Identification of driving genes of familial adenomatous polyposis by differential gene expression analysis and weighted gene co-expression network analysis.

BACKGROUND: Despite the advancement of new screening strategies and the advances in pharmacological therapies, the cancerization rates of familial adenomatous polyposis (FAP) are stable and even increased in the last years. Therefore, it necessitates additional research to characterize and understand the underlying mechanisms of FAP. OBJECTIVE: To determine the genes that drive the pathogenesis of familial adenomatous polyposis (FAP). METHODS: We performed on a cohort (GSE111156) gene profile, which consist of four group of gene expressions (the gene expressions of cancer, adenoma and normal tissue of duodenal cancer from patients with FAP were defined as Case N, Case A and Case C respectively, while that of adenoma tissue from patients with FAP who did not have duodenal cancer was Ctrl A). Tracking Tumor Immunophenotype (TIP) website was applied to reveal immune infiltration profile and signature genes of FAP. We merged the genes of key module (pink and midnight module) with signature genes to obtained the biomarkers related with FAP pathogenesis. The expression of these five biomarkers in FAP intratumoral region (IT) and tumor rim (TR) was detected with Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR). RESULTS: In total, 220, 23 and 63 DEGs were determined in Cases C, A and N, in comparison to Ctrl A. In total, 196 and 10 DEGs were determined in Cases C and A, separately, as compared to Case N. A total of four biomarkers including CCL5, CD3G, CD2 and TLR3 were finally identified associated with pink module, while only one biomarker (KLF2) associated with midnight module was identified. All biomarkers were evidently raised in FAP IT tissues utilizing qRT-PCR. CONCLUSION: We identified five potential biomarkers for pathogenesis of FAP to understand the fundamental mechanisms of FAP progression and revealed some probable targets for the diagnosis or treatment of FAP.

Somatic mutation profiling, tumor-infiltrating leukocytes, tertiary lymphoid structures and PD-L1 protein expression in HER2-amplified colorectal cancer.

The status of human epidermal growth factor receptor 2 (HER2) for the prognosis in colorectal cancer (CRC) is controversial, and the characteristics of the somatic mutation spectrum, tumor-infiltrating leukocytes, tertiary lymphoid structures and PD-L1 protein are unknown in HER2-amplified colorectal cancer (HACC). In order to explore these characteristics along with their correlation with clinicopathological factors and prognosis in HACC. Samples of 812 CRC patients was collected. After immunohistochemistry (IHC), 59 of 812 were found to be HER2-positive, then 26 of 59 samples were further determined to be HER2 amplification by fluorescence in situ hybridization (FISH). Somatic mutation profiling of HACC was analysed using whole exome sequencing (WES). Multiplex fluorescence immunohistochemistry (mIHC) was used for tumor-infiltrating leukocytes and tertiary lymphoid structures (TLSs), while PD-L1 protein was detected by IHC. Our results indicate that the detection rates of HER2 positivity by IHC and FISH were 7.3% and 3.2% respectively, and HER2 amplification is correlated with distant tumour metastasis. The somatic mutation profiling revealed no differences between HACC and HER2-negative CRC. However, TP 53 strongly correlated with poor prognosis in HACC. Furthermore, tumor-infiltrating T cells and TLSs in the tumor immune microenvironment, as well as PD-L1 expression, were higher in HACC than in HER2-negative controls. However, none of them were associated with the prognosis of HACC. In all, HER2 amplification is correlated with distant metastasis and TP53 gene mutation may be a potential protective mechanism of HACC.

Rare-Earth-Zirconate Porous High-Entropy Ceramics with Unique Pore Structures for Thermal Insulating Applications.

