Druggability properties of a L309K mutation in the antibody CH2 domain.

In the early stages of antibody drug development, it is imperative to conduct a comprehensive assessment and enhancement of the druggability attributes of potential molecules by considering their fundamental physicochemical properties. This study specifically concentrates on the surface-exposed hydrophobic region of the candidate antibody aPDL1-WT and explores the effectiveness of the L309K mutation strategy. The resulting aPDL1-LK variant demonstrates a notable enhancement over the original antibody in addressing the issue of aggregation and formation of large molecular impurities under accelerated high-temperature conditions. The mutated molecule, aPDL1-LK, exhibits excellent physicochemical properties such as hydrophilicity, conformational stability, charge variant stability, post-translational modifications, and serum stability. In terms of biological function, aPDL1-LK maintains the same glycosylation pattern as the original antibody and shows no significant difference in affinity for antigen hPDL1 protein, CD16a-F158, CD64, CD32a-H131, and complement C1q, compared to aPDL1-WT. The L309K mutation results in an approximately twofold reduction in its affinity for CD16a-V158 and CD32a-R131. In vitro biological assays, including antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-dependent cellular phagocytosis (ADCP), and complement-dependent cytotoxicity (CDC), reveal that the L309K mutation may decrease CD16a-V158-mediated ADCC activity due to the mutation-induced decrease in ligand affinity, while not affect CD32a-R131-mediated ADCP activity. In conclusion, the L309K mutation offers a promising strategy to enhance the druggability properties of candidate antibodies.

Integrated Metabolome and Transcriptome Analysis of Gibberellins Mediated the Circadian Rhythm of Leaf Elongation by Regulating Lignin Synthesis in Maize.

Plant growth exhibits rhythmic characteristics, and gibberellins (GAs) are involved in regulating cell growth, but it is still unclear how GAs crosstalk with circadian rhythm to regulate cell elongation. The study analyzed growth characteristics of wild-type (WT), zmga3ox and zmga3ox with GA3 seedlings. We integrated metabolomes and transcriptomes to study the interaction between GAs and circadian rhythm in mediating leaf elongation. The rates of leaf growth were higher in WT than zmga3ox, and zmga3ox cell length was shorter when proliferated in darkness than light, and GA3 restored zmga3ox leaf growth. The differentially expressed genes (DEGs) between WT and zmga3ox were mainly enriched in hormone signaling and cell wall synthesis, while DEGs in zmga3ox were restored to WT by GA3. Moreover, the number of circadian DEGs that reached the peak expression in darkness was more than light, and the upregulated circadian DEGs were mainly enriched in cell wall synthesis. The differentially accumulated metabolites (DAMs) were mainly attributed to flavonoids and phenolic acid. Twenty-two DAMs showed rhythmic accumulation, especially enriched in lignin synthesis. The circadian DEGs ZmMYBr41/87 and ZmHB34/70 were identified as regulators of ZmHCT8 and ZmBM1, which were enzymes in lignin synthesis. Furthermore, GAs regulated ZmMYBr41/87 and ZmHB34/70 to modulate lignin biosynthesis for mediating leaf rhythmic growth.

Synergistic regulation of chloride anion recognition using a triple-functional sites receptor with two different cationic effectors.

The synergistic regulation of the multi-functional sites on one receptor molecule with different cationic effectors for anion recognition is scarce to be well understood from the experiment and theory. In this work, a new anion receptor with three functional zones including ether hole, biurea and double bipyridine groups (EUPR) is designed expecting to enhance the chloride anion recognition together with a rational synthesis path being proposed based on four simple and mature organic reaction steps. The conformational structures of the designed receptor EUPR and the binding behaviors for three kinds of ions (Cl-, Na+, and Ag+) are deeply investigated by using density functional theoretical calculations. It is found that Cl- binding via the hydrogen bond interaction can be significantly enhanced and synergistically regulated by the two kinds of cations and the corresponding conformational changes of receptor EUPR. Especially, the conformational pre-organization of receptor caused by the encapsulation of sodium ion into ether hole is benefit to the binding for Cl- in both thermodynamics and kinetics. Na+ binding, in turn, can ever be enhanced by chloride anion, whereas it seems that Ag+ binding cannot always be enhanced by chloride anion, reflecting an electrical complementary matching and mutual enhancement effect for different counter ions. Moreover, solvent effect calculations indicate that EUPR may be an ideal candidate structure for Cl- recognition by strategy of counter ion enhancement in water. Additionally, a visual study of intermolecular noncovalent interaction (NCI) and molecular electrostatic potential (ESP) are used for the analysis on the nature of interactions between receptor and bound ions.

