Biomolecules

16 pages, 3206 KiB

Open AccessArticle

Compound Prunetin Induces Cell Death in Gastric Cancer Cell with Potent Anti-Proliferative Properties: In Vitro Assay, Molecular Docking, Dynamics, and ADMET Studies

by Preethi Vetrivel, Seong Min Kim, Sang Eun Ha, Hun Hwan Kim, Pritam Bhagwan Bhosale, Kalaiselvi Senthil and Gon Sup Kim

Biomolecules 2020, 10(7), 1086; https://doi.org/10.3390/biom10071086 - 21 Jul 2020

Cited by 31 | Viewed by 4224

Abstract

Gastric cancer is the common type of malignancy positioned at second in mortality rate causing burden worldwide with increasing treatment options. Prunetin (PRU) is an O-methylated flavonoid that belongs to the group of isoflavone executing beneficial activities. In the present study, we investigated [...] Read more.

Gastric cancer is the common type of malignancy positioned at second in mortality rate causing burden worldwide with increasing treatment options. Prunetin (PRU) is an O-methylated flavonoid that belongs to the group of isoflavone executing beneficial activities. In the present study, we investigated the anti-proliferative and cell death effect of the compound PRU in AGS gastric cancer cell line. The in vitro cytotoxic potential of PRU was evaluated and significant proliferation was observed. We identified that the mechanism of cell death was due to necroptosis through double staining and was confirmed by co-treatment with inhibitor necrostatin (Nec-1). We further elucidated the mechanism of action of necroptosis via receptor interacting protein kinase 3 (RIPK3) protein expression and it has been attributed by ROS generation through JNK activation. Furthermore, through computational analysis by molecular docking and dynamics simulation, the efficiency of compound prunetin against RIPK3 binding was validated. In addition, we also briefed the pharmacokinetic properties of the compound by in silico ADMET analysis. Full article

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22 pages, 1246 KiB

Open AccessArticle

The GDP-Bound State of Mitochondrial Mfn1 Induces Membrane Adhesion of Apposing Lipid Vesicles through a Cooperative Binding Mechanism

by Andrés Tolosa-Díaz, Víctor G. Almendro-Vedia, Paolo Natale and Iván López-Montero

Biomolecules 2020, 10(7), 1085; https://doi.org/10.3390/biom10071085 - 21 Jul 2020

Cited by 7 | Viewed by 3654

Abstract

Mitochondria are double-membrane organelles that continuously undergo fission and fusion. Outer mitochondrial membrane fusion is mediated by the membrane proteins mitofusin 1 (Mfn1) and mitofusin 2 (Mfn2), carrying a GTP hydrolyzing domain (GTPase) and two coiled-coil repeats. The detailed mechanism on how the [...] Read more.

Mitochondria are double-membrane organelles that continuously undergo fission and fusion. Outer mitochondrial membrane fusion is mediated by the membrane proteins mitofusin 1 (Mfn1) and mitofusin 2 (Mfn2), carrying a GTP hydrolyzing domain (GTPase) and two coiled-coil repeats. The detailed mechanism on how the GTP hydrolysis allows Mfns to approach adjacent membranes into proximity and promote their fusion is currently under debate. Using model membranes built up as giant unilamellar vesicles (GUVs), we show here that Mfn1 promotes membrane adhesion of apposing lipid vesicles. The adhesion forces were sustained by the GDP-bound state of Mfn1 after GTP hydrolysis. In contrast, the incubation with the GDP:AlF

_{4}^{-}

, which mimics the GTP transition state, did not induce membrane adhesion. Due to the flexible nature of lipid membranes, the adhesion strength depended on the surface concentration of Mfn1 through a cooperative binding mechanism. We discuss a possible scenario for the outer mitochondrial membrane fusion based on the modulated action of Mfn1. Full article

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19 pages, 2016 KiB

Open AccessEditor’s ChoiceReview

The Disordered Cellular Multi-Tasker WIP and Its Protein–Protein Interactions: A Structural View

by Chana G. Sokolik, Nasrin Qassem and Jordan H. Chill

Biomolecules 2020, 10(7), 1084; https://doi.org/10.3390/biom10071084 - 21 Jul 2020

Cited by 7 | Viewed by 4079

Abstract

WASp-interacting protein (WIP), a regulator of actin cytoskeleton assembly and remodeling, is a cellular multi-tasker and a key member of a network of protein–protein interactions, with significant impact on health and disease. Here, we attempt to complement the well-established understanding of WIP function [...] Read more.

WASp-interacting protein (WIP), a regulator of actin cytoskeleton assembly and remodeling, is a cellular multi-tasker and a key member of a network of protein–protein interactions, with significant impact on health and disease. Here, we attempt to complement the well-established understanding of WIP function from cell biology studies, summarized in several reviews, with a structural description of WIP interactions, highlighting works that present a molecular view of WIP’s protein–protein interactions. This provides a deeper understanding of the mechanisms by which WIP mediates its biological functions. The fully disordered WIP also serves as an intriguing example of how intrinsically disordered proteins (IDPs) exert their function. WIP consists of consecutive small functional domains and motifs that interact with a host of cellular partners, with a striking preponderance of proline-rich motif capable of interactions with several well-recognized binding partners; indeed, over 30% of the WIP primary structure are proline residues. We focus on the binding motifs and binding interfaces of three important WIP segments, the actin-binding N-terminal domain, the central domain that binds SH3 domains of various interaction partners, and the WASp-binding C-terminal domain. Beyond the obvious importance of a more fundamental understanding of the biology of this central cellular player, this approach carries an immediate and highly beneficial effect on drug-design efforts targeting WIP and its binding partners. These factors make the value of such structural studies, challenging as they are, readily apparent. Full article

(This article belongs to the Special Issue The Amazing World of IDPs in Human Diseases)

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9 pages, 1282 KiB

Open AccessArticle

Sphingolipid Analysis Indicate Lactosylceramide as a Potential Biomarker of Inflammatory Bowel Disease in Children

by Aleksandra Filimoniuk, Agnieszka Blachnio-Zabielska, Monika Imierska, Dariusz Marek Lebensztejn and Urszula Daniluk

Biomolecules 2020, 10(7), 1083; https://doi.org/10.3390/biom10071083 - 21 Jul 2020

Cited by 18 | Viewed by 3635

Abstract

An altered ceramide composition in patients with inflammatory bowel disease (IBD) has been reported recently. The aim of this study was to evaluate the concentrations of sphingolipids in the serum of treatment-naive children with newly diagnosed IBD and to determine the diagnostic value [...] Read more.

An altered ceramide composition in patients with inflammatory bowel disease (IBD) has been reported recently. The aim of this study was to evaluate the concentrations of sphingolipids in the serum of treatment-naive children with newly diagnosed IBD and to determine the diagnostic value of the tested lipids in pediatric IBD. The concentrations of sphingolipids in serum samples were evaluated using a quantitative method, an ultra-high-performance liquid chromatography-tandem mass spectrometry in children with Crohn’s disease (CD) (n=34), ulcerative colitis (UC) (n = 39), and controls (Ctr) (n = 24). Among the study groups, the most significant differences in concentrations were noted for C16:0-LacCer, especially in children with CD compared to Ctr or even to UC. Additionally, the relevant increase in C20:0-Cer and C18:1-Cer concentrations were detected in both IBD groups compared to Ctr. The enhanced C24:0-Cer level was observed only in UC, while C18:0-Cer only in the CD group. The highest area under the curve (AUC), specificity, and sensitivity were determined for C16:0-LacCer in CD diagnosis. Our results suggest that the serum LacC16-Cer may be a potential biomarker that distinguishes children with IBD from healthy controls and differentiates IBD subtypes. In addition, C20:0-Cer and C18:0-Cer levels also seem to be closely connected with IBD. Full article

(This article belongs to the Section Molecular Medicine)

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11 pages, 648 KiB

Open AccessReview

The COP9 Signalosome: A Multi-DUB Complex

by Wolfgang Dubiel, Supattra Chaithongyot, Dawadschargal Dubiel and Michael Naumann

Biomolecules 2020, 10(7), 1082; https://doi.org/10.3390/biom10071082 - 21 Jul 2020

Cited by 53 | Viewed by 7590

Abstract

The COP9 signalosome (CSN) is a signaling platform controlling the cellular ubiquitylation status. It determines the activity and remodeling of ~700 cullin-RING ubiquitin ligases (CRLs), which control more than 20% of all ubiquitylation events in cells and thereby influence virtually any cellular pathway. [...] Read more.

