Development of a GC-376 Based Peptidomimetic PROTAC as a Degrader of 3-Chymotrypsin-like Protease of SARS-CoV-2.

We have applied a proteolysis targeting chimera (PROTAC) technology to obtain a peptidomimetic molecule able to trigger the degradation of SARS-CoV-2 3-chymotrypsin-like protease (3CL). The PROTAC molecule was designed by conjugating a GC-376 based dipeptidyl 3CL ligand to a pomalidomide moiety through a piperazine-piperidine linker. NMR and crystallographic data complemented with enzymatic and cellular studies showed that (i) the dipeptidyl moiety of PROTAC binds to the active site of the dimeric state of SARS-CoV-2 3CL forming a reversible covalent bond with the sulfur atom of catalytic Cys145, (ii) the linker and the pomalidomide cereblon-ligand of PROTAC protrude from the protein, displaying a high degree of flexibility and no interactions with other regions of the protein, and (iii) PROTAC reduces the protein levels of SARS-CoV-2 3CL in cultured cells.

Espresso Coffee Mitigates the Aggregation and Condensation of Alzheimer's Associated Tau Protein.

Espresso coffee is among the most consumed beverages in the world. Recent studies report a protective activity of the coffee beverage against neurodegenerative disorders such as Alzheimer's disease. Alzheimer's disease belongs to a group of disorders, called tauopathies, which are characterized by the intraneuronal accumulation of the microtubule-associated protein tau in fibrillar aggregates.

Ultrasmall Gold Nanoparticles as Clients of Biomolecular Condensates.

Liquid-liquid phase separation (LLPS) of biopolymers to form condensates is a widespread phenomenon in living cells. Agents that target or alter condensation can help uncover elusive physiological and pathological mechanisms. Owing to their unique material properties and modes of interaction with biomolecules, nanoparticles represent attractive condensate-targeting agents.

Site-directed double monoubiquitination of the repeat domain of the amyloid-forming protein tau impairs self-assembly and coacervation.

In Alzheimer's disease and related disorders called tauopathies, the microtubule-associated protein tau accumulates in the brain in the form of amyloid-like supramolecular filaments. As an intrinsically disordered protein, tau undergoes many post-translational modifications, including ubiquitination. Alterations to the levels of ubiquitination of tau have been observed at various stages of neurodegenerative conditions.

New Paradigm for Nano-Bio Interactions: Multimolecular Assembly of a Prototypical Disordered Protein with Ultrasmall Nanoparticles.

Understanding the interactions between nanoparticles (NPs) and proteins is crucial for the successful application of NPs in biological contexts. Protein adsorption is dependent on particle size, and protein binding to ultrasmall (1-3 nm) NPs is considered to be generally weak. However, most studies have involved structured biomacromolecules, while the interactions of ultrasmall NPs with intrinsically disordered proteins (IDPs) have remained elusive.

Camouflaged Fluorescent Silica Nanoparticles Target Aggregates and Condensates of the Amyloidogenic Protein Tau.

Intrinsically disordered proteins (IDPs) are increasingly found to be associated with irreversible neurodegenerative disorders. The protein tau is a prototypical IDP whose abnormal aggregation into insoluble filaments is a major hallmark of Alzheimer's disease. The view has emerged that aggregation may proceed via alternative pathways involving oligomeric intermediates or phase-separated liquid droplets.

Alterations in calmodulin-cardiac ryanodine receptor molecular recognition in congenital arrhythmias.

Calmodulin (CaM), a ubiquitous and highly conserved Ca-sensor protein involved in the regulation of over 300 molecular targets, has been recently associated with severe forms of lethal arrhythmia. Here, we investigated how arrhythmia-associated mutations in CaM localized at the C-terminal lobe alter the molecular recognition with Ryanodine receptor 2 (RyR2), specifically expressed in cardiomyocytes. We performed an extensive structural, thermodynamic, and kinetic characterization of the variants D95V/H in the EF3 Ca-binding motif and of the D129V and D131H/E variants in the EF4 motif, and probed their interaction with RyR2.

Ubiquitination of Alzheimer's-related tau protein affects liquid-liquid phase separation in a site- and cofactor-dependent manner.

The formation of biomolecular condensates has emerged as a crucial player both in neuronal physiology and neurodegeneration. Phase separation of the Alzheimer's related protein tau into liquid condensates is facilitated by polyanions and is regulated by post-translational modifications. Given the central role of ubiquitination in proteostasis regulation and signaling, we investigated the behavior of monoubiquitinated tau during formation of condensates.

Semisynthetic Modification of Tau Protein with Di-Ubiquitin Chains for Aggregation Studies.

Ubiquitin, a protein modifier that regulates diverse essential cellular processes, is also a component of the protein inclusions characteristic of many neurodegenerative disorders. In Alzheimer's disease, the microtubule associated tau protein accumulates within damaged neurons in the form of cross-beta structured filaments. Both mono- and polyubiquitin were found linked to several lysine residues belonging to the region of tau protein that forms the structured core of the filaments.

Unsaturated Fatty Acid-Induced Conformational Transitions and Aggregation of the Repeat Domain of Tau.

Background: The intrinsically disordered, amyloidogenic protein Tau associates with diverse classes of molecules, including proteins, nucleic acids, and lipids. Mounting evidence suggests that fatty acid molecules could play a role in the dysfunction of this protein, however, their interaction with Tau remains poorly characterized.

Methods: In a bid to elucidate the association of Tau with unsaturated fatty acids at the sub-molecular level, we carried out a variety of solution NMR experiments in combination with circular dichroism and fluorescence measurements.

Dynamic molecular exchange and conformational transitions of alpha-synuclein at the nano-bio interface.

The notion that nanoscale surfaces influence protein conformational transitions stimulates the investigation of fibrillogenic polypeptides adsorbing to nanomaterials. Alpha-synuclein (αS) is a prototypical amyloidogenic protein whose aggregation is associated with severe neurodegenerative disorders. We explored the interaction of αS with silica nanoparticles (SNPs) in diverse solution conditions, ranging from protein-free to protein-rich media.