Pharmacokinetics, metabolism, excretion and safety of iruplinalkib (WX-0593), a novel ALK inhibitor, in healthy subjects: a phase I human radiolabeled mass balance study.

BACKGROUND: Iruplinalkib is a novel anaplastic lymphoma kinase (ALK) inhibitor for the treatment of ALK-positive crizotinib-resistant NSCLC. RESEARCH DESIGN AND METHODS: A single oral dose of 120 mg/3.7 MBq [14C]iruplinalkib was administered to healthy subjects. Blood, urine and fecal samples were collected and analyzed for iruplinalkib and its metabolites. The safety of iruplinalkib was also assessed. RESULTS: Iruplinalkib was absorbed quickly and eliminated slowly from plasma, with a Tmax of 1.5 h and t1/2 of 28.6 h. About 88.85% of iruplinalkib was excreted at 312 h, including 20.23% in urine and 68.63% in feces. Seventeen metabolites of iruplinalkib were identified, and M3b (demethylation), M7 (cysteine conjugation), M11 (oxidative dehydrogenation and cysteine conjugation of M3b) and M12 (oxidative dehydrogenation and cysteine conjugation) were considered the prominent metabolites in humans. Iruplinalkib-related compounds were found to be covalently bound to proteins, accounting for 7.70% in plasma and 17.96% in feces, which suggested chemically reactive metabolites were formed. There were no serious adverse events observed in the study. CONCLUSIONS: Iruplinalkib was widely metabolized and excreted mainly through feces in humans. Unchanged iruplinalkib, cysteine conjugates and covalent protein binding products were the main drug-related compounds in circulation. Iruplinalkib was well tolerated at the study dose. TRIAL REGISTRATION: The trial is registered at ClinicalTrials.gov (Identifier: Anonymized).

Cuproptosis-related gene SLC31A1: prognosis values and potential biological functions in cancer.

Cuproptosis is a unique type of cell death that may influence tumour formation by targeting lipoylated tricarboxylic acid cycle proteins. Solute carrier family 31 member 1 (SLC31A1), an important copper transporter, influences dietary copper absorption in the cell membrane. However, various SLC31A1 properties in pan-cancer profiles remain unknown. This study investigated the role of SLC31A1 in human malignancies and analysed its prognostic value. Raw data were obtained from The Cancer Genome Atlas database and processed using numerous internet databases, including UALCAN, GEPIA, cBioPortal, TIMER2.0, and Human Protein Atlas. SLC31A1 expression was found to be elevated in cervical, endometrial, and breast cancers compared to that in normal tissues, but reduced in clear cell renal cell carcinoma, liver hepatocellular carcinoma, and lung adenocarcinoma. Furthermore, SLC31A1 expression was strongly associated with overall survival and disease-free survival in several cancers. SLC31A1 gene mutations and methylations were identified in 33 cancers. SLC31A1 expression was positively correlated with immune cells in immune infiltration data. Single-cell sequencing revealed that SLC31A1 may play key roles in DNA repair, DNA damage, and proliferation. These findings may lead to better understanding of SLC31A1 in pan-cancer profiles and suggest that SLC31A1 could be a viable predictive biomarker, particularly in gynaecological cancers.

Expression of concern: Bacterial infection microenvironment-responsive enzymatically degradable multilayer films for multifunctional antibacterial properties.