The COP9 signalosome (CSN) is a signaling platform controlling the cellular ubiquitylation status. It determines the activity and remodeling of ~700 cullin-RING ubiquitin ligases (CRLs), which control more than 20% of all ubiquitylation events in cells and thereby influence virtually any cellular pathway. In addition, it is associated with deubiquitylating enzymes (DUBs) protecting CRLs from autoubiquitylation and rescuing ubiquitylated proteins from degradation. The coordination of ubiquitylation and deubiquitylation by the CSN is presumably important for fine-tuning the precise formation of defined ubiquitin chains. Considering its intrinsic DUB activity specific for deneddylation of CRLs and belonging to the JAMM family as well as its associated DUBs, the CSN represents a multi-DUB complex. Two CSN-associated DUBs, the ubiquitin-specific protease 15 (USP15) and USP48 are regulators in the NF-κB signaling pathway. USP15 protects CRL1^β-TrCP responsible for IκBα ubiquitylation, whereas USP48 stabilizes the nuclear pool of the NF-κB transcription factor RelA upon TNF stimulation by counteracting CRL2^SOCS1. Moreover, the CSN controls the neddylation status of cells by its intrinsic DUB activity and by destabilizing the associated deneddylation enzyme 1 (DEN1). Thus, the CSN is a master regulator at the intersection between ubiquitylation and neddylation. Full article

(This article belongs to the Special Issue Ubiquitin-Like Modifiers and Their Diverse Impact on Cell Signaling)

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13 pages, 2714 KiB

Open AccessArticle

DLBCL Cells with Acquired Resistance to Venetoclax Are Not Sensitized to BIRD-2 But Can Be Resensitized to Venetoclax through Bcl-XL Inhibition

by Martijn Kerkhofs, Tamara Vervloessem, Kinga B. Stopa, Victoria M. Smith, Meike Vogler and Geert Bultynck

Biomolecules 2020, 10(7), 1081; https://doi.org/10.3390/biom10071081 - 21 Jul 2020

Cited by 11 | Viewed by 3892

Abstract

Anti-apoptotic Bcl-2-family members are frequently dysregulated in both blood and solid cancers, contributing to their survival despite ongoing oncogenic stress. Yet, such cancer cells often are highly dependent on Bcl-2 for their survival, a feature that is exploited by so-called BH3-mimetic drugs. Venetoclax [...] Read more.

Anti-apoptotic Bcl-2-family members are frequently dysregulated in both blood and solid cancers, contributing to their survival despite ongoing oncogenic stress. Yet, such cancer cells often are highly dependent on Bcl-2 for their survival, a feature that is exploited by so-called BH3-mimetic drugs. Venetoclax (ABT-199) is a selective BH3-mimetic Bcl-2 antagonist that is currently used in the clinic for treatment of chronic lymphocytic leukemia patients. Unfortunately, venetoclax resistance has already emerged in patients, limiting the therapeutic success. Here, we examined strategies to overcome venetoclax resistance. Therefore, we used two diffuse large B-cell lymphoma (DLBCL) cell lines, Riva WT and venetoclax-resistant Riva (VR). The latter was obtained by prolonged culturing in the presence of venetoclax. We report that Riva VR cells did not become more sensitive to BIRD-2, a peptide targeting the Bcl-2 BH4 domain, and established cross-resistance towards BDA-366, a putative BH4-domain antagonist of Bcl-2. However, we found that Bcl-XL, another Bcl-2-family protein, is upregulated in Riva VR, while Mcl-1 expression levels are not different in comparison with Riva WT, hinting towards an increased dependence of Riva VR cells to Bcl-XL. Indeed, Riva VR cells could be resensitized to venetoclax by A-1155463, a selective BH3 mimetic Bcl-XL inhibitor. This is underpinned by siRNA experiments, demonstrating that lowering Bcl-XL-expression levels also augmented the sensitivity of Riva VR cells to venetoclax. Overall, this work demonstrates that Bcl-XL upregulation contributes to acquired resistance of DLBCL cancer cells towards venetoclax and that antagonizing Bcl-XL can resensitize such cells towards venetoclax. Full article

(This article belongs to the Special Issue BCL-2 Family in Health and Diseases)

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14 pages, 2964 KiB

Open AccessCommunication

Analysis of Protein Disorder Predictions in the Light of a Protein Structural Alphabet

by Alexandre G. de Brevern

Biomolecules 2020, 10(7), 1080; https://doi.org/10.3390/biom10071080 - 20 Jul 2020

Cited by 11 | Viewed by 3490

Abstract

Intrinsically-disordered protein (IDP) characterization was an amazing change of paradigm in our classical sequence-structure-function theory. Moreover, IDPs are over-represented in major disease pathways and are now often targeted using small molecules for therapeutic purposes. This has had created a complex continuum from order-that [...] Read more.

Intrinsically-disordered protein (IDP) characterization was an amazing change of paradigm in our classical sequence-structure-function theory. Moreover, IDPs are over-represented in major disease pathways and are now often targeted using small molecules for therapeutic purposes. This has had created a complex continuum from order-that encompasses rigid and flexible regions-to disorder regions; the latter being not accessible through classical crystallographic methodologies. In X-ray structures, the notion of order is dictated by access to resolved atom positions, providing rigidity and flexibility information with low and high experimental B-factors, while disorder is associated with the missing (non-resolved) residues. Nonetheless, some rigid regions can be found in disorder regions. Using ensembles of IDPs, their local conformations were analyzed in the light of a structural alphabet. An entropy index derived from this structural alphabet allowed us to propose a continuum of states from rigidity to flexibility and finally disorder. In this study, the analysis was extended to comparing these results to disorder predictions, underlying a limited correlation, and so opening new ideas to characterize and predict disorder. Full article

(This article belongs to the Special Issue Computational Perspectives on Intrinsic Disorder-Based Functionality)

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24 pages, 1666 KiB

Open AccessReview

POSCAbilities: The Application of the Prion Organotypic Slice Culture Assay to Neurodegenerative Disease Research

by Hailey Pineau and Valerie Sim

Biomolecules 2020, 10(7), 1079; https://doi.org/10.3390/biom10071079 - 20 Jul 2020

Cited by 12 | Viewed by 6182

Abstract

Prion diseases are fatal, transmissible neurodegenerative disorders whose pathogenesis is driven by the misfolding, self-templating and cell-to-cell spread of the prion protein. Other neurodegenerative diseases such as Alzheimer’s disease, Parkinson’s disease, amyotrophic lateral sclerosis and Huntington’s disease, share some of these prion-like features, [...] Read more.

Prion diseases are fatal, transmissible neurodegenerative disorders whose pathogenesis is driven by the misfolding, self-templating and cell-to-cell spread of the prion protein. Other neurodegenerative diseases such as Alzheimer’s disease, Parkinson’s disease, amyotrophic lateral sclerosis and Huntington’s disease, share some of these prion-like features, with different aggregation-prone proteins. Consequently, researchers have begun to apply prion-specific techniques, like the prion organotypic slice culture assay (POSCA), to these disorders. In this review we explore the ways in which the prion phenomenon has been used in organotypic cultures to study neurodegenerative diseases from the perspective of protein aggregation and spreading, strain propagation, the role of glia in pathogenesis, and efficacy of drug treatments. We also present an overview of the advantages and disadvantages of this culture system compared to in vivo and in vitro models and provide suggestions for new directions. Full article

(This article belongs to the Special Issue Prion Diseases: A Model for Neurodegenerative Disorders)

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9 pages, 911 KiB

Open AccessArticle

Coupling Ion Specificity of the Flagellar Stator Proteins MotA1/MotB1 of Paenibacillus sp. TCA20

by Sakura Onoe, Myu Yoshida, Naoya Terahara and Yoshiyuki Sowa

Biomolecules 2020, 10(7), 1078; https://doi.org/10.3390/biom10071078 - 20 Jul 2020

Cited by 4 | Viewed by 2773

Abstract

The bacterial flagellar motor is a reversible rotary molecular nanomachine, which couples ion flux across the cytoplasmic membrane to torque generation. It comprises a rotor and multiple stator complexes, and each stator complex functions as an ion channel and determines the ion specificity [...] Read more.

