter essential for magnetite biomineralization within magnetosome vesicles. The mutations' effects on magnetite biomineralization and iron transport within magnetosome vesicles were tested in vivo. Additionally, by combining several in vitro and in silico methodologies we provide new mechanistic insights for ZnT-8 polymorphism at position 325, located at a crucial dimerization site important for CDF regulation and activation. Overall, by following differentiated, easily measurable, magnetism-related phenotypes we can utilize magnetotactic bacteria for future research of CDF-related human diseases.

Cation diffusion facilitators transport initiation and regulation is mediated by cation induced conformational changes of the cytoplasmic domain.

Cation diffusion facilitators (CDF) are part of a highly conserved protein family that maintains cellular divalent cation homeostasis in all domains of life. CDF's were shown to be involved in several human diseases, such as Type-II diabetes and neurodegenerative diseases. In this work, we employed a multi-disciplinary approach to study the activation mechanism of the CDF protein family. For this we used MamM, one of the main ion transporters of magnetosomes--bacterial organelles that enable magnetotactic bacteria to orientate along geomagnetic fields. Our results reveal that the cytosolic domain of MamM forms a stable dimer that undergoes distinct conformational changes upon divalent cation binding. MamM conformational change is associated with three metal binding sites that were identified and characterized. Altogether, our results provide a novel auto-regulation mode of action model in which the cytosolic domain's conformational changes upon ligand binding allows the priming of the CDF into its transport mode.