Expression of concern for 'Bacterial infection microenvironment-responsive enzymatically degradable multilayer films for multifunctional antibacterial properties' by Qingqing Yao et al., J. Mater. Chem. B, 2017, 5, 8532-8541, https://doi.org/10.1039/C7TB02114C.

A Dual-Targeted Metal-Organic Framework Based Nanoplatform for the Treatment of Rheumatoid Arthritis by Restoring the Macrophage Niche.

Inflammatory infiltration and bone destruction are important pathological features of rheumatoid arthritis (RA), which originate from the disturbed niche of macrophages. Here, we identified a niche-disrupting process in RA: due to overactivation of complement, the barrier function of VSIg4+ lining macrophages is disrupted and mediates inflammatory infiltration within the joint, thereby activating excessive osteoclastogenesis and bone resorption. However, complement antagonists have poor biological applications due to superphysiologic dose requirements and inadequate effects on bone resorption. Therefore, we developed a dual-targeted therapeutic nanoplatform based on the MOF framework to achieve bone-targeted delivery of the complement inhibitor CRIg-CD59 and pH-responsive sustained release. The surface-mineralized zoledronic acid (ZA) of ZIF8@CRIg-CD59@HA@ZA targets the skeletal acidic microenvironment in RA, and the sustained release of CRIg-CD59 can recognize and prevent the complement membrane attack complex (MAC) from forming on the surface of healthy cells. Importantly, ZA can inhibit osteoclast-mediated bone resorption, and CRIg-CD59 can promote the repair of the VSIg4+ lining macrophage barrier to achieve sequential niche remodeling. This combination therapy is expected to treat RA by reversing the core pathological process, circumventing the pitfalls of traditional therapy.

Targeting ZDHHC9 potentiates anti-programmed death-ligand 1 immunotherapy of pancreatic cancer by modifying the tumor microenvironment.

Immune checkpoint blockade (ICB) therapy targeting the programmed death 1/programmed death-ligand 1 (PD-1/PD-L1) axis has achieved considerable success in treating a wide range of cancers. However, most patients with pancreatic cancer remain resistant to ICB. Moreover, there is a lack of optimal biomarkers for the prediction of response to this therapy. Palmitoylation is mediated by a family of 23 S-acyltransferases, termed zinc finger Asp-His-His-Cys-type palmitoyltransferases (ZDHHC), which precisely control various cancer-related protein functions and represent promising drug targets for cancer therapy. Here, we revealed that tumor cell-intrinsic ZDHHC9 was overexpressed in pancreatic cancer tissues and associated with impaired anti-tumor immunity. In syngeneic pancreatic tumor models, the knockdown of ZDHHC9 expression suppressed tumor progression and prolonged survival time of mice by modifying the immunosuppressive ('cold') to proinflammatory ('hot') tumor microenvironment. Furthermore, ZDHHC9 deficiency sensitized anti-PD-L1 immunotherapy mainly in a CD8+ T cell dependent manner. Lastly, we employed the ZDHHC9-siRNA nanoparticle system to efficiently silence ZDHHC9 in pancreatic tumors. Collectively, our findings indicate that ZDHHC9 overexpression in pancreatic tumors is a mechanism involved in the inhibition of host anti-tumor immunity and highlight the importance of inactivating ZDHHC9 as an effective immunotherapeutic strategy and booster for anti-PD-L1 therapy against pancreatic cancer.

Multifunctional porous poly (L-lactic acid) nanofiber membranes with enhanced anti-inflammation, angiogenesis and antibacterial properties for diabetic wound healing.