The bacterial flagellar motor is a reversible rotary molecular nanomachine, which couples ion flux across the cytoplasmic membrane to torque generation. It comprises a rotor and multiple stator complexes, and each stator complex functions as an ion channel and determines the ion specificity of the motor. Although coupling ions for the motor rotation were presumed to be only monovalent cations, such as H⁺ and Na⁺, the stator complex MotA1/MotB1 of Paenibacillus sp. TCA20 (MotA1^TCA/MotB1^TCA) was reported to use divalent cations as coupling ions, such as Ca²⁺ and Mg²⁺. In this study, we initially aimed to measure the motor torque generated by MotA1^TCA/MotB1^TCA under the control of divalent cation motive force; however, we identified that the coupling ion of MotA1^TCAMotB1^TCA is very likely to be a monovalent ion. We engineered a series of functional chimeric stator proteins between MotB1^TCA and Escherichia coli MotB. E. coli ΔmotAB cells expressing MotA1^TCA and the chimeric MotB presented significant motility in the absence of divalent cations. Moreover, we confirmed that MotA1^TCA/MotB1^TCA in Bacillus subtilis ΔmotABΔmotPS cells generates torque without divalent cations. Based on two independent experimental results, we conclude that the MotA1^TCA/MotB1^TCA complex directly converts the energy released from monovalent cation flux to motor rotation. Full article

(This article belongs to the Special Issue Perspectives on Bacterial Flagellar Motor)

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16 pages, 2910 KiB

Open AccessFeature PaperArticle

Dual-Functioning Antibacterial Eugenol-Derived Plasticizers for Polylactide

by Wenxiang Xuan, Karin Odelius and Minna Hakkarainen

Biomolecules 2020, 10(7), 1077; https://doi.org/10.3390/biom10071077 - 20 Jul 2020

Cited by 15 | Viewed by 4006

Abstract

Dual-functioning additives with plasticizing and antibacterial functions were designed by exploiting the natural aromatic compound eugenol and green platform chemical levulinic acid or valeric acid that can be produced from biobased resources. One-pot synthesis methodology was utilized to create three ester-rich plasticizers. The [...] Read more.

Dual-functioning additives with plasticizing and antibacterial functions were designed by exploiting the natural aromatic compound eugenol and green platform chemical levulinic acid or valeric acid that can be produced from biobased resources. One-pot synthesis methodology was utilized to create three ester-rich plasticizers. The plasticizers were thoroughly characterized by several nuclear magnetic resonance techniques (¹H NMR, ¹³C NMR, ³¹P NMR, HSQC, COSY, HMBC) and by electrospray ionization-mass spectrometry (ESI-MS) and their performances, as plasticizers for polylactide (PLA), were evaluated. The eugenyl valerate was equipped with a strong capability to depress the glass transition temperature (T_g) of PLA. Incorporating 30 wt% plasticizer led to a reduction of the T_g by 43 °C. This was also reflected by a remarkable change in mechanical properties, illustrated by a strain at break of 560%, almost 110 times the strain for the breaking of neat PLA. The two eugenyl levulinates also led to PLA with significantly increased strain at breaking. The eugenyl levulinates portrayed higher thermal stabilities than eugenyl valerate, both neat and in PLA blends. The different concentrations of phenol, carboxyl and alcohol functional groups in the three plasticizers caused different bactericidal activities. The eugenyl levulinate with the highest phenol-, carboxyl- and alcohol group content significantly inhibited the growth of Staphylococcus aureus and Escherichia coli, while the other two plasticizers could only inhibit the growth of Staphylococcus aureus. Thus, the utilization of eugenol as a building block in plasticizer design for PLA illustrated an interesting potential for production of additives with dual functions, being both plasticizers and antibacterial agents. Full article

(This article belongs to the Special Issue Biodegradable Polyesters: From Synthesis to Application)

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16 pages, 1777 KiB

Open AccessArticle

The Long-Acting D3 Partial Agonist MC-25-41 Attenuates Motivation for Cocaine in Sprague-Dawley Rats

by Gregory L. Powell, Mark D. Namba, Annika Vannan, John Paul Bonadonna, Andrew Carlson, Rachel Mendoza, Peng-Jen Chen, Robert R. Luetdke, Benjamin E. Blass and Janet L. Neisewander

Biomolecules 2020, 10(7), 1076; https://doi.org/10.3390/biom10071076 - 18 Jul 2020

Cited by 9 | Viewed by 3066

Abstract

The dopamine D3 receptor is a prime target for developing treatments for cocaine use disorders (CUDs). In this study, we conducted a pre-clinical investigation of the therapeutic potential of a long-acting, D3 receptor partial agonist, MC-25-41. Male rats were pre-treated with MC-25-41 (vehicle, [...] Read more.

The dopamine D3 receptor is a prime target for developing treatments for cocaine use disorders (CUDs). In this study, we conducted a pre-clinical investigation of the therapeutic potential of a long-acting, D3 receptor partial agonist, MC-25-41. Male rats were pre-treated with MC-25-41 (vehicle, 1.0, 3.0, 5.6, or 10 mg/kg, intraperitoneal (IP)) five minutes prior to tests of cocaine or sucrose intake on either a progressive ratio schedule of reinforcement or a variable interval 60 s multiple schedule consisting of 4, 15-min components with sucrose or cocaine available in alternating components. A separate cohort of rats was tested on a within-session, dose-reduction procedure to determine the effects of MC-25-41 on demand for cocaine using a behavioral economics analysis. Finally, rats were tested for effects of MC-25-41 on spontaneous and cocaine-induced locomotion. MC-25-41 failed to alter locomotion, but reduced reinforcement rates for both cocaine and sucrose on the low-effort, multiple schedule. However, on the higher-effort, progressive ratio schedule of cocaine reinforcement, MC-25-41 reduced infusions, and active lever presses at doses that did not alter sucrose intake. The behavioral economics analysis showed that MC-25-41 also increased cocaine demand elasticity compared to vehicle, indicating a reduction in consumption as price increases. Together, these results suggest that similar to other D3-selective antagonists and partial agonists, MC-25-41 reduces motivation for cocaine under conditions of high cost but has the added advantage of a long half-life (>10 h). These findings suggest that MC-25-41 may be a suitable pre-clinical lead compound for development of medications to treat CUDs. Full article

(This article belongs to the Special Issue Dopamine D3 Receptor: Contemporary Views of Its Function and Pharmacology for Neuropsychiatric Diseases)

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17 pages, 1774 KiB

Open AccessArticle

Structural Elucidation of Irish Ale Bioactive Polar Lipids with Antithrombotic Properties

by Alexandros Tsoupras, Ronan Lordan, Eoin O'Keefe, Katie Shiels, Sushanta Kumar Saha and Ioannis Zabetakis

Biomolecules 2020, 10(7), 1075; https://doi.org/10.3390/biom10071075 - 18 Jul 2020

Cited by 19 | Viewed by 4291

Abstract

The structures of bioactive polar lipids (PLs) of Irish ale with potent antithrombotic and cardioprotective properties were elucidated. Ale PL was fractionated by preparative thin layer chromatography (TLC) into subclasses, and their antithrombotic effect was assessed against human platelet aggregation induced by the [...] Read more.

