ABSTRACT
Starch is an attractive feedstock in biorefinery processes, while the low natural conversion rate of most microorganisms limits its applications. Herein, starch metabolic pathway was systematically investigated using Bacillus licheniformis DW2 as the host organism. Initially, the effects of overexpressing amylolytic enzymes on starch hydrolysis were evaluated. Subsequently, the transmembrane transport system and intracellular degradation module were modified to accelerate the uptake of hydrolysates and their further conversion to glucose-6-phosphate. The DW2-derived strains exhibited robust growth in starch medium, and productivity of bacitracin and subtilisin were improved by 38.5% and 32.6%, with an 32.3% and 22.9% increase of starch conversion rate, respectively. Lastly, the employment of engineering strategies enabled another B. licheniformis WX-02 to produce poly-γ-glutamic acid from starch with a 2.1-fold increase of starch conversion rate. This study not only provided excellent B. licheniformis chassis for sustainable bioproduction from starch, but shed light on researches of substrate utilization.
Subject(s)
Bacillus licheniformis , Starch , Starch/metabolism , Bacillus licheniformis/metabolism , Hydrolysis , Polyglutamic Acid/analogs & derivatives , Polyglutamic Acid/biosynthesis , Industrial Microbiology/methodsABSTRACT
BACKGROUND: Significant efforts have been devoted to assess the effects of the poly-gamma-glutamic acid (γ-PGA) on crop growth, yield and quality, soil water retention and fertilizer use efficiency. However, few studies have evaluated the effects of γ-PGA on greenhouse gas (GHG) emissions and grain yield from paddy fields with different rice varieties. METHODS: In the present study, a split-plot field experiment was performed to comprehensively evaluate the effects of γ-PGA concentrations (i.e., no application [P0] and 25.0 kg ha-1 of γ-PGA fermentation solution [P1]) and rice varieties (i.e., conventional rice [Huanghuazhan, H], red rice [Gangteyou 8024, R] and black rice [Black indica rice, B]) on the grain yield, GHG emissions, global warming potential (GWP), greenhouse gas intensity (GHGI), net ecosystem economic profit (NEEP) and carbon footprint (CF) during 2022 and 2023 rice-growing seasons in central China. RESULTS: Application of γ-PGA significantly affected the GHGs emissions, NEEP and CF. Compared with P0 treatments, P1 treatments significantly increased the NEEP by 1.2-11.2 %, and decreased the GWP by 12.9-35.4 %, the GHGI by 16.5-35.9 % and the CF by 13.8-26.2 % in 2022-2023. Application of γ-PGA showed a tendency to increase the yield. Under γ-PGA application condition, R treatment exhibited the lowest GWP, GHGI and CF, and the highest yield and NEEP compared with B and H treatments. CONCLUSION: Our results suggest that γ-PGA application is an ecological agricultural management to increase rice yield, reduce greenhouse gas emission and increase economic benefit, and its advantage is more significant for red rice than for other rice varieties.
Subject(s)
Greenhouse Gases , Oryza , Oryza/growth & development , Greenhouse Gases/analysis , China , Polyglutamic Acid/analogs & derivatives , Agriculture/methods , Fertilizers , Edible Grain/growth & development , Global WarmingABSTRACT
Inspired by the unique functionalities of biomolecular membraneless organelles (MLOs) formed via liquid-liquid phase separation (LLPS) of intrinsically disordered proteins (IDPs) and nucleic acids, a great deal of effort has been devoted to devising phase-separated artificial subcellular dynamic compartments. These endeavors aim to unravel the molecular mechanism underlying the formation and intracellular delivery of susceptible macromolecular therapeutics. We report herein pyroglutamic acid (PGA)-based well-defined homopolymers featuring stimuli-tunable reversible self-coacervation ability. The polymer exhibits an upper critical solution temperature (UCST) transition in aqueous solutions and has the propensity to undergo cooling-induced LLPS, producing micrometer-sized liquid droplets. This phase separation phenomenon could be modulated by various factors, including polymer concentration, chain length, solution pH, and types and concentrations of different additives. These micrometer droplets are thermally reversible and encapsulate a wide variety of cargoes, including small hydrophobic fluorescent molecules, hydrophilic anticancer drugs, and fluorophore-labeled macromolecular proteins (bovine serum albumin and lysozyme). The payloads were released by exploiting the thermo/pH-mediated disassembly behavior of the coacervates, preserving the bioactivity of the sensitive therapeutics. This environmentally responsive, simple yet versatile artificial MLO model system will provide insights into the biomolecular nonionic condensates and pave the way for the de novo design of dynamic biomolecule depots.