Porous high-entropy ceramics are a new alternative material for thermal insulation. Their better stability and low thermal conductivity are due to lattice distortion and unique pore structures. In this work, rare-earth-zirconate ((La0.25Eu0.25Gd0.25Yb0.25)2(Zr0.75Ce0.25)2O7) porous high-entropy ceramics were fabricated by a tert-butyl alcohol (TBA)-based gel-casting method. The regulation of pore structures was realized through changing different initial solid loadings. The XRD, HRTEM, and SAED results showed that the porous high-entropy ceramics had a single fluorite phase without impurity phases, exhibiting high porosity (67.1-81.5%), relatively high compressive strength (1.02-6.45 MPa) and low thermal conductivity (0.0642-0.1213 W/(m·K)) at room temperature. Porous high-entropy ceramics with 81.5% porosity demonstrated excellent thermal properties, showing a thermal conductivity of 0.0642 W/(m·K) at room temperature and 0.1467 W/(m·K) at 1200 °C. The unique pore structure with a micron size contributed to their excellent thermal insulating performance. The present work provides the prospect that rare-earth-zirconate porous high-entropy ceramics with tailored pore structures are expected to be thermal insulation materials.

STAU1 promotes adipogenesis by regulating the alternative splicing of Pparγ2 mRNA.

During adipocyte differentiation, specific genes such as peroxisome proliferator-activated receptor γ (PPARγ) are transcribed and post-transcriptional pre-mRNA is processed into mature mRNA. Since Pparγ2 pre-mRNAs contain putative binding sites for STAUFEN1 (STAU1), which can affect the alternative splicing of pre-mRNA, we hypothesized that STAU1 might regulate the alternative splicing of Pparγ2 pre-mRNA. In this study, we found that STAU1 affects the differentiation of 3 T3-L1 pre-adipocytes. Through RNA-seq analysis, we confirmed that STAU1 can regulate alternative splicing events during adipocyte differentiation, mainly through exon skipping, which suggests that STAU1 is mainly involved in exon splicing. In addition, gene annotation and cluster analysis revealed that the genes affected by alternative splicing were enriched in lipid metabolism pathways. We further demonstrated that STAU1 can regulate the alternative splicing of Pparγ2 pre-mRNA and affect the splicing of exon E1 through RNA immuno-precipitation, photoactivatable ribonucleotide enhanced crosslinking and immunoprecipitation, and sucrose density gradient centrifugation assays. Finally, we confirmed that STAU1 can regulate the alternative splicing of Pparγ2 pre-mRNA in stromal vascular fraction cells. In summary, this study improves our understanding of the function of STAU1 in adipocyte differentiation and the regulatory network of adipocyte differentiation-related gene expression.

Interferon regulatory factor 1-triggered free ubiquitin protects the intestines against radiation-induced injury via CXCR4/FGF2 signaling.

Radiation-induced intestinal injury is a serious concern during abdominal and pelvic cancers radiotherapy. Ubiquitin (Ub) is a highly conserved protein found in all eukaryotic cells. This study aims to explore the role and mechanism of free Ub against radiogenic intestinal injury. We found that free Ub levels of irradiated animals and human patients receiving radiotherapy were upregulated. Radiation-induced Ub expression was associated with the activation of interferon regulatory factor 1 (IRF1). Intraperitoneal injection of free Ub significantly reduced the mortality of mice following 5-9 Gy total body irradiation (TBI) through the Akt pathway. Free Ub facilitates small intestinal regeneration induced by TBI or abdominal irradiation. At the cellular level, free Ub or its mutants significantly alleviated cell death and enhanced the survival of irradiated intestinal epithelial cells. The radioprotective role of free Ub depends on its receptor CXCR4. Mechanistically, free Ub increased fibroblast growth factor-2 (FGF2) secretion and consequently activated FGFR1 signaling following radiation in vivo and in vivo. Thus, free Ub confers protection against radiation-induced intestinal injury through CXCR4/Akt/FGF2 axis, which provides a novel therapeutic option.

The Molecular Mechanism of Human Voltage-Dependent Anion Channel 1 Blockade by the Metallofullerenol Gd@C82(OH)22: An In Silico Study.

