Lcn2 is a host defense protein induced
via the innate immune response
to sequester iron-loaded bacterial siderophores. However, excess or
prolonged elevation of Lcn2 levels can induce adverse cellular effects,
including oxidative stress and inflammation. In this work, we use
Hydrogen–Deuterium eXchange (HDX) and Isothermal Titration
Calorimetry (ITC) to characterize the binding interaction between
Lcn2 and siderophores enterobactin and 2,3-DHBA, in the presence and
absence of iron. Our results indicate a rare “Type II”
interaction in which binding of siderophores drives the protein conformational
equilibrium toward an unfolded state. Linking our molecular model
to cellular assays, we demonstrate that this “distorted binding
mode” facilitates a deleterious cellular accumulation of reactive
oxygen species that could represent the molecular origin of Lcn2 pathology.
These results add important insights into mechanisms of Lcn2 action
and have implications in Lcn2-mediated effects including inflammation.
A major hallmark of Alzheimer's disease (AD) is the aggregation of amyloid-β peptides in the brains of people afflicted by the disease. The exact pathway to this catastrophic event is unknown. In this work, a total of 9.5 μs molecular dynamics simulations have been performed to investigate the structure and dynamics of the smallest form of toxic Aβ oligomers, i.e., the Aβ dimers. This study suggests that specific hydrophobic regions are vital in the aggregation process. Different possible structures for Aβ dimers are reported along with their relative binding affinity. These data may be used to design better Aβ-aggregation inhibitors. The diversity of the dimer structures suggests several aggregation pathways.
Neurodegeneration in Alzheimer’s disease (AD) is defined by pathology featuring amyloid-β (Aβ) deposition in the brain. Aβ monomers themselves are generally considered to be nontoxic, but misfold into β-sheets and aggregate to form neurotoxic oligomers. One suggested strategy to treat AD is to prevent the formation of toxic oligomers. The SG inhibitors are a class of pseudopeptides designed and optimized using molecular dynamics (MD) simulations for affinity to Aβ and experimentally validated for their ability to inhibit amyloid-amyloid binding using single molecule force spectroscopy (SMFS). In this work, we provide a review of our previous MD and SMFS studies of these inhibitors and present new cell viability studies that demonstrate their neuroprotective effects against Aβ(1–42) oligomers using mouse hippocampal-derived HT22 cells. Two of the tested SG inhibitors, predicted to bind Aβ in anti-parallel orientation, demonstrated neuroprotection against Aβ(1–42). A third inhibitor, predicted to bind parallel to Aβ, was not neuroprotective. Myristoylation of SG inhibitors, intended to enhance delivery across the blood-brain barrier (BBB), resulted in cytotoxicity. This is the first use of HT22 cells for the study of peptide aggregation inhibitors. Overall, this work will inform the future development of peptide aggregation inhibitors against Aβ toxicity.
The region encompassing residues 13–23 of the amyloid beta peptide (Aβ(13–23)) of Alzheimer’s disease is the self-recognition site that initiates toxic oligomerization and fibrillization and also is the site of interaction of Aβ with many other proteins. We describe herein a study by molecular dynamics of the complexes formed by R (= N-AcAβ(13–23)NH2(N-CH3C(O)HHQKLVFFAEDNH2)) with several pseudopeptides designed to form β-sheets with Aβ(1-40,42) and prevent oligomer and fibril formation. Adhesion to both edges of the R β-strand is examined by structure analysis. Umbrella sampling along a dissociation pathway reveals approximate free energies of binding in the submicromolar range. One of the three pseudopeptides binds strongly to one edge of the R β-strand and another to the opposite edge, while the third displays strong binding to both edges. It is desirable to block both edges of the self-recognition site of Aβ to prevent oligomer formation. The study reveals that this may be accomplished by a single pseudopeptide or two in combination. Thus the pseudopeptides, used singly or in pairs, may be competitive inhibitors of Aβ oligomerization at stoichiometric concentrations.
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