The structures of bioactive polar lipids (PLs) of Irish ale with potent antithrombotic and cardioprotective properties were elucidated. Ale PL was fractionated by preparative thin layer chromatography (TLC) into subclasses, and their antithrombotic effect was assessed against human platelet aggregation induced by the pro-inflammatory mediator, platelet-activating factor (PAF). The fatty acid content and the overall structures of ale PL were elucidated by liquid chromatography mass spectrometry (LC-MS). Phosphatidylcholines (PC) and molecules of the sphingomyelin (SM) family exhibited the strongest anti-PAF effects, followed by phosphatidylethanolamines (PE). PC contained higher amounts of omega-3 polyunsaturated fatty acids (n-3 PUFA) and thus the lowest n-6/n-3 ratio. Bioactive diacyl and alkyl-acyl PC and PE molecules bearing n-3 PUFA at their sn-2 position, especially docosahexaenoic acid (DHA) and α-linolenic acid (ALA) but mostly oleic acid (OA), were identified in both PC and PE subclasses. Eicosapentaenoic acid (EPA) was present only in bioactive PC molecules and not in PE, explaining the lower anti-PAF effects of PE. Bioactive sphingolipid and glycolipid molecules with reported anti-inflammatory and anti-tumour properties, such as specific ceramides and glucosylcerebrosides with sphingosine, phytosphingosine and dihydrosphingosine bases but also specific monogalactodiglycerides and SM species bearing ALA at their sn-2 position, were identified in the SM subclass, providing a rational for its strong bioactivities against the PAF pathway. Further studies are required on the health benefits of bioactive PL from beer and brewery by-products. Full article

(This article belongs to the Section Natural and Bio-derived Molecules)

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10 pages, 1061 KiB

Open AccessArticle

Identification of a Biosynthetic Gene Cluster Responsible for the Production of a New Pyrrolopyrimidine Natural Product—Huimycin

by Hui Shuai, Maksym Myronovskyi, Suvd Nadmid and Andriy Luzhetskyy

Biomolecules 2020, 10(7), 1074; https://doi.org/10.3390/biom10071074 - 18 Jul 2020

Cited by 13 | Viewed by 3501

Abstract

Pyrrolopyrimidines are an important class of natural products with a broad spectrum of biological activities, including antibacterial, antifungal, antiviral, anticancer or anti-inflammatory. Here, we present the identification of a biosynthetic gene cluster from the rare actinomycete strain Kutzneria albida DSM 43870, which leads [...] Read more.

Pyrrolopyrimidines are an important class of natural products with a broad spectrum of biological activities, including antibacterial, antifungal, antiviral, anticancer or anti-inflammatory. Here, we present the identification of a biosynthetic gene cluster from the rare actinomycete strain Kutzneria albida DSM 43870, which leads to the production of huimycin, a new member of the pyrrolopyrimidine family of compounds. The huimycin gene cluster was successfully expressed in the heterologous host strain Streptomyces albus Del14. The compound was purified, and its structure was elucidated by means of nuclear magnetic resonance spectroscopy. The minimal huimycin gene cluster was identified through sequence analysis and a series of gene deletion experiments. A model for huimycin biosynthesis is also proposed in this paper. Full article

(This article belongs to the Section Natural and Bio-derived Molecules)

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20 pages, 1916 KiB

Open AccessArticle

Isoprostanoid Profiling of Marine Microalgae

by Claire Vigor, Camille Oger, Guillaume Reversat, Amandine Rocher, Bingqing Zhou, Amandyne Linares-Maurizi, Alexandre Guy, Valérie Bultel-Poncé, Jean-Marie Galano, Joseph Vercauteren, Thierry Durand, Philippe Potin, Thierry Tonon and Catherine Leblanc

Biomolecules 2020, 10(7), 1073; https://doi.org/10.3390/biom10071073 - 18 Jul 2020

Cited by 24 | Viewed by 3510

Abstract

Algae result from a complex evolutionary history that shapes their metabolic network. For example, these organisms can synthesize different polyunsaturated fatty acids, such as those found in land plants and oily fish. Due to the presence of numerous double-bonds, such molecules can be [...] Read more.

Algae result from a complex evolutionary history that shapes their metabolic network. For example, these organisms can synthesize different polyunsaturated fatty acids, such as those found in land plants and oily fish. Due to the presence of numerous double-bonds, such molecules can be oxidized nonenzymatically, and this results in the biosynthesis of high-value bioactive metabolites named isoprostanoids. So far, there have been only a few studies reporting isoprostanoid productions in algae. To fill this gap, the current investigation aimed at profiling isoprostanoids by liquid chromatography -mass spectrometry/mass spectrometry (LC-MS/MS) in four marine microalgae. A good correlation was observed between the most abundant polyunsaturated fatty acids (PUFAs) produced by the investigated microalgal species and their isoprostanoid profiles. No significant variations in the content of oxidized derivatives were observed for Rhodomonas salina and Chaetoceros gracilis under copper stress, whereas increases in the production of C18-, C20- and C22-derived isoprostanoids were monitored in Tisochrysis lutea and Phaeodactylum tricornutum. In the presence of hydrogen peroxide, no significant changes were observed for C. gracilis and for T. lutea, while variations were monitored for the other two algae. This study paves the way to further studying the physiological roles of isoprostanoids in marine microalgae and exploring these organisms as bioresources for isoprostanoid production. Full article

(This article belongs to the Special Issue Lipids of Marine Algae)

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17 pages, 1668 KiB

Open AccessArticle

Regulated Iron Siderophore Production of the Halophilic Archaeon Haloferax volcanii

by Natalie Niessen and Jörg Soppa

Biomolecules 2020, 10(7), 1072; https://doi.org/10.3390/biom10071072 - 17 Jul 2020

Cited by 13 | Viewed by 3943

Abstract

Iron is part of many redox and other enzymes and, thus, it is essential for all living beings. Many oxic environments have extremely low concentrations of free iron. Therefore, many prokaryotic species evolved siderophores, i.e., small organic molecules that complex Fe³⁺ with [...] Read more.

Iron is part of many redox and other enzymes and, thus, it is essential for all living beings. Many oxic environments have extremely low concentrations of free iron. Therefore, many prokaryotic species evolved siderophores, i.e., small organic molecules that complex Fe³⁺ with very high affinity. Siderophores of bacteria are intensely studied, in contrast to those of archaea. The haloarchaeon Haloferax volcanii contains a gene cluster that putatively encodes siderophore biosynthesis genes, including four iron uptake chelate (iuc) genes. Underscoring this hypothesis, Northern blot analyses revealed that a hexacistronic transcript is generated that is highly induced under iron starvation. A quadruple iuc deletion mutant was generated, which had a growth defect solely at very low concentrations of Fe³⁺, not Fe²⁺. Two experimental approaches showed that the wild type produced and exported an Fe³⁺-specific siderophore under low iron concentrations, in contrast to the iuc deletion mutant. Bioinformatic analyses revealed that haloarchaea obtained the gene cluster by lateral transfer from bacteria and enabled the prediction of enzymatic functions of all six gene products. Notably, a biosynthetic pathway is proposed that starts with aspartic acid, uses several group donors and citrate, and leads to the hydroxamate siderophore Schizokinen. Full article

(This article belongs to the Collection Archaea: Diversity, Metabolism and Molecular Biology)

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22 pages, 1526 KiB

Open AccessReview

Critical Roles of N⁶-Methyladenosine (m⁶A) in Cancer and Virus Infection

by Ken Asada, Amina Bolatkan, Ken Takasawa, Masaaki Komatsu, Syuzo Kaneko and Ryuji Hamamoto

Biomolecules 2020, 10(7), 1071; https://doi.org/10.3390/biom10071071 - 17 Jul 2020

Cited by 16 | Viewed by 4935

Abstract

Studies have shown that epigenetic abnormalities are involved in various diseases, including cancer. In particular, in order to realize precision medicine, the integrated analysis of genetics and epigenetics is considered to be important; detailed epigenetic analysis in the medical field has been becoming [...] Read more.