The endohedral metallofullerenol Gd@C82(OH)22 has been identified as a possible antineoplastic agent that can inhibit both the growth and metastasis of cancer cells. Despite these potentially important effects, our understanding of the interactions between Gd@C82(OH)22 and biomacromolecules remains incomplete. Here, we study the interaction between Gd@C82(OH)22 and the human voltage-dependent anion channel 1 (hVDAC1), the most abundant porin embedded in the mitochondrial outer membrane (MOM), and a potential druggable target for novel anticancer therapeutics. Using in silico approaches, we observe that Gd@C82(OH)22 molecules can permeate and form stable interactions with the pore of hVDAC1. Further, this penetration can occur from either side of the MOM to elicit blockage of the pore. The binding between Gd@C82(OH)22 and hVDAC1 is largely driven by long-range electrostatic interactions. Analysis of the binding free energies indicates that it is thermodynamically more favorable for Gd@C82(OH)22 to bind to the hVDAC1 pore when it enters the channel from inside the membrane rather than from the cytoplasmic side of the protein. Multiple factors contribute to the preferential penetration, including the surface electrostatic landscape of hVDAC1 and the unique physicochemical properties of Gd@C82(OH)22. Our findings provide insights into the potential molecular interactions of macromolecular biological systems with the Gd@C82(OH)22 nanodrug.

The prognostic and clinicopathological roles of microsatellite instability, PD-L1 expression and tumor-infiltrating leukocytes in familial adenomatous polyposis.

BACKGROUND: Microsatellite instability, programmed death-ligand 1 and tumor-infiltrating leukocytes are prognostic biomarkers in colorectal cancer but unknown toward familial adenomatous polyposis. AIM: To investigate the prognostic and clinicopathological roles of microsatellite instability, programmed death-ligand 1 and tumor-infiltrating leukocytes in familial adenomatous polyposis. METHODS: Clinical data and paraffin embedded tissues from 45 familial adenomatous polyposis patients were collected. Microsatellite instability was detected by immunohistochemistry and polymerase chain reaction. Programmed death-ligand 1 was detected by immunohistochemistry. Tumor-infiltrating leukocytes comprising CD8+ T cells, M1 and M2 tumor associated macrophages, CD56bright and CD56dim natural killer cells were analyzed using multiple fluorescence immunohistochemistry. RESULTS: Microsatellite instability high was noted in 6 samples but not associated with overall survival or progression-free survival. Programmed death-ligand 1 is negative on tumor cells but positive on tumor-infiltrating leukocytes, and positive programmed death-ligand 1 expression on tumor-infiltrating leucocytes is associated with overall survival. Low CD56bright natural killer cell infiltration was associated with longer progression-free survival and was an independent prognostic factor in FAP. CONCLUSION: For familial adenomatous polyposis, microsatellite instability high can be found but has no correlation with prognosis; programmed death-ligand 1 on tumor-infiltrating leukocytes is related with overall survival; CD56bright natural killer cell is an independent prognostic factor associating with longer progression-free survival.

Staufen1 unwinds the secondary structure and facilitates the translation of fatty acid binding protein 4 mRNA during adipogenesis.

Adipogenesis is regulated by genetic interactions, in which post-transcriptional regulation plays an important role. Staufen double-stranded RNA binding protein 1 (Staufen1 or STAU1) plays diverse roles in RNA processing and adipogenesis. Previously, we found that the downregulation of STAU1 affects the expression of fatty acid-binding protein 4 (FABP4) at the protein level but not at the mRNA level. This study aimed to determine the mechanism underlying the regulation of FABP4 expression by STAU1, explaining the inconsistency between FABP4 mRNA and protein levels. We used RNA interference, photoactivatable ribonucleoside enhanced cross-linking and immunoprecipitation, and an adeno-associated virus to examine the functions of STAU1 in adipogenesis. Our results indicate that STAU1 binds to the coding sequences of FABP4, thereby regulating the translation of FABP4 mRNA by unwinding the double-stranded structure. Furthermore, STAU1 mediates adipogenesis by regulating the secretion of free fatty acids. However, STAU1 knockdown decreases the fat weight/body weight ratio but does not affect the plasma triglyceride levels. These findings describe the mechanisms involved in STAU1-mediated regulation of FABP4 expression at the translational level during adipogenesis.

Multifaceted Regulation of Potassium-Ion Channels by Graphene Quantum Dots.