Subject(s)
Hydrogen Bonding , Humans , Serum Albumin, Bovine/chemistry , Muramidase/chemistry , Polyglutamic Acid/chemistry , Polyglutamic Acid/analogs & derivatives , Antineoplastic Agents/chemistry , Hydrogen-Ion Concentration , Drug Liberation , Temperature , Polymers/chemistry , Hydrophobic and Hydrophilic InteractionsABSTRACT
Drought stress is the main factor restricting maize yield. Poly-γ-glutamic acid (γ-PGA), as a water-retaining agent and fertilizer synergist, could significantly improve the drought resistance and yield of many crops. However, its high production costs and unclear long-term impact on soil ecology limit its large-scale application. In this study, an environmentally friendly green material γ-PGA was heterologous synthesized in maize for the first time using the synthetic biology method. The genes (PgsA, PgsB, PgsC) participated in γ-PGA synthesis were cloned from Bacillus licheniformis and transformed into maize to produce γ-PGA for the first time. Under drought stress, transgenic maize significantly increased the ear length, ear weight and grain weight by 50 % compared to the control, whereas the yield characteristic of ear weight, grain number per ear, grain weight per ear and 100-grain weight increased by 1.67 %-2.33 %, 3.78 %-13.06 %, 8.41 %-22.06 %, 6.03 %-19.28 %, and 11.85 %-18.36 %, respectively under normal growth conditions. γ-PGA was mainly expressed in the mesophyll cells of maize leaf rosette structure and improved drought resistance and yield by protecting and increasing the expression of genes for the photosynthetic and carbon fixation. This study is an important exploration for maize drought stress molecular breeding and building resource-saving agriculture.
Subject(s)
Droughts , Plants, Genetically Modified , Polyglutamic Acid , Zea mays , Zea mays/genetics , Polyglutamic Acid/analogs & derivatives , Polyglutamic Acid/biosynthesis , Plants, Genetically Modified/genetics , Stress, Physiological/genetics , Gene Expression Regulation, Plant , Bacillus licheniformis/genetics , Bacillus licheniformis/metabolism , Plant Leaves/genetics , Drought ResistanceABSTRACT
Cryopreservation is highly desired for long-term maintenance of the viability of living biosamples, while effective cell cryopreservation still relies heavily on the addition of dimethyl sulfoxide (DMSO) and fetal bovine serum (FBS). However, the intrinsic toxicity of DMSO is still a bottleneck, which could not only cause the clinical side effect but also induce cell genetic variants. In the meantime, the addition of FBS may bring potentially the risk of pathogenic microorganism contamination. The liquid marbles (LMs), a novel biotechnology tool for cell cryopreservation, which not only have a small volume system that facilitated recovery, but the hydrophobic shell also resisted the harm to cells caused by adverse environments. Previous LM-based cell cryopreservation relied heavily on the addition of FBS. In this work, we introduced acidic polyaspartic acid and polyglutamic acid as cryoprotectants to construct LM systems. LMs could burst in an instant to facilitate and achieve ultrarapid recovery process, and the hydrophilic carboxyl groups of the cryoprotectants could form hydrogen bonds with water molecules and further inhibit ice growth/formation to protect cells from cryoinjuries. The L929 cells could be well cryopreserved by acidic polyamino acid-based LMs. This new biotechnology platform is expected to be widely used for cell cryopreservation, which has the potential to propel LMs for the preservation of various functional cells in the future.
