Compounds harboring high acidity and oxidizability of thiol groups permit tuning the redox equilibrium constants of CxxC sites of members of the protein disulphide isomerase (PDI) family and thus can ...be used to accelerate folding processes and increase the production of native proteins by minimal loading in comparison to glutathione.
Compounds harboring high acidity and oxidizability of thiol groups permit tuning the redox equilibrium constants of CxxC sites of the PDI family enzymes and thus can be used to promote oxidative folding and increase the native protein productions by minimal loading as compared to glutathione.
Malignant pleural mesothelioma (MPM) is an aggressive malignant tumor of mesothelial origin associated with asbestos exposure. MPM has a limited response to conventional chemotherapy and ...radiotherapy, thus early diagnosis of MPM is extremely critical. CT scans have limited accuracy in the differentiation between benign and malignant pleural disease. Several studies have reported that 18-fluorodeoxyglucose (FDG) positron emission tomography (PET) plays an important role in the assessment of thoracic malignancy such as lung cancer. Here, we investigated the clinical utility of PET in patients with MPM. The maximum SUV (SUVmax) of 18F-FDG was measured in 47 MPM patients and 29 non-MPM patients including those with pleural thickening. We demonstrated that patients with MPM had significantly higher SUVmax levels than a population with non-malignant pleural disease. The Kaplan-Meier method revealed significant differences in overall survival between groups with SUVmax levels lower and higher than the assumed cut-off. Our data suggest that SUVmax levels are useful as an aid for diagnosis and prognosis of MPM.
The misfolding, amyloid aggregation, and fibril formation of intrinsically disordered proteins/peptides (or amyloid proteins) have been shown to cause a number of disorders. The underlying mechanisms ...of amyloid fibrillation and structural properties of amyloidogenic precursors, intermediates, and amyloid fibrils have been elucidated in detail; however, in-depth examinations on physiologically relevant contributing factors that induce amyloidogenesis and lead to cell death remain challenging. A large number of studies have attempted to characterize the roles of biomembranes on protein aggregation and membrane-mediated cell death by designing various membrane components, such as gangliosides, cholesterol, and other lipid compositions, and by using various membrane mimetics, including liposomes, bicelles, and different types of lipid-nanodiscs.
We herein review the dynamic effects of membrane curvature on amyloid generation and the inhibition of amyloidogenic proteins and peptides, and also discuss how amyloid formation affects membrane curvature and integrity, which are key for understanding relationships with cell death. Small unilamellar vesicles with high curvature and large unilamellar vesicles with low curvature have been demonstrated to exhibit different capabilities to induce the nucleation, amyloid formation, and inhibition of amyloid-β peptides and α-synuclein. Polymorphic amyloidogenesis in small unilamellar vesicles was revealed and may be viewed as one of the generic properties of interprotein interaction-dominated amyloid formation. Several mechanical models and phase diagrams are comprehensively shown to better explain experimental findings. The negative membrane curvature-mediated mechanisms responsible for the toxicity of pancreatic β cells by the amyloid aggregation of human islet amyloid polypeptide (IAPP) and binding of the precursors of the semen-derived enhancer of viral infection (SEVI) are also described. The curvature-dependent binding modes of several types of islet amyloid polypeptides with high-resolution NMR structures are also discussed.
•The relationships between membrane curvature and amyloid aggregation were surveyed.•SUV promoted amyloidogenesis and LUV induced amorphous-like amyloids of Aβ(1–40).•SUV caused polymorphic Aβ(1–42) amyloid formation and LUV blocked amyloid generation.•The initial structures of αSN in synaptic mimic SUV are key to its amyloid fibrillation.•IAPP aggregation and the SEVI precursor affected curvature and resulting cytotoxicity.
We report the first example of a synthetic thiol-based compound that promotes oxidative protein folding upon 1-equivalent loading to the disulfide bonds in the client protein to afford the native ...form in over 70% yield.
