Biology subjects

Martins, P. M.

Publications and source records attributed to Martins, P. M..

5 recordsLinked to original sources

bioRxiv2025

Allosteric Modulation of Pathological Ataxin-3 Aggregation: A Path to Spinocerebellar Ataxia Type-3 Therapies

Spinocerebellar ataxia type 3 (SCA3) is a rare inherited neurodegenerative disease caused by the expansion of a polyglutamine repeat in the protease ataxin-3 (Atx3). Despite extensive knowledge of the downstream pathophysiology, no disease-modifying therapies are currently available to halt disease progression. The accumulation of protein inclusions enriched in the polyQ-expanded Atx3 in neurons suggests that inhibiting its self-assembly may yield targeted therapeutic approaches. Here it is shown that a supramolecular tweezer, CLR01, binds to a lysine residue on a positively charged surface patch of the Atx3 catalytic Josephin domain. At this site, the binding of CLR01 decreases the conformational fluctuations of the distal flexible hairpin. This results in reduced exposure of the nearby aggregation-prone region, which overlaps with the substrate ubiquitin binding site and primes Atx3 self-assembly, ultimately delaying Atx3 amyloid fibril formation and reducing the secondary nucleation rate, a process linked to fibril proliferation and toxicity. These effects translate into the reversal of synapse loss in a SCA3 cultured cortical neuron model, an improved locomotor function in a C. elegans SCA3 model, and a delay in disease onset, accompanied by reduced severity of motor symptoms in a SCA3 mouse model. This study provides critical insights into Atx3 self-assembly, revealing a novel allosteric site for designing CLR01-inspired therapies targeting pathological aggregation pathways while sparing essential functional sites. These findings emphasize that targeting allosteric sites in amyloid-forming proteins may offer unique opportunities to develop safe therapeutic strategies for various protein misfolding disorders.

biophysics↗

bioRxiv2023

Transthyretin has conformation-selective proteolytic activity against α-synuclein

Transthyretin (TTR) is a plasma protein known as a transporter of thyroxine and retinol but also can inhibit the formation of amyloid {beta}-peptide (A{beta}) fibrils and catalyze the proteolysis of apolipoprotein A-I and A{beta}. Here, recombinant TTR is shown to have proteolytic activity against specific conformations of -synuclein (aSyn), a protein that accumulates in intraneuronal inclusions characteristic of Parkinsons disease (PD). Our discovery stemmed from the observation of a marked decrease in aSyn aggregation in the presence of submicromolar concentrations of TTR. Using integrated biophysical techniques, including advanced microscopy and mass spectrometry, we found that aSyn aggregation is prevented due to the proteolytic cleavage of free aSyn by TTR, in a reaction that is slower for S-glutathionylated TTR and faster for TTR preparations containing vestigial amounts of [~]70 kDa TTR oligomers (oTTR). Interestingly, this proteolysis-trigger effect is not unique to oTTR since it can also be induced by Teflon and glass surfaces independently of which TTR preparation is tested. Our results indicate that a change in aSyn conformation must precede the proteolysis step. The proteolysis of misfolded aSyn emerges as a possible TTR function with implications for the understanding of different neurodegenerative disorders.

biochemistry↗

bioRxiv2022

Drug repurposing of dopaminergic drugs to inhibit Ataxin-3 aggregation

The accumulation of mutant ataxin-3 (Atx3) in neuronal nuclear inclusions is a pathological hallmark of Machado-Joseph disease (MJD), also known as Spinocerebellar Ataxia Type 3. Decreasing the protein aggregation burden is a possible disease-modifying strategy to tackle MJD and other neurodegenerative disorders for which only symptomatic treatments are currently available. We performed a drug repurposing screening to identify inhibitors of Atx3 aggregation with known toxicological and pharmacokinetic profiles. Interestingly, dopamine hydrochloride and other catecholamines are among the most potent inhibitors of Atx3 aggregation in vitro. Our results indicate that low micromolar concentrations of dopamine markedly delay the formation of mature amyloid fibrils of mutant Atx3 through the inhibition of the earlier oligomerization steps. Although dopamine itself does not pass the blood-brain barrier, dopamine levels in the brain can be increased by low doses of dopamine precursors and dopamine agonists commonly used to treat Parkinsonian symptoms. These findings disclose a possible application of dopaminergic drugs to halt or reduce Atx3 accumulation in the brains of MJD patients.

neuroscience↗

bioRxiv2022

Quantification of surface tension effects and nucleation-and-growth rates during self-assembly of biological condensates

Liquid-solid and liquid-liquid phase separation (PS) drives the formation of functional and disease-associated biological assemblies. Principles of phase equilibrium are here employed to derive a general kinetic solution that predicts the evolution of the mass and size of biological assemblies. Thermodynamically, protein PS is determined by two measurable concentration limits: the saturation concentration and the critical solubility. Due to surface tension effects, the critical solubility can be higher than the saturation concentration for small, curved nuclei. Kinetically, PS is characterized by the primary nucleation rate constant and a combined rate constant accounting for growth and secondary nucleation. It is demonstrated that the formation of a few number of large condensates is possible without active mechanisms of size control and in the absence of coalescence phenomena. Our exact analytical solution can be used to interrogate how the elementary steps of PS are affected by candidate drugs.

biophysics↗

bioRxiv2019

A simple linearization method unveils hidden enzymatic assay interferences

Enzymes are among the most important drug targets in the pharmaceutical industry. The bioassays used to screen enzyme modulators can be affected by unaccounted interferences such as time-dependent inactivation and inhibition effects. Using procaspase-3, caspase-3, and -thrombin as model enzymes, we show that some of these effects are not eliminated by merely ignoring the reaction phases that follow initial-rate measurements. We thus propose a linearization method (LM) for detecting spurious changes of enzymatic activity based on the representation of progress curves in modified coordinates. This method is highly sensitive to signal readout distortions, thereby allowing rigorous selection of valid kinetic data. The method allows the detection of assay interferences even when their occurrence is not suspected a priori. By knowing the assets and liabilities of the bioassay, enzymology results can be reported with enhanced reproducibility and accuracy. Critical analysis of full progress curves is expected to help discriminating experimental artifacts from true mechanisms of enzymatic inhibition.

biophysics↗