Graphene quantum dots (GQDs) are emerging as a versatile nanomaterial with numerous proposed biomedical applications. Despite the explosion in potential applications, the molecular interactions between GQDs and complex biomolecular systems, including potassium-ion (K+) channels, remain largely unknown. Here, we use molecular dynamics (MD) simulations and electrophysiology to study the interactions between GQDs and three representative K+ channels, which participate in a variety of physiological processes and are closely related to many disease states. Using MD simulations, we observed that GQDs adopt distinct contact poses with each of the three structurally distinct K+ channels. Our electrophysiological characterization of the effects of GQDs on channel currents revealed that GQDs interact with the extracellular voltage-sensing domain (VSD) of a Kv1.2 channel, augmenting current by left-shifting the voltage dependence of channel activation. In contrast, GQDs form a "lid" cluster over the extracellular mouth of inward rectifier Kir3.2, blocking the channel pore and decreasing the current in a concentration-dependent manner. Meanwhile, GQDs accumulate on the extracellular "cap domain" of K2P2 channels and have no apparent impact on the K+ flux through the channel. These results reveal a surprising multifaceted regulation of K+ channels by GQDs, which might help de novo design of nanomaterial-based channel probe openers/inhibitors that can be used to further discern channel function.

Molecular Dynamics Simulation Study on Interactions of Cycloviolacin with Different Phospholipids.

Cyclotides are disulfide-rich cyclic peptides isolated from plants, which are extremely stable against thermal and proteolytic degradation, with a variety of biological activities including antibacterial, hemolytic, anti-HIV, and anti-tumor. Most of these bioactivities are related to their preference for binding to certain types of phospholipids and subsequently disrupt lipid membranes. In the present study, we use a cyclotide, cycloviolacin O2 (cyO2), as a model system to investigate its interactions with three lipid bilayers 1-palmitoyl-2-oleoylphosphatidylethanolamine (POPE), 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol (POPG)-doped POPE, and 1-palmitoyl-2-oleoylphosphatidylcholine (POPC), to help understand its potential mechanism of action toward the membranes at the molecular level using molecular dynamics simulations. In our simulations, cyO2 repeatedly forms stable binding complexes with the POPE-containing bilayers, while within the same simulation time scale, it "jumps" back and forth on the surface of the POPC bilayer without a strong binding. Detailed analyses reveal that the electrostatic attraction is the main driving force for the initial bindings between cyO2 and the lipids, but with strikingly different strengths in different bilayers. For the POPE-containing bilayers, the charged residues of cyO2 attract both POPE amino and phosphate head groups favorably; meanwhile, its hydrophobic residues are deeply inserted into the lipid hydrophobic tails (core) of the membrane, thus forming stable binding complexes. In contrast, POPC lipids with three methyl groups on the amino head group create a steric hindrance when interacting with cyO2, thus resulting in a relatively difficult binding of cyO2 on POPC compared to POPE. Our current findings provide additional insights for a better understanding of how cyO2 binds to the POPE-containing membrane, which should shed light on the future cyclotide-based antibacterial agent design.

Ad36 promotes differentiation of hADSCs into brown adipocytes by up-regulating LncRNA ROR.

AIMS: This study is to investigate the role of adenovirus type 36 (Ad36) in inducing differentiation of human adipose-derived stem cells (hADSCs) into brown adipocytes. MAIN METHODS: The hADSCs were induced to differentiate into adipocytes by a cocktail method and Ad36, respectively. They were collected on the 2nd, 4th, 6th, and 8th day, respectively. LncRNA ROR was silenced by siRNA. RT-qPCR and Western-blot were used to detect the mRNA and protein levels. Transmission electron microscopy was used to observe the mitochondria. KEY FINDINGS: The mRNA and protein expression levels of LncRNA ROR, Cidea, Dio2, Fgf21, Ucp1, Prdm16, Cox5b, Atp5o, Atp6, and Nd2 in the Ad36 induction group were significantly higher than those in the cocktail induction group. The expression levels of Leptin mRNA and protein in the Ad36 induction group were significantly lower than those in the cocktail induction group. After siRNA knockdown of LncRNA ROR, mRNA and protein expression levels of Cidea, Dio2, Fgf21, Ucp1, Prdm16, Cox5b, Atp5o, Atp6 and Nd2 were significantly lower than the control group during the induction of hADSC differentiation into adipocytes by Ad36. Additionally, mitochondria in the Ad36 induction group was increased compared to that in the cocktail induction group. SIGNIFICANCE: Ad36 may promote the differentiation of hADSCs into brown adipocytes by up-regulating LncRNA ROR.