Studies have shown that epigenetic abnormalities are involved in various diseases, including cancer. In particular, in order to realize precision medicine, the integrated analysis of genetics and epigenetics is considered to be important; detailed epigenetic analysis in the medical field has been becoming increasingly important. In the epigenetics analysis, DNA methylation and histone modification analyses have been actively studied for a long time, and many important findings were accumulated. On the other hand, recently, attention has also been focused on RNA modification in the field of epigenetics; now it is known that RNA modification is associated with various biological functions, such as regulation of gene expression. Among RNA modifications, functional analysis of N⁶-methyladenosine (m⁶A), the most abundant RNA modification found from humans to plants is actively progressing, and it has also been known that m⁶A abnormality is involved in cancer and other diseases. Importantly, recent studies have shown that m⁶A is related to viral infections. Considering the current world situation under threat of viral infections, it is important to deepen knowledge of RNA modification from the viewpoint of viral diseases. Hence, in this review, we have summarized the recent findings regarding the roles of RNA modifications in biological functions, cancer biology, and virus infection, particularly focusing on m⁶A in mRNA. Full article

(This article belongs to the Special Issue Epigenetics in Cancer)

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16 pages, 2192 KiB

Open AccessFeature PaperArticle

Adipose Tissue-Derived Stem Cells Retain Their Adipocyte Differentiation Potential in Three-Dimensional Hydrogels and Bioreactors

by Benjamen T. O'Donnell, Sara Al-Ghadban, Clara J. Ives, Michael P. L'Ecuyer, Tia A. Monjure, Monica Romero-Lopez, Zhong Li, Stuart B. Goodman, Hang Lin, Rocky S. Tuan and Bruce A. Bunnell

Biomolecules 2020, 10(7), 1070; https://doi.org/10.3390/biom10071070 - 17 Jul 2020

Cited by 30 | Viewed by 4998

Abstract

Osteoarthritis (OA) is a common joint disorder with a significant economic and healthcare impact. The knee joint is composed of cartilage and the adjoining bone, a synovial capsule, the infrapatellar fat pad (IPFP), and other connective tissues such as tendons and ligaments. Adipose [...] Read more.

Osteoarthritis (OA) is a common joint disorder with a significant economic and healthcare impact. The knee joint is composed of cartilage and the adjoining bone, a synovial capsule, the infrapatellar fat pad (IPFP), and other connective tissues such as tendons and ligaments. Adipose tissue has recently been highlighted as a major contributor to OA through strong inflammation mediating effects. In this study, methacrylated gelatin (GelMA) constructs seeded with adipose tissue-derived mesenchymal stem cells (ASCs) and cultured in a 3D printed bioreactor were investigated for use in microphysiological systems to model adipose tissue in the knee joint. Four patient-derived ASC populations were seeded at a density of 20 million cells/mL in GelMA. Live/Dead and boron-dipyrromethene/4′,6-diamidino-2-phenylindole (BODIPY/DAPI) staining of cells within the constructs demonstrated robust cell viability after 28 days in a growth (control) medium, and robust cell viability and lipid accumulation in adipogenic differentiation medium. qPCR gene expression analysis and protein analysis demonstrated an upregulated expression of key adipogenesis-associated genes. Overall, these data indicate that ASCs retain their adipogenic potential when seeded within GelMA hydrogels and cultured within perfusion bioreactors, and thus can be used in a 3D organ-on-a-chip system to study the role of the IPFP in the pathobiology of the knee OA. Full article

(This article belongs to the Special Issue Recent Advances in Translational Adipose-Derived Stem Cell Biology)

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20 pages, 15251 KiB

Open AccessArticle

The Droserasin 1 PSI: A Membrane-Interacting Antimicrobial Peptide from the Carnivorous Plant Drosera capensis

by Marc A. Sprague-Piercy, Jan C. Bierma, Marquise G. Crosby, Brooke P. Carpenter, Gemma R. Takahashi, Joana Paulino, Ivan Hung, Rongfu Zhang, John E. Kelly, Natalia Kozlyuk, Xi Chen, Carter T. Butts and Rachel W. Martin

Biomolecules 2020, 10(7), 1069; https://doi.org/10.3390/biom10071069 - 17 Jul 2020

Cited by 8 | Viewed by 5513

Abstract

The Droserasins, aspartic proteases from the carnivorous plant Drosera capensis, contain a 100-residue plant-specific insert (PSI) that is post-translationally cleaved and independently acts as an antimicrobial peptide. PSIs are of interest not only for their inhibition of microbial growth, but also because [...] Read more.

The Droserasins, aspartic proteases from the carnivorous plant Drosera capensis, contain a 100-residue plant-specific insert (PSI) that is post-translationally cleaved and independently acts as an antimicrobial peptide. PSIs are of interest not only for their inhibition of microbial growth, but also because they modify the size of lipid vesicles and strongly interact with biological membranes. PSIs may therefore be useful for modulating lipid systems in NMR studies of membrane proteins. Here we present the expression and biophysical characterization of the Droserasin 1 PSI (D1 PSI.) This peptide is monomeric in solution and maintains its primarily

α

-helical secondary structure over a wide range of temperatures and pH values, even under conditions where its three disulfide bonds are reduced. Vesicle fusion assays indicate that the D1 PSI strongly interacts with bacterial and fungal lipids at pH 5 and lower, consistent with the physiological pH of D. capensis mucilage. It binds lipids with a variety of head groups, highlighting its versatility as a potential stabilizer for lipid nanodiscs. Solid-state NMR spectra collected at a field strength of 36 T, using a unique series-connected hybrid magnet, indicate that the peptide is folded and strongly bound to the membrane. Molecular dynamics simulations indicate that the peptide is stable as either a monomer or a dimer in a lipid bilayer. Both the monomer and the dimer allow the passage of water through the membrane, albeit at different rates. Full article

(This article belongs to the Special Issue Advances in Membrane Proteins)

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18 pages, 991 KiB

Open AccessEditor’s ChoiceReview

The Multifaceted Pyruvate Metabolism: Role of the Mitochondrial Pyruvate Carrier

by Joséphine Zangari, Francesco Petrelli, Benoît Maillot and Jean-Claude Martinou

Biomolecules 2020, 10(7), 1068; https://doi.org/10.3390/biom10071068 - 17 Jul 2020

Cited by 73 | Viewed by 22590

Abstract

Pyruvate, the end product of glycolysis, plays a major role in cell metabolism. Produced in the cytosol, it is oxidized in the mitochondria where it fuels the citric acid cycle and boosts oxidative phosphorylation. Its sole entry point into mitochondria is through the [...] Read more.

Pyruvate, the end product of glycolysis, plays a major role in cell metabolism. Produced in the cytosol, it is oxidized in the mitochondria where it fuels the citric acid cycle and boosts oxidative phosphorylation. Its sole entry point into mitochondria is through the recently identified mitochondrial pyruvate carrier (MPC). In this review, we report the latest findings on the physiology of the MPC and we discuss how a dysfunctional MPC can lead to diverse pathologies, including neurodegenerative diseases, metabolic disorders, and cancer. Full article

(This article belongs to the Special Issue Mitochondrial Transport Proteins)

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9 pages, 2163 KiB

Open AccessArticle

Residue and Dissipation Kinetics of Metsulfuron-Methyl Herbicide in Soil: A Field Assessment at an Oil Palm Plantation

by Zainol Maznah, B. Sahid Ismail and Oii Kok Eng

Biomolecules 2020, 10(7), 1067; https://doi.org/10.3390/biom10071067 - 16 Jul 2020

Cited by 6 | Viewed by 3383

Abstract

A field trial experiment was conducted to investigate the degradation of metsulfuron-methyl at two application dosages, 15 g a.i/ha and 30 g a.i/ha, at an oil palm plantation. Soil samples were collected at ‒1, 0, 1, 3, 7, 14, and 21 days after [...] Read more.