With increased diabetes incidence, diabetic wound healing is one of the most common diabetes complications and is characterized by easy infection, chronic inflammation, and reduced vascularization. To address these issues, biomaterials with multifunctional antibacterial, immunomodulatory, and angiogenic properties must be developed to improve overall diabetic wound healing for patients. In our study, we prepared porous poly (L-lactic acid) (PLA) nanofiber membranes using electrospinning and solvent evaporation methods. Then, sulfated chitosan (SCS) combined with polydopamine-gentamicin (PDA-GS) was stepwise modified onto porous PLA nanofiber membrane surfaces. Controlled GS release was facilitated via dopamine self-polymerization to prevent early stage infection. PDA was also applied to PLA nanofiber membranes to suppress inflammation. In vitro cell tests results showed that PLA/SCS/PDA-GS nanofiber membranes immuomodulated macrophage toward the M2 phenotype and increased endogenous vascular endothelial growth factor secretion to induce vascularization. Moreover, SCS-contained PLA nanofiber membranes also showed good potential in enhancing macrophage trans-differentiation to fibroblasts, thereby improving wound healing processes. Furthermore, our in vitro antibacterial studies against Staphylococcus aureus indicated the effective antibacterial properties of the PLA/SCS/PDA-GS nanofiber membranes. In summary, our novel porous PLA/SCS/PDA-GS nanofiber membranes possessing enhanced antibacterial, anti-inflammatory, and angiogenic properties demonstrate promising potential in diabetic wound healing processes.

Auxin efflux carrier ZmPIN1a modulates auxin reallocation involved in nitrate-mediated root formation.

BACKGROUND: Auxin plays a crucial role in nitrate (NO3-)-mediated root architecture, and it is still unclear that if NO3- supply modulates auxin reallocation for regulating root formation in maize (Zea mays L.). This study was conducted to investigate the role of auxin efflux carrier ZmPIN1a in the root formation in response to NO3- supply. RESULTS: Low NO3- (LN) promoted primary root (PR) elongation, while repressed the development of lateral root primordia (LRP) and total root length. LN modulated auxin levels and polar transport and regulated the expression of auxin-responsive and -signaling genes in roots. Moreover, LN up-regulated the expression level of ZmPIN1a, and overexpression of ZmPIN1a enhanced IAA efflux and accumulation in PR tip, while repressed IAA accumulation in LRP initiation zone, which consequently induced LN-mediated PR elongation and LR inhibition. The inhibition rate of PR length, LRP density and number of ZmPIN1a-OE plants was higher than that of wild-type plants after auxin transport inhibitor NPA treatment under NN and LN conditions, and the degree of inhibition of root growth in ZmPIN1a-OE plants was more obvious under LN condition. CONCLUSION: These findings suggest that ZmPIN1a was involved in modulating auxin levels and transport to alter NO3--mediated root formation in maize.

Aptamer-assisted two-point immobilized agonist-bound angiotensin II type 1 receptor for a second-site modulator discovery.

Methods of immobilized proteins are challenged by the way how to capture the proteins in their intact functional states. Here we present a two-point, high-specific method for the immobilization of conformationally specific angiotensin II type 1 receptor (AT1R) on amino-functionalized polystyrene microspheres. We identified a selective DNA aptamer of AT1R by a column-based SELEX approach with micromolar affinity. Two single-stranded DNA strands were utilized to introduce the AT1R aptamer and angiotensin II 3-8 peptide to the microsphere surface, resulting in the two surface-positioned sites. The two-point immobilized AT1R exhibited enhanced ligand-binding activity and stability in comparison with that prepared by a one-positioned site. Ginsenoside Rg1 and rosmarinic acid were screened from the herbal extract and proved to bind with AT1R through the allosteric and orthosteric sites of the receptor, respectively. These provide a generally applicable approach for functional protein immobilization with enhanced conformation stability, ligand binding activity, and screening efficiency.

Immunomodulation of MiRNA-223-based nanoplatform for targeted therapy in retinopathy of prematurity.