Binding patterns and dynamics of double-stranded DNA on the phosphorene surface.

Phosphorene, a monolayer of black phosphorus, has emerged as one of the most promising two-dimensional (2D) nanomaterials for various applications in the post-graphene-discovery period due to its highly anisotropic structure and novel properties. In order to apply phosphorene in biomedical fields, it is crucial to understand how it interacts with biomolecules. Herein, we use both molecular dynamics (MD) simulations and experimental techniques to investigate the interactions of phosphorene with a dsDNA segment. Our results reveal that dsDNA can form a stable binding on the phosphorene surface through the terminal base pairs and adopt an upright orientation regardless of its initial configurations. Moreover, the binding strength of dsDNA with phosphorene is found to be mild and does not cause significant distortion in the internal structure of dsDNA. This phenomenon is attributed to the weaker dispersion interaction between dsDNA and phosphorene. Further analysis of the free energy profile calculated by the umbrella sampling technique suggests that the puckered surface morphology significantly reduces the adsorption free energy of DNA bases to phosphorene. Compared to graphene, phosphorene is found to show a milder attraction to DNA, which is confirmed by our electrophoresis experiments. We believe that these findings provide valuable insight into the molecular interactions between phosphorene and dsDNA which may prompt further investigation of phosphorene for future biomedical applications.

Potential blockade of the human voltage-dependent anion channel by MoS2 nanoflakes.

Despite significant interest in molybdenum disulfide (MoS2) nanomaterials, particularly in biomedicine, their biological effects have been understudied. Here, we explored the effect of MoS2 nanoflakes on the ubiquitous mitochondrial porin voltage-dependent anion channel (VDAC1), using a combined computational and functional approach. All-atomic molecular dynamics simulations suggest that MoS2 nanoflakes make specific contact interactions with human VDAC1. We show that the initial contacts between hVDAC1 and the nanoflake are hydrophobic but are subsequently enhanced by a complex interplay of van der Waals (vdW), hydrophobic and electrostatic interactions in the equilibrium state. Moreover, the MoS2 nanoflake can insert into the lumen of the hVDAC1 pore. Free-energy calculations computed by the potential of mean force (PMF) verify that the blocked configuration of the MoS2-hVDAC1 complex is more energetically favorable than the non-blocked binding mode. Consistent with these predictions, we showed that MoS2 depolarizes the mitochondrial membrane potential (Ψm) and causes a decrease in the viability of mammalian tissue culture cells. These findings might shed new light on the potential biological effect of MoS2 nanomaterials.

Molecular mechanism of phosphoinositides' specificity for the inwardly rectifying potassium channel Kir2.2.

Phosphoinositides are essential signaling lipids that play a critical role in regulating ion channels, and their dysregulation often results in fatal diseases including cardiac arrhythmia and paralysis. Despite decades of intensive research, the underlying molecular mechanism of lipid agonism and specificity remains largely unknown. Here, we present a systematic study of the binding mechanism and specificity of a native agonist, phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) and two of its variants, PI(3,4)P2 and PI(3,4,5)P3, on inwardly rectifying potassium channel Kir2.2, using molecular dynamics simulations and free energy perturbations (FEPs). Our results demonstrate that the major driving force for the PI(4,5)P2 specificity on Kir2.2 comes from the highly organized salt-bridge network formed between the charged inositol head and phosphodiester linker of PI(4,5)P2. The unsaturated arachidonic chain is also shown to contribute to the stable binding through hydrophobic interactions with nearby Kir2.2 hydrophobic residues. Consistent with previous experimental findings, our FEP results confirmed that non-native ligands, PI(3,4)P2 and PI(3,4,5)P3, show significant loss in binding affinity as a result of the substantial shift from the native binding mode and unfavorable local solvation environment. However, surprisingly, the underlying molecular pictures for the unfavorable binding of both ligands are quite distinctive: for PI(3,4)P2, it is due to a direct destabilization in the bound state, whereas for PI(3,4,5)P3, it is due to a relative stabilization in its free state. Our findings not only provide a theoretical basis for the ligand specificity, but also generate new insights into the allosteric modulation of ligand-gated ion channels.

Dynamic imine bond cross-linked self-healing thermosensitive hydrogels for sustained anticancer therapy via intratumoral injection.

