Browsing by Subject "Cryo-EM"

Now showing 1 - 4 of 4
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    Cryo-EM Structures of Chronic Traumatic Encephalopathy Tau Filaments with PET Ligand Flortaucipir
    (Elsevier, 2023) Shi, Yang; Ghetti, Bernardino; Goedert, Michel; Scheres, Sjors H. W.; Pathology and Laboratory Medicine, School of Medicine
    Positron emission tomography (PET) imaging allows monitoring the progression of amyloid aggregation in the living brain. [18F]-Flortaucipir is the only approved PET tracer compound for the visualisation of tau aggregation. Here, we describe cryo-EM experiments on tau filaments in the presence and absence of flortaucipir. We used tau filaments isolated from the brain of an individual with Alzheimer's disease (AD), and from the brain of an individual with primary age-related tauopathy (PART) with a co-pathology of chronic traumatic encephalopathy (CTE). Unexpectedly, we were unable to visualise additional cryo-EM density for flortaucipir for AD paired helical or straight filaments (PHFs or SFs), but we did observe density for flortaucipir binding to CTE Type I filaments from the case with PART. In the latter, flortaucipir binds in a 1:1 molecular stoichiometry with tau, adjacent to lysine 353 and aspartate 358. By adopting a tilted geometry with respect to the helical axis, the 4.7 Å distance between neighbouring tau monomers is reconciled with the 3.5 Å distance consistent with π-π-stacking between neighbouring molecules of flortaucipir.
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    Cryo-EM structures of cotton wool plaques' amyloid β and of tau filaments in dominantly inherited Alzheimer disease
    (Springer, 2024-08-15) Hoq, Md Rejaul; Fernandez, Anllely; Vago, Frank S.; Hallinan, Grace I.; Bharath, Sakshibeedu R.; Li, Daoyi; Ozcan, Kadir A.; Garringer, Holly J.; Jiang, Wen; Vidal, Ruben; Ghetti, Bernardino; Pathology and Laboratory Medicine, School of Medicine
    Cotton wool plaques (CWPs) have been described as features of the neuropathologic phenotype of dominantly inherited Alzheimer disease (DIAD) caused by some missense and deletion mutations in the presenilin 1 (PSEN1) gene. CWPs are round, eosinophilic amyloid-β (Aβ) plaques that lack an amyloid core and are recognizable, but not fluorescent, in Thioflavin S (ThS) preparations. Amino-terminally truncated and post-translationally modified Aβ peptide species are the main component of CWPs. Tau immunopositive neurites may be present in CWPs. In addition, neurofibrillary tangles coexist with CWPs. Herein, we report the structure of Aβ and tau filaments isolated from brain tissue of individuals affected by DIAD caused by the PSEN1 V261I and A431E mutations, with the CWP neuropathologic phenotype. CWPs are predominantly composed of type I Aβ filaments present in two novel arrangements, type Ic and type Id; additionally, CWPs contain type I and type Ib Aβ filaments. Tau filaments have the AD fold, which has been previously reported in sporadic AD and DIAD. The formation of type Ic and type Id Aβ filaments may be the basis for the phenotype of CWPs. Our data are relevant for the development of PET imaging methodologies to best detect CWPs in DIAD.
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    Structural Basis of Arrestin Binding to Cell Membranes
    (2024-04) Miller, Kyle Warren; Chen, Qiuyan; Takagi, Yuichiro; Georgiadis, Millie M.; Hurley, Thomas D.
    Two non-visual arrestins, arrestin2 (Arr2) and arrestin3 (Arr3), selectively interact with activated and phosphorylated G protein-coupled receptors (GPCRs) and play crucial roles in regulating many important physiological processes. Arrestins also engage the lipid bilayer surrounding activated GPCRs, which further potentiates arrestin activation and regulates GPCR trafficking in cells. Because of this, structural and functional understanding of arrestins would provide insight in enhancing arrestin’s GPCR desensitization for various diseases where constitutively active GPCR mutants play a role including congenital endocrine disorders and familial gestational hyperthyroidism. To better understand the membrane binding role of arrestins, we performed in vitro binding assays and demonstrated that Arr2 selectively binds to nanodiscs containing Phosphatidylinositol 4,5-bisphosphate (PIP2) even in the absence of different binding sites. Our cryo-electron microscopy (Cryo-EM) structure of Arr2 in complex with PIP2 nanodisc reveals that multiple structural elements of Arr2, including the finger loop, C domain and C-edge loop, contribute to membrane binding. Eliminating one individual site does not significantly impact Arr2 binding to the nanodisc. Moreover, a preactivated variant of Arr2 shows increased binding to the nanodisc than wildtype. We also labeled four potential membrane binding sites with monobromobimane (mBrB) and detected different levels of fluorescence increase in the presence of nanodisc containing various types of phospholipids. Overall, our study provides detailed structural evidence on how arrestins engage the membrane via multiple contact points and how this can impact arrestin-mediated signaling.
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    The structural mechanism of human glycogen synthesis by the GYS1-GYG1 complex
    (Elsevier, 2022) Fastman, Nathan M.; Liu, Yuxi; Ramanan, Vyas; Merritt, Hanne; Ambing, Eileen; DePaoli-Roach, Anna A.; Roach, Peter J.; Hurley, Thomas D.; Mellem, Kevin T.; Ullman, Julie C.; Green, Eric; Morgans, David, Jr.; Tzitzilonis, Christos; Biochemistry and Molecular Biology, School of Medicine
    Glycogen is the primary energy reserve in mammals, and dysregulation of glycogen metabolism can result in glycogen storage diseases (GSDs). In muscle, glycogen synthesis is initiated by the enzymes glycogenin-1 (GYG1), which seeds the molecule by autoglucosylation, and glycogen synthase-1 (GYS1), which extends the glycogen chain. Although both enzymes are required for proper glycogen production, the nature of their interaction has been enigmatic. Here, we present the human GYS1:GYG1 complex in multiple conformations representing different functional states. We observe an asymmetric conformation of GYS1 that exposes an interface for close GYG1 association, and propose this state facilitates handoff of the GYG1-associated glycogen chain to a GYS1 subunit for elongation. Full activation of GYS1 widens the GYG1-binding groove, enabling GYG1 release concomitant with glycogen chain growth. This structural mechanism connecting chain nucleation and extension explains the apparent stepwise nature of glycogen synthesis and suggests distinct states to target for GSD-modifying therapeutics.