A field trial experiment was conducted to investigate the degradation of metsulfuron-methyl at two application dosages, 15 g a.i/ha and 30 g a.i/ha, at an oil palm plantation. Soil samples were collected at ‒1, 0, 1, 3, 7, 14, and 21 days after treatment (DAT) at the following depths: 0–10, 10–20, 20–30, 30–40, and 40–50 cm. The results showed rapid degradation of metsulfuron-methyl in the soil, with calculated half-life (t_½) values ranging from 6.3 and 7.9 days. The rates of degradation of metsulfuron-methyl followed first-order reaction kinetics (R² = 0.91–0.92). At the spray dosage of 15 g a.i/ha, metsulfuron-methyl residue was detected at up to 20–30 cm soil depth, at 3.56% to 1.78% at 3 and 7 DAT, respectively. Doubling the dosage to 30 g a.i/ha increased the metsulfuron-methyl residue in up to 30–40 cm soil depth at 3, 7, and 14 DAT, with concentrations ranging from 1.90% to 1.74%. These findings suggest that metsulfuron-methyl has a low impact on the accumulation of the residues in the soil at application dosages of 15 g a.i/ha and 30 g a.i/ha, due to rapid degradation, and the half-life was found to be 6.3 to 7.9 days. Full article

(This article belongs to the Special Issue Biomolecules from Plant Residues)

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36 pages, 3692 KiB

Open AccessArticle

Proteomic and Bioinformatic Investigations of Heat-Treated Anisakis simplex Third-Stage Larvae

by Maciej Kochanowski, Mirosław Różycki, Joanna Dąbrowska, Aneta Bełcik, Jacek Karamon, Jacek Sroka and Tomasz Cencek

Biomolecules 2020, 10(7), 1066; https://doi.org/10.3390/biom10071066 - 16 Jul 2020

Cited by 11 | Viewed by 4339

Abstract

Anisakis simplex third-stage larvae are the main source of hidden allergens in marine fish products. Some Anisakis allergens are thermostable and, even highly processed, could cause hypersensitivity reactions. However, Anisakis proteome has not been studied under autoclaving conditions of 121 °C for 60 [...] Read more.

Anisakis simplex third-stage larvae are the main source of hidden allergens in marine fish products. Some Anisakis allergens are thermostable and, even highly processed, could cause hypersensitivity reactions. However, Anisakis proteome has not been studied under autoclaving conditions of 121 °C for 60 min, which is an important process in the food industry. The aim of the study was the identification and characterization of allergens, potential allergens, and other proteins of heat-treated A. simplex larvae. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to identify 470 proteins, including allergens—Ani s 1, Ani s 2, Ani s 3, Ani s 4, Ani s 5—and 13 potential allergens that were mainly homologs of Anisakis spp., Ascaris spp., and Acari allergens. Ani s 2, Ani s 3, Ani s 5, and three possible allergens were found among the top 25 most abundant proteins. The computational analysis allowed us to detect allergen epitopes, assign protein families, and domains as well as to annotate the localization of proteins. The predicted 3D models of proteins revealed similarities between potential allergens and homologous allergens. Despite the partial degradation of heated A. simplex antigens, their immunoreactivity with anti-A. simplex IgG antibodies was confirmed using a Western blot. In conclusion, identified epitopes of allergenic peptides highlighted that the occurrence of Anisakis proteins in thermally processed fish products could be a potential allergic hazard. Further studies are necessary to confirm the IgE immunoreactivity and thermostability of identified proteins. Full article

(This article belongs to the Special Issue Advances on Allergens Identification and Characterization)

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19 pages, 3049 KiB

Open AccessArticle

The Role of Cardiac N-Methyl-D-Aspartate Receptors in Heart Conditioning—Effects on Heart Function and Oxidative Stress

by Natalia Govoruskina, Vladimir Jakovljevic, Vladimir Zivkovic, Isidora Milosavljevic, Jovana Jeremic, Jovana Bradic, Sergey Bolevich, Israpil Alisultanovich Omarov, Dragan Djuric, Katarina Radonjic, Marijana Andjic, Nevena Draginic, Aleksandra Stojanovic and Ivan Srejovic

Biomolecules 2020, 10(7), 1065; https://doi.org/10.3390/biom10071065 - 16 Jul 2020

Cited by 9 | Viewed by 2923

Abstract

As well as the most known role of N-methyl-D-aspartate receptors (NMDARs) in the nervous system, there is a plethora of evidence that NMDARs are also present in the cardiovascular system where they participate in various physiological processes, as well as pathological conditions. The [...] Read more.

As well as the most known role of N-methyl-D-aspartate receptors (NMDARs) in the nervous system, there is a plethora of evidence that NMDARs are also present in the cardiovascular system where they participate in various physiological processes, as well as pathological conditions. The aim of this study was to assess the effects of preconditioning and postconditioning of isolated rat heart with NMDAR agonists and antagonists on heart function and release of oxidative stress biomarkers. The hearts of male Wistar albino rats were subjected to global ischemia for 20 min, followed by 30 min of reperfusion, using the Langendorff technique, and cardiodynamic parameters were determined during the subsequent preconditioning with the NMDAR agonists glutamate (100 µmol/L) and (RS)-(Tetrazol-5-yl)glycine (5 μmol/L) and the NMDAR antagonists memantine (100 μmol/L) and MK-801 (30 μmol/L). In the postconditioning group, the hearts were perfused with the same dose of drugs during the first 3 min of reperfusion. The oxidative stress biomarkers were determined spectrophotometrically in samples of coronary venous effluent. The NMDAR antagonists, especially MK-801, applied in postconditioning had a marked antioxidative effect with a most pronounced protective effect. The results from this study suggest that NMDARs could be a potential therapeutic target in the prevention and treatment of ischemic and reperfusion injury of the heart. Full article

(This article belongs to the Special Issue NMDA Receptor in Health and Diseases)

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10 pages, 1201 KiB

Open AccessBrief Report

Over-Expression of Allograft Inflammatory Factor-1 (AIF-1) in Patients with Rheumatoid Arthritis

by Katarzyna Piotrowska, Sylwia Słuczanowska-Głabowska, Mateusz Kurzawski, Violetta Dziedziejko, Patrycja Kopytko, Edyta Paczkowska, Dorota Rogińska, Krzysztof Safranow, Bogusław Machaliński and Andrzej Pawlik

Biomolecules 2020, 10(7), 1064; https://doi.org/10.3390/biom10071064 - 16 Jul 2020

Cited by 18 | Viewed by 2793

Abstract

Allograft inflammatory factor-1 (AIF-1) is a cytoplasmic protein that is encoded by the AIF1 gene. The main functions of AIF-1 are the activation of macrophages and enhancing the production of pro-inflammatory cytokines. To date, three different AIF-1 isoforms have been identified. In this [...] Read more.

Allograft inflammatory factor-1 (AIF-1) is a cytoplasmic protein that is encoded by the AIF1 gene. The main functions of AIF-1 are the activation of macrophages and enhancing the production of pro-inflammatory cytokines. To date, three different AIF-1 isoforms have been identified. In this study, we examined the expression of AIF-1 isoforms on the level of mRNA, and we compared the percentage of AIF-1-positive white blood cells (WBCs) in blood and AIF-1/CD68 cells in the synovial membranes in patients with rheumatoid arthritis (RA) and osteoarthritis (OA). We examined 15 patients with RA and 15 patients with OA who had previously undergone knee arthroplasty. Peripheral blood and synovial membranes (SMs) were collected from these patients during knee arthroplasty. We identified three AIF-1 mRNA expression variants in peripheral mononuclear cells (PBMCs) and SMs from patients in both groups. Spearman’s rank correlation coefficient tests showed strong, positive, and significant correlations between the three AIF-1 mRNA expression variants in PBMCs and/or SMs in patients with RA and OA. There were no statistically significant correlations for any of the AIF-1 mRNA expression variants between PBMCs and SMs in patients with RA and OA. We observed a statistically significant increased percentage of AIF-1-positive cells in patients with RA in comparison to patients with OA. The percentage of AIF-1-positive cells in the blood of patients with RA and OA was 1.35 ± 0.81% and 0.71 ± 0.25% (p < 0.01), respectively, whereas the percentage of AIF-1/CD68-positive WBC cells in the SMs was 24.05 ± 7.17% and 4.78 ± 1.52% (p < 0.001), respectively. In conclusion, three AIF-1 mRNA expression variants occurred in PBMCs and SM cells in patients with RA and OA. The AIF-1 mRNA expression levels of the variants correlated with each other in PBMCs and SM cells, but there were no statistically significant correlations for AIF-1 mRNA expression variants between PBMCs and SM cells in patients with RA and OA. Both in the blood and SMs, we observed an increased percentage of AIF-1-positive cells in patients with RA in comparison to patients with OA. The above results suggested that AIF-1 was the cytokine involved in the pathogenesis of RA. The precise knowledge of the role of AIF-1 in RA pathogenesis and the development of inflammatory response requires further investigations. Full article