-Methylation is a central post-translational processing of proteins
for regulating functions including chaperone activities. Despite the universally observed biochemical reactions in nature,
-methylation has hardly been utilized in the design, functionalization, and switching of synthetic bioregulatory agents, particularly folding promotors. As a biomimetic approach, we developed pyridinylmethanethiols to investigate the effects of
-methylation on the promotion of oxidative protein folding. For a comprehensive study on the geometrical effects, constitutional isomers of pyridinylmethanethiols with
-,
-, and
-substitutions have been synthesized. Among the constitutional isomers,
-substituted pyridinylmethanethiol showed the fastest disulfide-bond formation of the client proteins to afford the native forms most efficiently.
-Methylation drastically increased the acidity and enhanced the oxidizability of the thiol groups in the pyridinylmethanethiols to enhance the folding promotion efficiencies. Among the isomers,
-substituted
-methylated pyridinylmethanethiol accelerated the oxidative protein folding reactions with the highest efficiency, allowing for protein folding promotion by 1-equivalent loading as a semi-enzymatic activity. This study will offer a novel bioinspired molecular design of synthetic biofunctional agents that are semi-enzymatically effective for the promotion of oxidative protein folding including biopharmaceuticals such as insulin
by minimum loading.
We report the first example of a synthetic thiol-based compound that promotes oxidative protein folding upon 1-equivalent loading to the disulfide bonds in the client protein to afford the native ...form in over 70% yield.
N
-Methylation is a central post-translational processing of proteins
in vivo
for regulating functions including chaperone activities. Despite the universally observed biochemical reactions in nature,
N
-methylation has hardly been utilized in the design, functionalization, and switching of synthetic bioregulatory agents, particularly folding promotors. As a biomimetic approach, we developed pyridinylmethanethiols to investigate the effects of
N
-methylation on the promotion of oxidative protein folding. For a comprehensive study on the geometrical effects, constitutional isomers of pyridinylmethanethiols with
ortho
-,
meta
-, and
para
-substitutions have been synthesized. Among the constitutional isomers,
para
-substituted pyridinylmethanethiol showed the fastest disulfide-bond formation of the client proteins to afford the native forms most efficiently.
N
-Methylation drastically increased the acidity and enhanced the oxidizability of the thiol groups in the pyridinylmethanethiols to enhance the folding promotion efficiencies. Among the isomers,
para
-substituted
N
-methylated pyridinylmethanethiol accelerated the oxidative protein folding reactions with the highest efficiency, allowing for protein folding promotion by 1-equivalent loading as a semi-enzymatic activity. This study will offer a novel bioinspired molecular design of synthetic biofunctional agents that are semi-enzymatically effective for the promotion of oxidative protein folding including biopharmaceuticals such as insulin
in vitro
by minimum loading.
We report the first example of a synthetic thiol-based compound that promotes oxidative protein folding upon 1-equivalent loading to the disulfide bonds in the client protein to afford the native form in over 70% yield.
Proteins form native structures through folding processes, many of which proceed through intramolecular hydrophobic effect, hydrogen bond and disulfide-bond formation. In vivo , protein aggregation ...is prevented even in the highly condensed milieu of a cell through folding mediated by molecular chaperones and oxidative enzymes. Chemical approaches to date have not replicated such exquisite mediation. Oxidoreductases efficiently promote folding by the cooperative effects of oxidative reactivity for disulfide-bond formation in the client unfolded protein and chaperone activity to mitigate aggregation. Conventional synthetic folding promotors mimic the redox-reactivity of thiol/disulfide units but do not address client-recognition units for inhibiting aggregation. Herein, we report thiol/disulfide compounds containing client-recognition units, which act as synthetic oxidoreductase-mimics. For example, compound βCD W SH/SS bears a thiol/disulfide unit at the wide rim of β-cyclodextrin as a client recognition unit. βCD W SH/SS shows promiscuous binding to client proteins, mitigates protein aggregation, and accelerates disulfide-bond formation. In contrast, positioning a thiol/disulfide unit at the narrow rim of β-cyclodextrin promotes folding less effectively through preferential interactions at specific residues, resulting in aggregation. The combination of promiscuous client-binding and redox reactivity is effective for the design of synthetic folding promoters. βCD W SH/SS accelerates oxidative protein folding at highly condensed sub-millimolar protein concentrations.
