Mechanical properties of BiP protein determined by nano‐rheology

Casanova-Morales, Nathalie; Quiroga-Roger, Diego; Alfaro-Valdés, Hilda M.; Alavi, Zahra; Lagos‐Espinoza, Miguel I.A.; Zocchi, Giovanni; Wilson, Christian A.M.

doi:10.1002/pro.3432

Cited by 7 publications

(10 citation statements)

References 34 publications

(93 reference statements)

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“…It is important to mention that structurally all nucleotides bind similarly to BiP based on docking simulations performed in this work at room temperature (Figure S6). The latter is supported with previous results from our lab obtained with nano-rheology (Casanova-Morales et al, 2018), as well as other studies (Wei & Hendershot, 1995). Interestingly, it has been reported that the affinity of nucleotides for the heat shock cognate 70 (Hsc70, member of the Hsp70 family) is modified around five times by changes in a range of temperatures similar to our work (Buxbaum & Woodman, 1996;Palleros et al, 1991).…”

Section: Discussionsupporting

confidence: 93%

Effect of temperature and nucleotide on the binding of BiP chaperone to a protein substrate

et al. 2023

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BiP (immunoglobulin heavy-chain binding protein) is a Hsp70 monomeric ATPase motor that plays broad and crucial roles in maintaining proteostasis inside the cell. Structurally, BiP is formed by two domains, a nucleotidebinding domain (NBD) with ATPase activity connected by a flexible hydrophobic linker to the substrate-binding domain. While the ATPase and substrate binding activities of BiP are allosterically coupled, the latter is also dependent on nucleotide binding. Recent structural studies have provided new insights into BiP's allostery; however, the influence of temperature on the coupling between substrate and nucleotide binding to BiP remains unexplored. Here, we study BiP's binding to its substrate at the single molecule level using thermoregulated optical tweezers which allows us to mechanically unfold the client protein and explore the effect of temperature and different nucleotides on BiP binding. Our results confirm that the affinity of BiP for its protein substrate relies on nucleotide binding, by mainly regulating the binding kinetics between BiP and its substrate. Interestingly, our findings also showed that the apparent affinity of BiP for its protein substrate in the presence of nucleotides remains invariable over a wide range of temperatures, suggesting that BiP may interact with its client proteins with similar affinities even when the temperature is not optimal. Thus, BiP could play a role as a "thermal buffer" in proteostasis.Maira Rivera and Francesca Burgos-Bravo contributed equally to this study.

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Section: Discussionsupporting

confidence: 93%

Effect of temperature and nucleotide on the binding of BiP chaperone to a protein substrate

et al. 2023

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“…The structural reason is because the lid is close [6]. Additionally, it was observed in presence of peptide the dissociation constant (KD) for ADP decreased 1000 times, demonstrating that peptide binding dramatically increases the affinity for ADP which evidences the allosteric coupling between SBD and NBD domains [66].…”

Section: Mechanical Aspectsmentioning

confidence: 89%

Mechanical Properties of Chaperone BiP, the Master Regulator of the Endoplasmic Reticulum

Alfaro-Valdés¹,

Burgos‐Bravo²,

Casanova-Morales³

et al. 2019

Endoplasmic Reticulum

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Immunoglobulin heavy-chain-binding protein (BiP protein) is a 75-kDa Hsp70 monomeric ATPase motor that plays broad and crucial roles maintaining proteostasis inside the cell. Its malfunction has been related with the appearance of many and important health problems such as neurodegenerative diseases, cancer, and heart diseases, among others. In particular, it is involved in many endoplasmic reticulum (ER) processes and functions, such as protein synthesis, folding, and assembly, and also it works in the posttranslational mechanism of protein translocation. However, it is unknown what kind of molecular motor BiP works like, since the mechanochemical mechanism that BiP utilizes to perform its work during posttranslational translocation across the ER is not fully understood. One novel approach to study both structural and catalytic properties of BiP considers that the viscoelastic regime behavior of the enzymes (considering them as a spring) and their mechanical properties are correlated with catalysis and ligand binding. Structurally, BiP is formed by two domains, and to establish a correlation between BiP structure and catalysis and how its conformational and viscoelastic changes are coupled to ligand binding, catalysis, and allosterism (information transmitted between the domains), optical tweezers and nano-rheology techniques have been essential in this regard.

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“…The activity of PDI has been linked to interactions with ubiquilin [58], thus allowing the uninhibited lysosomal protein degradation [59]. GRP78/BiP is another pivotal protein in the chaperone function [60,61] of the ER that was analyzed. The preservation of the observed 60 KDa fragment by the PDT-treated cells was indicative of degradation of functional GRP78/BiP, but also of the presence of ATP [61] displaying priming of oncolytic activity.…”

Section: Discussionmentioning

confidence: 99%

Metalloporphyrin Pd(T4) Exhibits Oncolytic Activity and Cumulative Effects with 5-ALA Photodynamic Treatment against C918 Cells

Leviskas

Vályi-Nagy

Munirathinam

et al. 2020

IJMS

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Photodynamic therapy is a non-invasive method where light activates a photosensitizer bound to cancer cells, generating reactive oxygen species and resulting in cell death. This study assessed the oncolytic potential of photodynamic therapy, comparing European Medicines Agency and United States Food and Drug Administration-approved 5-aminolevulinic acid (5-ALA) to a metalloporphyrin, Pd(T4), against a highly invasive uveal melanoma cell line (C918) in two- and three-dimensional models in vitro. Epithelial monolayer studies displayed strong oncolytic effects (>70%) when utilizing Pd(T4) at a fraction of the concentration, and reduced pre-illumination time compared to 5-ALA post-405 nm irradiance. When analyzed at sub-optimal concentrations, application of Pd(T4) and 5-ALA with 405 nm displayed cumulative effects. Lethality from Pd(T4)-photodynamic therapy was maintained within a three-dimensional model, including the more resilient vasculogenic mimicry-forming cells, though at lower rates. At high concentrations, modality of cell death exhibited necrosis partially dependent on reactive oxygen species. However, sub-optimal concentrations of photosensitizer exhibited an apoptotic protein expression profile characterized by increased Bax/Bcl-2 ratio and endoplasmic stress-related proteins, along with downregulation of apoptotic inhibitors CIAP-1 and -2. Together, our results indicate Pd(T4) as a strong photosensitizer alone and in combination with 5-ALA against C918 cells.

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Mechanical properties of BiP protein determined by nano‐rheology

Cited by 7 publications

References 34 publications

Effect of temperature and nucleotide on the binding of BiP chaperone to a protein substrate

Effect of temperature and nucleotide on the binding of BiP chaperone to a protein substrate

Mechanical Properties of Chaperone BiP, the Master Regulator of the Endoplasmic Reticulum

Metalloporphyrin Pd(T4) Exhibits Oncolytic Activity and Cumulative Effects with 5-ALA Photodynamic Treatment against C918 Cells

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