Retinopathy of prematurity (ROP) is characterized by pathological angiogenesis and associated inflammation in the retina and is the leading cause of childhood blindness. MiRNA-223 (miR-223) drives microglial polarization toward the anti-inflammatory phenotype and offers a therapeutic approach to suppress inflammation and consequently pathological neovascularization. However, miRNA-based therapy is hindered by the low stability and non-specific cell-targeting ability of delivery systems. In the present study, we developed folic acid-chitosan (FA-CS)-modified mesoporous silica nanoparticles (PMSN) loaded with miR-223 to regulate retinal microglial polarization. The FA-CS/PMSN/miR-223 nanoparticles exhibited high stability and loading efficiency, achieved targeted delivery, and successfully escaped from lysosomes. In cultured microglial cells, treatment with FA-CS/PMSN/miR-223 nanoparticles upregulated the anti-inflammatory gene YM1/2 and IL-4RA, and downregulated the proinflammatory genes iNOS, IL-1ß, and IL-6. Notably, in a mouse oxygen-induced retinopathy model of ROP, intravitreally injected FA-CS/PMSN/miR-223 nanoparticles (1 µg) decreased the retinal neovascular area by 52.6%. This protective effect was associated with the reduced and increased levels of pro-inflammatory (M1) and anti-inflammatory (M2) cytokines, respectively. Collectively, these findings demonstrate that FA-CS/PMSN/miR-223 nanoparticles provide an effective therapeutic strategy for the treatment of ROP by modulating the miR-223-mediated microglial polarization to the M2 phenotype.

Early potential evaluation of lead compounds from a DNA-encoded library by the determination of their thermodynamics through a chromatographic method based on immobilized ß2-adrenoceptor.

Early potential evaluation of lead compounds is critical to decrease downstream lead-optimization cycle times and clinical attrition rates for drug development. This increasingly necessitates the methodologies for accurately evaluating the potential compounds. This work immobilized ß2-adrenoceptor (ß2-AR) onto microspheres through Halo-tag mediated reaction. Characterizing the resulting microspheres by elemental and functional analysis, we utilized the immobilized receptor to determine the thermodynamics of terbutaline, tulobuterol, clorprenaline, salbutamol, and methoxyphenamine. The association constants correlated to their capacity factors on the column containing the immobilized ß2-AR, thus providing a possibility for early potential evaluation of lead compounds from complex matrices like a DNA-encoded library. By this model, the lead compound (XC267) was predicted to have an association constant higher than terbutaline, salbutamol, and methoxyphenamine, but lower than tulobuterol and clorprenaline. The binding interaction between XC267 and ß2-AR is a spontaneous endothermic process with an association constant of (6.62 ± 0.13) × 104 M-1 at 37 °C. The change of Gibbs free energy(ΔGθ), enthalpy change (ΔHθ), and entropy change (ΔSθ) was -28.49 kJ/mol, -10.58 kJ/mol, and 57.79 J/moL·K at 37 °C. By the semi-empirical rule of Ross, the driving force of the interaction between XC267 and ß2-AR was electrostatic interaction. Such binding force was also achieved by molecular docking. These results suggested that XC267 is a candidate to treat asthma by specific binding to ß2-AR. We reasoned that receptor chromatography is able to the early potential evaluation of lead compounds from complex matrices.

Development of an Allostery Responsive Chromatographic Method for Screening Potential Allosteric Modulator of Beta2-adrenoceptor from a Natural Product-Derived DNA-Encoded Chemical Library.

Allosteric ligands are promising drugs owing to their remote regulations of the orthosteric ligand signaling pathway. There are few allosteric ligands due to the lack of handy and efficacious method for the screening. Herein, we developed an affinity chromatographic method for allosteric ligand screening by immobilizing purified beta2 adrenoceptor (ß2-AR) onto macroporous silica gel by a two-point tethering method. The method relies on the occupation of the orthosteric site by an antagonist and the chelation of N-terminal His-tag of the receptor and Ni2+ coated on the gel. The immobilized ß2-AR demonstrated the greatest allosteric responsive feature when Cmpd-15 (0.25 µM) was included in the mobile phase. Under the same conditions, the association constants of three agonists (salbutamol, terbutaline, and tulobuterol) reduced to 47%, 19%, and 27% compared with the data without the inclusion of Cmpd-15 in the mobile phase. APF was screened as a potential allosteric modulator of ß2-AR by applying the immobilized receptor in a natural product-derived DNA-encoded chemical library (DEL). Relying on these results, we reasoned that the current method has potential in screening allosteric ligands of the receptor. We expect that it is applicable for the discovery of new allosteric binding sites of a target protein and screening allosteric modulators of the other receptors from complex samples.