Subject(s)
Cell Survival , Cryopreservation , Cryoprotective Agents , Cryopreservation/methods , Animals , Mice , Cell Survival/drug effects , Cryoprotective Agents/pharmacology , Cryoprotective Agents/chemistry , Cell Line , Hydrophobic and Hydrophilic Interactions , Dimethyl Sulfoxide/chemistry , Dimethyl Sulfoxide/pharmacology , Peptides/chemistry , Peptides/pharmacology , Polyglutamic Acid/chemistry , Polyglutamic Acid/analogs & derivatives , Polyglutamic Acid/pharmacologyABSTRACT
Anionic synthetic polypeptides are promising candidates as standalone bone-targeting drug carriers. Nevertheless, the structure-property relationship of the bone-targeting ability of polypeptides remains largely unexplored. Herein we report the optimization of the in vitro and in vivo bone-targeting ability of poly(glutamic acid)s (PGAs) by altering their chain lengths and backbone chirality. PGA 100-mers exhibited higher hydroxyapatite affinity in vitro, but their rapid macrophage clearance limited their targeting ability. Shorter PGA was therefore favored in terms of in vivo bone targeting. Meanwhile, the backbone chirality showed less significant impact on the in vitro and in vivo targeting behavior. This study highlights the modulation of structural parameters on the bone-targeting performance of anionic polypeptides, shedding light on the future design of polypeptide-based carriers.
Subject(s)
Bone and Bones , Polyglutamic Acid , Polyglutamic Acid/chemistry , Polyglutamic Acid/analogs & derivatives , Animals , Bone and Bones/drug effects , Bone and Bones/metabolism , Mice , Durapatite/chemistry , RAW 264.7 Cells , Drug Carriers/chemistry , Macrophages/drug effects , Macrophages/metabolismABSTRACT
Kinema, a traditional fermented soybean food from the Himalayas, is well-liked for its sticky texture and flavourful umami taste. Among 175 bacterial strains from spontaneously fermented kinema samples, Bacillus subtilis Tamang strain stood out for its high stickiness and viscosity. The strain's Poly-γ-glutamic acid (γ-PGA) contains various groups of glutamic acid and has a molecular weight of 660 kDa. It demonstrates the ability to solubilize iron, preserve ferritin in Caco-2 cells, and exhibit antibacterial properties. The genome of B. subtilis Tamang is devoid of plasmid elements but does feature nine insert elements. Noteworthy is the presence of unique secondary metabolites with potential antimicrobial effects, such as amyloliquecidin GF610, bogorol A, and thermoactinoamide A. A total of 132 carbohydrate-active enzymes (CAZy) were identified, hinting at possible prebiotic characteristics. The genome analysis revealed genes responsible for γ-PGA production via the capBCA complex. Furthermore, genes associated with fibrinolytic activity, taste enhancement, biopeptides, immunomodulators, and vitamins like B12 and K2 were found, along with probiotics and various health benefits. The genetic material for L-asparaginase production, known for its anti-cancer properties, was also detected, as well as CRISPR-Cas systems. The absence of virulence factors and antimicrobial resistance genes confirms the safety of consuming B. subtilis Tamang as a food-grade bacterium.
Subject(s)
Bacillus subtilis , Fermentation , Genome, Bacterial , Polyglutamic Acid , Whole Genome Sequencing , Bacillus subtilis/genetics , Bacillus subtilis/metabolism , Polyglutamic Acid/analogs & derivatives , Polyglutamic Acid/metabolism , Caco-2 Cells , Humans , Food Microbiology , Fermented Foods/microbiology , Soy Foods/microbiology , Anti-Bacterial Agents/pharmacologyABSTRACT
Farmland mercury (Hg) pollution poses a significant threat to human health, but there is a lack of highly efficient phytoextraction for its remediation at present. This study investigates the impact of poly-γ-glutamic acid (γ-PGA) on the phytoextraction capabilities of Pennisetum giganteum (P. giganteum) in Hg-contaminated soil. Our research indicates that amending γ-PGA to soil markedly enhances the assimilation of soil Hg by P. giganteum and transformation of Hg within itself, with observed increases in Hg concentrations in roots, stems, and leaves by 1.1, 4.3, and 18.9 times, respectively, compared to the control. This enhancement is attributed to that γ-PGA can facilitate the hydrophilic and bioavailable of soil Hg. Besides, γ-PGA can stimulate the abundance of Hg-resistance bacteria Proteobacteria in the rhizosphere of P. giganteum, thus increasing the mobility and uptake of soil Hg by P. giganteum roots. Moreover, the hydrophilic nature of Hg-γ-PGA complexes supports their transport via the apoplastic pathway, across the epidermis, and through the Casparian strip, eventually leading to immobilization in the mesophyll tissues. This study provides novel insights into the mechanisms of Hg phytoextraction, demonstrating that γ-PGA significantly enhances the effectiveness of P. giganteum in Hg uptake and translocation. The findings suggest a promising approach for the remediation of Hg-contaminated soil, offering a sustainable and efficient strategy for environmental management and health risk mitigation.