P5, also known as PDIA6, is a PDI family member involved in the ER quality control. Here, we revealed that P5 dimerizes via a unique adhesive motif contained in the N-terminal thioredoxin-like ...domain. Unlike conventional leucine zipper motifs with leucine residues every two helical turns on ∼30-residue parallel α helices, this adhesive motif includes periodic repeats of leucine/valine residues at the third or fourth position spanning five helical turns on 15-residue anti-parallel α helices. The P5 dimerization interface is further stabilized by several reciprocal salt bridges and C-capping interactions between protomers. A monomeric P5 mutant with the impaired adhesive motif showed structural instability and local unfolding, and behaved as aberrant proteins that induce the ER stress response. Disassembly of P5 to monomers compromised its ability to inactivate IRE1α via intermolecular disulfide bond reduction and its Ca2+-dependent regulation of chaperone function in vitro. Thus, the leucine-valine adhesive motif supports structure and function of P5.
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•P5 dimerizes via a unique leucine-valine adhesive motif in the first Trx-like domain•The adhesive motif is radically different from conventional coiled-coil motifs•The dimeric structure serves to promote P5-mediated inactivation of IRE1α•Ca2+-binding regions and Ca2+-dependent functional regulation were clarified for P5
Using SAXS and X-ray crystallography, Okumura et al. determined the structure of a protein disulfide isomerase P5, in which the N-terminal thioredoxin-like domain dimerizes via a unique leucine-valine adhesive motif. A monomeric P5 mutant displayed structural instability and compromised functions, indicating the structural and functional essentiality of this motif.
Malignant pleural mesothelioma (MPM) is an aggressive malignant tumor of mesothelial origin that shows a limited response to conventional chemotherapy and radiotherapy. Therefore, diagnosing MPM ...early is very important. Some researchers have previously reported that high-mobility group box 1 (HMGB1) was correlated with pulmonary fibrosis. MPM involves the malignant transformation of mesothelial cells, which originate from mesenchymal cells similar to lung fibroblasts. Here, we investigated serum levels of HMGB1 in patients with MPM and compared them with those of a population that had been exposed to asbestos without developing MPM.
HMGB1 production from MPM cell lines was measured using ELISA. Serum HMGB1 levels were also examined in 61 MPM patients and 45 individuals with benign asbestos-related diseases.
HMGB1 concentrations of 2 out of 4 MPM cell lines were higher than that of normal mesothelial cell line, Met-5A. We demonstrated that patients with MPM had significantly higher serum levels of HMGB1 than the population who had been exposed to asbestos but had not developed MPM. The difference in overall survival between groups with serum HMGB1 levels that were lower and higher than assumed cut-off values was significant.
Our data suggest that serum HMGB1 concentration is a useful prognostic factor for MPM.
In the mammalian endoplasmic reticulum, oxidoreductin-1α (Ero1α) generates protein disulfide bonds and transfers them specifically to canonical protein-disulfide isomerase (PDI) to sustain oxidative ...protein folding. This oxidative process is coupled to the reduction of O2 to H2O2 on the bound flavin adenine dinucleotide cofactor. Because excessive thiol oxidation and H2O2 generation cause cell death, Ero1α activity must be properly regulated. In addition to the four catalytic cysteines (Cys94, Cys99, Cys104, and Cys131) that are located in the flexible active site region, the Cys208–Cys241 pair located at the base of another flexible loop is necessary for Ero1α regulation, although the mechanistic basis is not fully understood. The present study revealed that the Cys208–Cys241 disulfide was reduced by PDI and other PDI family members during PDI oxidation. Differential scanning calorimetry and small angle X-ray scattering showed that mutation of Cys208 and Cys241 did not grossly affect the thermal stability or overall shape of Ero1α, suggesting that redox regulation of this cysteine pair serves a functional role. Moreover, the flexible loop flanked by Cys208 and Cys241 provides a platform for functional interaction with PDI, which in turn enhances the oxidative activity of Ero1α through reduction of the Cys208–Cys241 disulfide. We propose a mechanism of dual Ero1α regulation by dynamic redox interactions between PDI and the two Ero1α flexible loops that harbor the regulatory cysteines.