In this study, we developed a self-healing thermosensitive gel, via dialdehyde-functionalized polyethylene glycol (DF-PEG) and ß-glycerophosphate (GP) cross-linked chitosan (CS) hydrogels, which enable autonomous self-healing upon damage and sustained release of doxorubicin hydrochloride (DOX) for antitumor therapy via intratumoral injection. The cross-linked gels could exhibit sol-gel transition at 37 °C within 5 min and satisfactory in vitro and in vivo healing ability by observing the rejoining and fusion process of scratched gels. Moreover, the in vitro release of DOX loaded cross-linked gels in PBS (pH 6.5 and 7.4) was found to be extended to 13 d. After intratumoral injection in Heps tumor-bearing mice, the drug loaded self-healing thermosensitive gels showed a superior tumor inhibition rate (66.12%) than CS thermosensitive hydrogels (53.23%), while the histology studies gave the relieved cardiotoxicity and good biocompatibility of the cross-linked gels. Overall, these cross-linked gels could become a potential local drug delivery system to achieve efficient sustained release, relieved side effects and enhanced therapy efficiency through localized administration.

The Heptahelical Domain of the Sweet Taste Receptor T1R2 Is a New Allosteric Binding Site for the Sweet Taste Modulator Amiloride That Modulates Sweet Taste in a Species-Dependent Manner.

The activity of sweet taste receptor (heterodimeric T1R2 and T1R3) can be modulated by sweet regulators. The compound amiloride can inhibit the sweet sensitivity of the human sweet taste receptor. This study describes the species-dependent regulation of the response of sweet taste receptors by this sweet inhibitor. Amiloride inhibited the sweet taste response of humans and mice but not that of squirrel monkeys. Using human/squirrel monkey/mouse chimeric T1R2 and T1R3 receptors as well as the agonist perillartine (which can activate the single heptahelical domain of T1R2), we found that the heptahelical domain of T1R2 is the molecular determinant that mediates the species-dependent sensitivity to this sweet regulator. Compared to the sweet inhibitor lactisole (which acts on T1R3), amiloride has a different allosteric binding site on the sweet receptor, which is important new information for the design of novel sweet taste modulators that act on T1R2.

Effect of lncRNA HOXA11-AS1 on adipocyte differentiation in human adipose-derived stem cells.

AIMS: To determine the role of lncRNA HOXA11-AS1 on adipocyte differentiation. METHODS: Human adipose-derived stem cells (hADSCs) were isolated from adipose tissues of patients and cultured in vitro, followed by knockdown of HOXA11-AS1. Then, adipocyte differentiation and expression of adipogenic-related genes (CEBP-α, DGAT2, CIDEC, and perilipin) were measured by RT-qPCR and Western blot. RESULTS: We demonstrated that knockdown of HOXA11-AS1 inhibited adipocyte differentiation, leading to suppression of adipogenic-related gene (CEBP-α, DGAT2, CIDEC, and perilipin) transcription, as well as decreased lipid accumulation in hADSCs. In addition, lncRNA HOXA11-AS1 was highly expressed in obese patients and significantly increased during the process of adipocyte differentiation. CONCLUSION: The results provide new insight into the molecular mechanism by which lncRNA HOXA11-AS1 is involved in adipogenesis and may have implications for the treatment of obesity and associated disorders.

Exploring the Nanotoxicology of MoS2: A Study on the Interaction of MoS2 Nanoflakes and K+ Channels.