Subject(s)
Biodegradation, Environmental , Mercury , Pennisetum , Polyglutamic Acid , Soil Pollutants , Soil Pollutants/metabolism , Mercury/metabolism , Pennisetum/metabolism , Polyglutamic Acid/analogs & derivatives , Polyglutamic Acid/metabolism , Soil/chemistryABSTRACT
BACKGROUND AND PURPOSE: Clinical studies showed that prolonged infusion of methotrexate (MTX) leads to more severe adverse reactions than short infusion of MTX at the same dose. We hypothesized that it is the saturation of folate polyglutamate synthetase (FPGS) at high MTX concentration that limits the intracellular synthesis rate of methotrexate polyglutamate (MTX-PG). Due to a similar accumulation rate, a longer infusion duration may increase the concentration of MTX-PG and, result in more serious adverse reactions. In this study, we validated this hypothesis. EXPERIMENTAL APPROACH: A549, BEL-7402 and MHCC97H cell lines were treated with MTX at gradient concentrations. Liquid chromatograph-mass spectrometer (UPLC-MS/MS) was used to quantify the intracellular concentration of MTX-PG and the abundance of FPGS and γ-glutamyl hydrolase (GGH). High quality data were used to fit the cell pharmacokinetic model. KEY RESULTS: Both cell growth inhibition rate and intracellular MTX-PG concentration showed a nonlinear relationship with MTX concentration. The parameter Vmax in the model, which represents the synthesis rate of MTX-PG, showed a strong correlation with the abundance of intracellular FPGS. CONCLUSION AND IMPLICATIONS: According to the model fitting results, it was confirmed that the abundance of FPGS is a decisive factor limiting the synthesis rate of MTX-PG. The proposed hypothesis was verified in this study. In addition, based on the intracellular metabolism, a reasonable explanation was provided for the correlation between the severity of adverse reactions of MTX and infusion time. This study provides a new strategy for the individualized treatment and prediction of efficacy/side effects of MTX.
Subject(s)
Methotrexate , Peptide Synthases , Polyglutamic Acid , gamma-Glutamyl Hydrolase , Methotrexate/pharmacokinetics , Methotrexate/analogs & derivatives , gamma-Glutamyl Hydrolase/metabolism , Peptide Synthases/metabolism , Humans , Cell Line, Tumor , Polyglutamic Acid/analogs & derivatives , Tandem Mass Spectrometry , Cell Proliferation/drug effects , Antimetabolites, Antineoplastic/pharmacokinetics , Antimetabolites, Antineoplastic/pharmacologyABSTRACT
Polymer-homopolypeptide block copolymers are a class of bioinspired materials that combine the processability and stability of synthetic polymers with the biocompatibility and unique secondary structures of peptides, such as α-helices and ß-sheets. These properties make them ideal candidates for a wide variety of applications, for example, in the pharmaceutical field, where they are frequently explored as building blocks for polymeric micelle drug delivery systems. While homopolypeptide side chains can be furnished with an array of different moieties to impart the copolymers with desirable properties, such as stimulus responsivity, pyridine derivatives represent an underutilized functional group for this purpose. Additionally, the interplay between polypeptide side chain structure, secondary conformation, and micelle morphology is not yet well understood, particularly in the case of structural regioisomers. Therefore, in this work, a series of polymer-homopolypeptide copolymers were prepared from a poly(ethylene glycol)-b-poly(glutamic acid) (PEG-b-PGA) backbone, where the pendant carboxylic acid groups were covalently conjugated to a series of pyridine regioisomers by carbodiimide coupling. These pyridine regioisomers differed only in the position of the nitrogen heteroatom, ortho, meta or para, relative to the linking group, generating a series of PEG-b-poly(pyridinylmethyl glutamate) (PEG-b-PMG) copolymers. Following self-assembly of the copolymers in aqueous solutions, dynamic light scattering (DLS) revealed differences in micelle hydrodynamic diameter (Dh) (ranging from â¼60 to 120 nm), while transmission electron microscopy (TEM) and small-angle X-ray scattering (SAXS) revealed distinctive morphologies ranging from ellipsoidal, to cylindrical, and disc-like, suggesting that subtle changes in positional isomers in the polypeptide block may influence the micelle structure. Analysis of the PEG-b-PMG copolymer micelles by circular dichroism (CD) and attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy revealed that differences in the morphology were associated with changes in polypeptide secondary structure, which in turn was influenced by the position of the pyridine heteroatom. Overall, these findings contribute to the broader understanding of the relationship between polypeptide structure and micelle morphology and serve as useful insight for the rational design of polymer-polypeptide nanoparticles.