Long-term induction of endogenous BMPs growth factor from antibacterial dual network hydrogels for fast large bone defect repair.

Osteoinductive, osteoconductive, and antibacterial properties of bone repair materials play important roles in regulating the successful bone regeneration. In the present work, we developed pH-sensitive gelatin methacryloyl (GelMA)-oxidized sodium alginate (OSA) hydrogels for dual-release of gentamicin sulfate (GS) and phenamil (Phe) to enhance the antibacterial activity and to promote large bone defect repair. Controlled release of GS was achieved through physical blending with GelMA-OSA solution before photo-polymeriaztion, while Phe was encapsulated into mesoporous silicate nanoparticles (MSN) within the hydrogels. In vitro antibacterial studies against Staphylococcus aureus and Escherichia coli indicated the broad-spectrum antibacterial property. Moreover, in vitro cell tests verified the synergistically enhanced osteogenic differentiation ability. Furthermore, in vivo studies revealed that the hydrogels significantly increased new bone formation in a critical-sized mouse cranial bone defect model. In summary, the novel dual-network hydrogels with both antibacterial and osteoinductive properties showed promising potential applications in bone tissue engineering.

Hypoxia-mimicking 3D bioglass-nanoclay scaffolds promote endogenous bone regeneration.

Large bone defect repair requires biomaterials that promote angiogenesis and osteogenesis. In present work, a nanoclay (Laponite, XLS)-functionalized 3D bioglass (BG) scaffold with hypoxia mimicking property was prepared by foam replication coupled with UV photopolymerization methods. Our data revealed that the incorporation of XLS can significantly promote the mechanical property of the scaffold and the osteogenic differentiation of human adipose mesenchymal stem cells (ADSCs) compared to the properties of the neat BG scaffold. Desferoxamine, a hypoxia mimicking agent, encourages bone regeneration via activating hypoxia-inducible factor-1 alpha (HIF-1α)-mediated angiogenesis. GelMA-DFO immobilization onto BG-XLS scaffold achieved sustained DFO release and inhibited DFO degradation. Furthermore, in vitro data demonstrated increased HIF-1α and vascular endothelial growth factor (VEGF) expressions on human adipose mesenchymal stem cells (ADSCs). Moreover, BG-XLS/GelMA-DFO scaffolds also significantly promoted the osteogenic differentiation of ADSCs. Most importantly, our in vivo data indicated BG-XLS/GelMA-DFO scaffolds strongly increased bone healing in a critical-sized mouse cranial bone defect model. Therefore, we developed a novel BG-XLS/GelMA-DFO scaffold which can not only induce the expression of VEGF, but also promote osteogenic differentiation of ADSCs to promote endogenous bone regeneration.

Novel three-dimensional bioglass functionalized gelatin nanofibrous scaffolds for bone regeneration.

The clinical use of FDA-approved bone morphogenetic proteins (BMPs) are impeded by high costs, super-high dosage requirement, short half-life, and other undesirable side effects. Therefore, designing a biomaterial that can promote new bone formation without using exogenous BMPs is highly desirable in clinical applications. In the present work, a new kind of nanofibrous scaffold composed of gelatin and 45S5 bioglass (GF/45S5 BG) was prepared through thermally induced phase separation method together with the particle leach technique (TIPS&P). In addition to the significantly higher mechanical strength, the composite scaffolds (GF/45S5 BG) significantly increased osteogenic differentiation of human mesenchymal stem cells (hMSCs) in vitro compared with the neat scaffold (GF) without adding other biological agents, for example, BMPs or hormones. Most importantly, our in vivo studies also indicated that GF/45S5 BG scaffolds could directly promote ectopic bone regeneration in SD rats without exogenous BMP2. In summary, both in vitro and in vivo results indicated that the novel 45S5 bioglass functionalized GF nanofibrous scaffold is a promising alternative for bone tissue engineering.