Molybdenum disulfide (MoS2) nanomaterial has recently found various applications in the biomedical field mainly due to its outstanding physicochemical properties. However, little is known about its interactions with biological systems at the atomic level, which intimately relates to the biocompatibility of the material. To provide insights into the effects of MoS2 in biological entities, we investigated the interactions of MoS2 with proteins from a functionally important membrane family, the ubiquitous potassium (K+) channels. Here, we study four representative K+ channels-KcsA, Kir3.2, the Kv1.2 paddle chimera, and K2P2-to investigate their interactions with a triangular MoS2 nanoflake using Molecular Dynamics (MD) simulations combined with electrophysiology experiments. These particular K+ channels were selected based on the diversity in their structure; that is, although these K+ channels display similar structural motifs, they also contain significant differences related to their particular function. Our results indicate that the MoS2 nanoflake is able to stably bind to three out of the four channels, albeit through distinct binding modes. The binding mode between each channel and MoS2 underlies the specific deleterious influence on the channel's basic physiological function: For KcsA, MoS2 binds on the extracellular loops, which indirectly destroys the delicate structure of the selectivity filter causing a strong leak of K+ ions. In the binding mode with Kir3.2, the MoS2 nanoflake completely covers the entrance to the channel pore affecting the normal ion conduction. For the Kv1.2 chimera, the MoS2 nanoflake prefers to bind into a crevice located at the extracellular side of the Voltage Sensor Domain (VSD). Interestingly, the crevice involves the N-terminal segment of S4, a crucial transmembrane helix which directly controls the gating process of the Kv1.2 chimera channel by electromechanical coupling the VSD to the transmembrane electric field. MoS2 in contact with S4 from the Kv1.2 chimera, potentially influences the channel's gating process from open to closed states. In all three systems, the van der Waals contribution to the total energy dominates the binding interactions; also, hydrophobic residues contribute the most contact points, which agrees with the strong hydrophobic character of the MoS2 nanomaterial. Electrophysiology recordings using two-electrode voltage-clamp show that currents of Kir3.2 and Kv1.2 are both blocked by the MoS2 nanoflakes in a concentration-dependent way. While the background K+ channel, K2P2 (TREK-1), identified as a negative control, is not blocked by the MoS2 nanoflakes. The large and rigid extracellular domain of K2P2 appears to protect the channel from disturbance by the nanoflakes. Intrinsic chemical properties of MoS2, together with the specific features of the channels, such as the electrostatic character and complex surface architecture, determine the critical details of the binding events. These findings might shed light on the potential nanotoxicology of MoS2 nanomaterials and help us to understand the underlying molecular mechanism.

An In Silico study of TiO2 nanoparticles interaction with twenty standard amino acids in aqueous solution.

Titanium dioxide (TiO2) is probably one of the most widely used nanomaterials, and its extensive exposure may result in potentially adverse biological effects. Yet, the underlying mechanisms of interaction involving TiO2 NPs and macromolecules, e.g., proteins, are still not well understood. Here, we perform all-atom molecular dynamics simulations to investigate the interactions between TiO2 NPs and the twenty standard amino acids in aqueous solution exploiting a newly developed TiO2 force field. We found that charged amino acids play a dominant role during the process of binding to the TiO2 surface, with both basic and acidic residues overwhelmingly preferred over the non-charged counterparts. By calculating the Potential Mean Force, we showed that Arg is prone to direct binding onto the NP surface, while Lys needs to overcome a ~2 kT free energy barrier. On the other hand, acidic residues tend to form "water bridges" between their sidechains and TiO2 surface, thus displaying an indirect binding. Moreover, the overall preferred positions and configurations of different residues are highly dependent on properties of the first and second solvation water. These molecular insights learned from this work might help with a better understanding of the interactions between biomolecules and nanomaterials.

The Molecular Mechanism of Opening the Helix Bundle Crossing (HBC) Gate of a Kir Channel.

Inwardly rectifying K(+) (Kir) channels, serving as natural molecular nanomachines, transport potassium ions across the plasma membrane of the cell. Along the ion permeation pathway, three relatively narrow regions (the selectivity filter (SF), the inner helix bundle crossing (HBC), and the cytosolic G loop) may serve as gates to control ion permeation. Our previous molecular dynamics simulations based on the crystal structure of a Kir3.1 chimera revealed the possible gating mechanism of the G loop gate. Here, we introduced a proline mutation in the inner helix and obtained a channel model of the open HBC gate. The open HBC gate reaches 0.6 nm in diameter, which allows partial hydrated K(+) ions to pass through. During the gating process, both the transmembrane helices TM1 and TM2 cooperatively rotate in a counterclockwise direction (viewed from the extracellular side) with the aid of the phospholipid PIP2. Only when all the transmembrane helices adopt a counterclockwise rotation, the HBC gate can be stabilized in the open state. We estimate that introduction of the proline mutation decreases the energy required to open the HBC gate by about 1.4 kcal/mol (ΔΔG).