Subject(s)
Micelles , Pyridines , Pyridines/chemistry , Polyethylene Glycols/chemistry , Peptides/chemistry , Protein Structure, Secondary , Stereoisomerism , Isomerism , Polyglutamic Acid/chemistry , Polyglutamic Acid/analogs & derivatives , Polymers/chemistryABSTRACT
Most hydrogels have poor mechanical properties, severely limiting their potential applications, and numerous approaches have been introduced to fabricate more robust and durable examples. However, these systems consist of nonbiodegradable polymers which limit their application in tissue engineering. Herein, we focus on the fabrication and investigate the influence of hydrophobic segments on ionic cross-linking properties for the construction of a tough, biodegradable hydrogel. A biodegradable, poly(γ-glutamic acid) polymer conjugated with a hydrophobic amino acid, l-phenylalanine ethyl ester (Phe), together with an ionic cross-linking group, alendronic acid (Aln) resulting in γ-PGA-Aln-Phe, was initially synthesized. Rheological assessments through time sweep oscillation testing revealed that the presence of hydrophobic domains accelerated gelation. Comparing gels with and without hydrophobic domains, the compressive strength of γ-PGA-Aln-Phe was found to be six times higher and exhibited longer stability properties in ethylenediaminetetraacetic acid solution, lasting for up to a month. Significantly, the contribution of the hydrophobic domains to the mechanical strength and stability of ionic cross-linking properties of the gel was found to be the dominant factor for the fabrication of a tough hydrogel. As a result, this study provides a new strategy for mechanical enhancement and preserves ionic cross-linked sites by the addition of hydrophobic domains. The development of tough, biodegradable hydrogels reported herein will open up new possibilities for applications in the field of biomaterials.
Subject(s)
Hydrogels , Hydrophobic and Hydrophilic Interactions , Hydrogels/chemistry , Hydrogels/chemical synthesis , Cross-Linking Reagents/chemistry , Polyglutamic Acid/chemistry , Polyglutamic Acid/analogs & derivatives , Rheology , Compressive Strength , Ions/chemistry , Biocompatible Materials/chemistry , Phenylalanine/chemistry , Phenylalanine/analogs & derivativesABSTRACT
With continuous advances in medical technology, non-invasive embolization has emerged as a minimally invasive treatment, offering new possibilities in cancer therapy. Fluorescent labeling can achieve visualization of therapeutic agents in vivo, providing technical support for precise treatment. This paper introduces a novel in situ non-invasive embolization composite material, Au NPs@(mPEG-PLGTs), created through the electrostatic combination of L-cysteine-modified gold nanoparticles (Au NPs) and methoxy polyethylene glycol amine-poly[(L-glutamic acid)-(L-tyrosine)] (mPEG-PLGTs). Experiments were undertaken to confirm the biocompatibility, degradability, stability and performance of this tumor therapy. The research results demonstrated a reduction in tumor size as early as the fifth day after the initial injection, with a significant 90% shrinkage in tumor volume observed after a 20-day treatment cycle, successfully inhibiting tumor growth and exhibiting excellent anti-tumor effects. Utilizing near-infrared in vivo imaging, Au NPs@(mPEG-PLGTs) displayed effective fluorescence tracking within the bodies of nude BALB-c mice. This study provides a novel direction for the further development and innovation of in situ non-invasive embolization in the field, highlighting its potential for rapid, significant therapeutic effects with minimal invasiveness and enhanced safety.