Streptomyces hygroscopicus OsiSh-2-induced mitigation of Fe deficiency in rice plants.

The limited availability of nutrient Fe severely impairs the health of almost all organisms. Endophytic actinobacteria can benefit the host plant in different ways. We previously inferred that the rice (Oryza) endophytic Streptomyces hygroscopicus OsiSh-2 possesses a highly efficient Fe-acquisition system. In this work, we first evaluated the effects of OsiSh-2 on the Fe-deficiency resilience of the host rice. The results demonstrated that the inoculation of OsiSh-2 considerably increased the plant biomass, Fe concentration and translocation factor, and chlorophyll content, and net leaf photosynthetic rate under Fe limiting condition. The expression of genes involved with Fe3+-reduction-related strategy in rice was up-regulated, while that involved with Fe3+-chelation-related strategy was down-regulated by OsiSh-2 treatment. Meanwhile, the OsiSh-2-rice symbiont showed enhancement of Fe3+-chelate reductase activity, total siderophore production, and acidification trend in the rhizosphere under Fe deficiency compared to plants without this endophyte. In conclusion, endophytic OsiSh-2 could protect plants against Fe-deficient stress by a sophisticated interaction with the host, including modulating Fe chelation, solubilization, reduction and translocation, ultimately leading to enhanced fitness of plant.

Identification and characterization of a novel bacterial carbohydrate esterase from the bacterium Pantoea ananatis Sd-1 with potential for degradation of lignocellulose and pesticides.

OBJECTIVE: Identification and characterization of a novel bacterial carbohydrate esterase (PaCes7) with application potential for lignocellulose and pesticide degradation. RESULTS: PaCes7 was identified from the lignocellulolytic bacterium, Pantoea ananatis Sd-1 as a new carbohydrate esterase. Recombinant PaCes7 heterologously expressed in Escherichia coli showed a clear preference for esters with short-chain fatty acids and exhibited maximum activity towards α-naphthol acetate at 37 °C and pH 7.5. Purified PaCes7 exhibited its catalytic activity under mesophilic conditions and retained more than 40% activity below 30 °C. It displayed a relatively wide pH stability from pH 6-11. Furthermore, the enzyme was strongly resistant to Mg2+, Pb2+, and Co2+ and activated by K+ and Ca2+. Both P. ananatis Sd-1 and PaCes7 could degrade the pesticide carbaryl. Additionally, PaCes7 was shown to work in combination with cellulase and/or xylanase in rice straw degradation. CONCLUSIONS: The data suggest that PaCes7 possesses promising biotechnological potential.

Nanoclay-functionalized 3D nanofibrous scaffolds promote bone regeneration.

Developing a biomaterial that can promote osteoblastic differentiation, thereby reducing the needs of exogenous osteogenic factors for large bone repair, has been a significant and long-term technical hurdle. In this study, we developed an innovative nanoclay (nanosilicate, NS)-functionalized 3D gelatin nanofibrous scaffold (GF/NS) through a thermally induced phase separation method together with the particle leaching technique (TIPS&P). In addition to the significantly higher mechanical strength, the composite scaffolds (GF/NS) demonstrated a significantly stronger ability to promote the osteogenic differentiation of human mesenchymal stem cells (hMSCs) in vitro compared to the GF scaffold. Our data further revealed that this intriguing pro-osteoblastic functionality was largely because of the unique features of NS, particularly, the strong binding ability to pro-osteoblastic factors (e.g., BMP2) as well as the intrinsic osteoinductivity of its bioactive degradation products. Most importantly, our in vivo studies indicated that GF/NS scaffolds significantly improved low-dose BMP2-induced ectopic bone regeneration in mice.