(This article belongs to the Special Issue Pathogenesis of Arthritis)

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17 pages, 5496 KiB

Open AccessArticle

Understanding the Biosynthetic Changes that Give Origin to the Distinctive Flavor of Sotol: Microbial Identification and Analysis of the Volatile Metabolites Profiles During Sotol (Dasylirion sp.) Must Fermentation

by Francisco Javier Zavala-Díaz de la Serna, Ricardo Contreras-López, L. Paola Lerma-Torres, Francisco Ruiz-Terán, Beatriz A. Rocha-Gutiérrez, Samuel B. Pérez-Vega, Leslie R. Elías-Ogaz and Ivan Salmerón

Biomolecules 2020, 10(7), 1063; https://doi.org/10.3390/biom10071063 - 16 Jul 2020

Cited by 3 | Viewed by 3360

Abstract

In northern Mexico, the distilled spirit sotol with a denomination of origin is made from species of Dasylirion. The configuration of the volatile metabolites produced during the spontaneous fermentation of Dasylirion sp. must is insufficiently understood. In this study, the aim was [...] Read more.

In northern Mexico, the distilled spirit sotol with a denomination of origin is made from species of Dasylirion. The configuration of the volatile metabolites produced during the spontaneous fermentation of Dasylirion sp. must is insufficiently understood. In this study, the aim was to investigate the composition of the microbial consortia, describe the variation of volatile metabolites, and relate such profiles with their particular flavor attributes during the fermentation of sotol (Dasylirion sp.) must. Ascomycota was the phylum of most strains identified with 75% of total abundance. The genus of fermenting yeasts constituted of 101 Pichia strains and 13 Saccharomyces strains. A total of 57 volatile metabolites were identified and grouped into ten classes. The first stage of fermentation was composed of diesel, green, fruity, and cheesy attributes due to butyl 2-methylpropanoate, octan-1-ol, ethyl octanoate, and butanal, respectively, followed by a variation to pungent and sweet descriptors due to 3-methylbutan-1-ol and butyl 2-methylpropanoate. The final stage was described by floral, ethereal-winey, and vinegar attributes related to ethyl ethanimidate, 2-methylpropan-1-ol, and 2-hydroxyacetic acid. Our results improve the knowledge of the variations of volatile metabolites during the fermentation of sotol must and their contribution to its distinctive flavor. Full article

(This article belongs to the Special Issue Function of Microorganism in Food Production)

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17 pages, 2824 KiB

Open AccessArticle

Nickel and GTP Modulate Helicobacter pylori UreG Structural Flexibility

by Annalisa Pierro, Emilien Etienne, Guillaume Gerbaud, Bruno Guigliarelli, Stefano Ciurli, Valérie Belle, Barbara Zambelli and Elisabetta Mileo

Biomolecules 2020, 10(7), 1062; https://doi.org/10.3390/biom10071062 - 16 Jul 2020

Cited by 8 | Viewed by 3480

Abstract

UreG is a P-loop GTP hydrolase involved in the maturation of nickel-containing urease, an essential enzyme found in plants, fungi, bacteria, and archaea. This protein couples the hydrolysis of GTP to the delivery of Ni(II) into the active site of apo-urease, interacting with [...] Read more.

UreG is a P-loop GTP hydrolase involved in the maturation of nickel-containing urease, an essential enzyme found in plants, fungi, bacteria, and archaea. This protein couples the hydrolysis of GTP to the delivery of Ni(II) into the active site of apo-urease, interacting with other urease chaperones in a multi-protein complex necessary for enzyme activation. Whereas the conformation of Helicobacter pylori (Hp) UreG was solved by crystallography when it is in complex with two other chaperones, in solution the protein was found in a disordered and flexible form, defining it as an intrinsically disordered enzyme and indicating that the well-folded structure found in the crystal state does not fully reflect the behavior of the protein in solution. Here, isothermal titration calorimetry and site-directed spin labeling coupled to electron paramagnetic spectroscopy were successfully combined to investigate HpUreG structural dynamics in solution and the effect of Ni(II) and GTP on protein mobility. The results demonstrate that, although the protein maintains a flexible behavior in the metal and nucleotide bound forms, concomitant addition of Ni(II) and GTP exerts a structural change through the crosstalk of different protein regions. Full article

(This article belongs to the Special Issue The Amazing World of IDPs in Human Diseases)

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30 pages, 1678 KiB

Open AccessReview

Epigenetic Mechanisms of Resistance to Immune Checkpoint Inhibitors

by Alexandre Perrier, Audrey Didelot, Pierre Laurent-Puig, Hélène Blons and Simon Garinet

Biomolecules 2020, 10(7), 1061; https://doi.org/10.3390/biom10071061 - 16 Jul 2020

Cited by 64 | Viewed by 6706

Abstract

Immune checkpoint inhibitors (ICIs) have demonstrated to be highly efficient in treating solid tumors; however, many patients have limited benefits in terms of response and survival. This rapidly led to the investigation of combination therapies to enhance response rates. Moreover, predictive biomarkers were [...] Read more.

Immune checkpoint inhibitors (ICIs) have demonstrated to be highly efficient in treating solid tumors; however, many patients have limited benefits in terms of response and survival. This rapidly led to the investigation of combination therapies to enhance response rates. Moreover, predictive biomarkers were assessed to better select patients. Although PD-L1 expression remains the only validated marker in clinics, molecular profiling has brought valuable information, showing that the tumor mutation load and microsatellite instability (MSI) status were associated to higher response rates in nearly all cancer types. Moreover, in lung cancer, EGFR and MET mutations, oncogene fusions or STK11 inactivating mutations were associated with low response rates. Cancer progression towards invasive phenotypes that impede immune surveillance relies on complex regulatory networks and cell interactions within the tumor microenvironment. Epigenetic modifications, such as the alteration of histone patterns, chromatin structure, DNA methylation status at specific promoters and changes in microRNA levels, may alter the cell phenotype and reshape the tumor microenvironment, allowing cells to grow and escape from immune surveillance. The objective of this review is to make an update on the identified epigenetic changes that target immune surveillance and, ultimately, ICI responses, such as histone marks, DNA methylation and miR signatures. Translational studies or clinical trials, when available, and potential epigenetic biomarkers will be discussed as perspectives in the context of combination treatment strategies to enhance ICI responses in patients with solid tumors. Full article

(This article belongs to the Collection Recent Advances in Cancer Immunotherapy)

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13 pages, 1243 KiB

Open AccessArticle

Probing Structure and Function of Alkali Sensor IRR with Monoclonal Antibodies

by Alexander S. Goryashchenko, Andrey A. Mozhaev, Oxana V. Serova, Tatiana N. Erokhina, Alexander N. Orsa, Igor E. Deyev and Alexander G. Petrenko

Biomolecules 2020, 10(7), 1060; https://doi.org/10.3390/biom10071060 - 16 Jul 2020

Cited by 5 | Viewed by 2408

Abstract

To study the structure and function of the pH-regulated receptor tyrosine kinase insulin receptor-related receptor (IRR), а member of the insulin receptor family, we obtained six mouse monoclonal antibodies against the recombinant IRR ectodomain. These antibodies were characterized in experiments with exogenously expressed [...] Read more.

To study the structure and function of the pH-regulated receptor tyrosine kinase insulin receptor-related receptor (IRR), а member of the insulin receptor family, we obtained six mouse monoclonal antibodies against the recombinant IRR ectodomain. These antibodies were characterized in experiments with exogenously expressed full-length IRR by Western blotting, immunoprecipitation, and immunocytochemistry analyses. Utilizing a previously obtained set of IRR/IR chimeras with swapped small structural domains and point amino acid substitutions, we mapped the binding sites of the obtained antibodies in IRR. Five of them showed specific binding to different IRR domains in the extracellular region, while one failed to react with the full-length receptor. Unexpectedly, we found that 4D5 antibody can activate IRR at neutral pH, and 4C2 antibody can inhibit activation of IRR by alkali. Our study is the first description of the instruments of protein nature that can regulate activity of the orphan receptor IRR and confirms that alkali-induced activation is an intrinsic property of this receptor tyrosine kinase. Full article

(This article belongs to the Section Molecular Biology)

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30 pages, 4611 KiB

Open AccessArticle

Inflammatory Breast Carcinoma: Elevated microRNA miR-181b-5p and Reduced miR-200b-3p, miR-200c-3p, and miR-203a-3p Expression as Potential Biomarkers with Diagnostic Value

by Sarah Atef Fahim, Mahmoud Salah Abdullah, Nancy A. Espinoza-Sánchez, Hebatallah Hassan, Ayman M. Ibrahim, Sarah Hamdy Ahmed, George Shakir, Mohamed A. Badawy, Nadia I. Zakhary, Burkhard Greve, Mohamed El-Shinawi, Martin Götte and Sherif Abdelaziz Ibrahim

Biomolecules 2020, 10(7), 1059; https://doi.org/10.3390/biom10071059 - 16 Jul 2020

Cited by 21 | Viewed by 4592

Abstract

Inflammatory breast cancer (IBC) is a rare yet aggressive breast cancer variant, associated with a poor prognosis. The major challenge for IBC is misdiagnosis due to the lack of molecular biomarkers. We profiled dysregulated expression of microRNAs (miRNAs) in primary samples of IBC [...] Read more.

Inflammatory breast cancer (IBC) is a rare yet aggressive breast cancer variant, associated with a poor prognosis. The major challenge for IBC is misdiagnosis due to the lack of molecular biomarkers. We profiled dysregulated expression of microRNAs (miRNAs) in primary samples of IBC and non-IBC tumors using human breast cancer miRNA PCR array. We discovered that 28 miRNAs were dysregulated (10 were upregulated, while 18 were underexpressed) in IBC vs. non-IBC tumors. We identified 128 hub genes, which are putative targets of the differentially expressed miRNAs and modulate important cancer biological processes. Furthermore, our qPCR analysis independently verified a significantly upregulated expression of miR-181b-5p, whereas a significant downregulation of miR-200b-3p, miR-200c-3p, and miR-203a-3p was detected in IBC tumors. Receiver operating characteristic (ROC) curves implied that the four miRNAs individually had a diagnostic accuracy in discriminating patients with IBC from non-IBC and that miR-203a-3p had the highest diagnostic value with an AUC of 0.821. Interestingly, a combination of miR-181b-5p, miR-200b-3p, and miR-200c-3p robustly improved the diagnostic accuracy, with an area under the curve (AUC) of 0.897. Intriguingly, qPCR revealed that the expression of zinc finger E box-binding homeobox 2 (ZEB2) mRNA, the putative target of miR-200b-3p, miR-200c-3p, and miR-203a-3p, was upregulated in IBC tumors. Overall, this study identified a set of miRNAs serving as potential biomarkers with diagnostic relevance for IBC. Full article

(This article belongs to the Special Issue microRNA Biomarkers in Clinical Study)

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17 pages, 3619 KiB

Open AccessArticle

Machine Learning-Empowered FTIR Spectroscopy Serum Analysis Stratifies Healthy, Allergic, and SIT-Treated Mice and Humans

by Elke Korb, Murat Bağcıoğlu, Erika Garner-Spitzer, Ursula Wiedermann, Monika Ehling-Schulz and Irma Schabussova

Biomolecules 2020, 10(7), 1058; https://doi.org/10.3390/biom10071058 - 16 Jul 2020

Cited by 14 | Viewed by 4467

Abstract

The unabated global increase of allergic patients leads to an unmet need for rapid and inexpensive tools for the diagnosis of allergies and for monitoring the outcome of allergen-specific immunotherapy (SIT). In this proof-of-concept study, we investigated the potential of Fourier-Transform Infrared (FTIR) [...] Read more.

The unabated global increase of allergic patients leads to an unmet need for rapid and inexpensive tools for the diagnosis of allergies and for monitoring the outcome of allergen-specific immunotherapy (SIT). In this proof-of-concept study, we investigated the potential of Fourier-Transform Infrared (FTIR) spectroscopy, a high-resolution and cost-efficient biophotonic method with high throughput capacities, to detect characteristic alterations in serum samples of healthy, allergic, and SIT-treated mice and humans. To this end, we used experimental models of ovalbumin (OVA)-induced allergic airway inflammation and allergen-specific tolerance induction in BALB/c mice. Serum collected before and at the end of the experiment was subjected to FTIR spectroscopy. As shown by our study, FTIR spectroscopy, combined with deep learning, can discriminate serum from healthy, allergic, and tolerized mice, which correlated with immunological data. Furthermore, to test the suitability of this biophotonic method for clinical diagnostics, serum samples from human patients were analyzed by FTIR spectroscopy. In line with the results from the mouse models, machine learning-assisted FTIR spectroscopy allowed to discriminate sera obtained from healthy, allergic, and SIT-treated humans, thereby demonstrating its potential for rapid diagnosis of allergy and clinical therapeutic monitoring of allergic patients. Full article

(This article belongs to the Special Issue Application of Artificial Intelligence for Medical Research)

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28 pages, 3716 KiB

Open AccessArticle

The Constitutive Lack of α7 Nicotinic Receptor Leads to Metabolic Disorders in Mouse

by Blandine Gausserès, Junjun Liu, Ewout Foppen, Cécile Tourrel-Cuzin, Ana Rodriguez Sanchez-Archidona, Etienne Delangre, Céline Cruciani-Guglielmacci, Stéphanie Pons, Uwe Maskos, Bernard Thorens, Christophe Magnan, Jamileh Movassat and Kamel Maouche

Biomolecules 2020, 10(7), 1057; https://doi.org/10.3390/biom10071057 - 16 Jul 2020

Cited by 11 | Viewed by 3412

Abstract

Objective: Type 2 diabetes (T2D) occurs by deterioration in pancreatic β-cell function and/or progressive loss of pancreatic β-cell mass under the context of insulin resistance. α7 nicotinic acetylcholine receptor (nAChR) may contribute to insulin sensitivity but its role in the pathogenesis of T2D [...] Read more.

Objective: Type 2 diabetes (T2D) occurs by deterioration in pancreatic β-cell function and/or progressive loss of pancreatic β-cell mass under the context of insulin resistance. α7 nicotinic acetylcholine receptor (nAChR) may contribute to insulin sensitivity but its role in the pathogenesis of T2D remains undefined. We investigated whether the systemic lack of α7 nAChR was sufficient to impair glucose homeostasis. Methods: We used an α7 nAChR knock-out (α7^−/−) mouse model fed a standard chow diet. The effects of the lack of α7 nAChR on islet mass, insulin secretion, glucose and insulin tolerance, body composition, and food behaviour were assessed in vivo and ex vivo experiments. Results: Young α7^−/− mice display a chronic mild high glycemia combined with an impaired glucose tolerance and a marked deficit in β-cell mass. In addition to these metabolic disorders, old mice developed adipose tissue inflammation, elevated plasma free fatty acid concentrations and presented glycolytic muscle insulin resistance in old mice. Finally, α7^−/− mice, fed a chow diet, exhibited a late-onset excessive gain in body weight through increased fat mass associated with higher food intake. Conclusion: Our work highlights the important role of α7 nAChR in glucose homeostasis. The constitutive lack of α7 nAChR suggests a novel pathway influencing the pathogenesis of T2D. Full article

(This article belongs to the Special Issue Pancreatic Islets of Langerhans: Not Only Beta-Cells)

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Biomolecules, Volume 10, Issue 7 (July 2020) – 119 articles

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