Subject(s)
Gold , Metal Nanoparticles , Mice, Inbred BALB C , Mice, Nude , Polyethylene Glycols , Gold/chemistry , Animals , Metal Nanoparticles/chemistry , Metal Nanoparticles/therapeutic use , Mice , Polyethylene Glycols/chemistry , Cell Line, Tumor , Humans , Hydrogen-Ion Concentration , Embolization, Therapeutic , Polyglutamic Acid/chemistry , Polyglutamic Acid/analogs & derivativesABSTRACT
Injectable adhesive hydrogels combining rapid gelling with robust adhesion to wet tissues are highly required for fast hemostasis in surgical and major trauma scenarios. Inspired by the cross-linking mechanism of mussel adhesion proteins, we developed a bionic double-crosslinked (BDC) hydrogel of poly (γ-glutamic acid) (PGA)/poly (N-(2-hydroxyethyl) acrylamide) (PHEA) fabricated through a combination of photo-initiated radical polymerization and hydrogen bonding cross-linking. The BDC hydrogel exhibited an ultrafast gelling process within 1 s. Its maximum adhesion strength to wet porcine skin reached 254.5 kPa (9 times higher than that of cyanoacrylate (CA) glue) and could withstand an ultrahigh burst pressure of 626.4 mmHg (24 times higher than that of CA glue). Notably, the BDC hydrogel could stop bleeding within 10 s from a rat liver incision 10 mm long and 5 mm deep. The wound treated with the BDC hydrogel healed faster than the control groups, underlining the potential for emergency rescue and wound care scenarios.
Subject(s)
Hydrogels , Polyglutamic Acid , Polyglutamic Acid/chemistry , Polyglutamic Acid/analogs & derivatives , Animals , Hydrogels/chemistry , Rats , Pressure , Cross-Linking Reagents/chemistry , Wound Healing/drug effects , Swine , Acrylamides/chemistryABSTRACT
The clinical application of 7-ethyl hydroxy-camptothecin (SN-38) maintains challenges not only due to its poor solubility and stability but also the lack of effective carriers to actively deliver SN-38 to deep tumor sites. Although SN-38-based nanomedicines could improve the solubility and stability from different aspects, the tumor targeting efficiency remains very low. Leveraging the hypoxic taxis of bifidobacteria bifidum (B. bifi) to the deep tumor area, we report SN-38-based nanomedicines-engineered bifidobacterial complexes for effective tumor-targeted delivery. Firstly, SN-38 was covalently coupled with poly-L-glutamic acid (L-PGA) and obtained soluble polymeric prodrug L-PGA-SN38 to improve its solubility and stability. To prolong the drug release, L-PGA-SN38 was mildly complexed with chitosan to form nanomedicines, and nanomedicines engineered B. bifi were further elaborated via electrostatic interaction of the excess of cationic chitosan shell from nanomedicines and anionic teichoic acid from B. bifi. The engineered B. bifi complexes inherited the bioactivity of native B. bifi and exhibited distinctly enhanced accumulation at the tumor site. More importantly, significantly elevated anti-tumor efficacy was achieved after the treatment of CS-L-PGA-SN38 NPs/B. bifi complexes, with favorable tumor suppression up to 80%. Such a B. bifi-mediated delivery system offers a promising platform for effective drug delivery and enhanced drug accumulation in the hypoxia deep tumor with superior anti-tumor efficacy.
Subject(s)
Chitosan , Colorectal Neoplasms , Irinotecan , Nanomedicine , Polyglutamic Acid , Irinotecan/administration & dosage , Irinotecan/pharmacology , Chitosan/chemistry , Colorectal Neoplasms/drug therapy , Animals , Polyglutamic Acid/chemistry , Polyglutamic Acid/analogs & derivatives , Humans , Nanomedicine/methods , Drug Liberation , Drug Carriers/chemistry , Drug Delivery Systems , Antineoplastic Agents, Phytogenic/administration & dosage , Antineoplastic Agents, Phytogenic/pharmacology , Mice , Camptothecin/administration & dosage , Camptothecin/analogs & derivatives , Camptothecin/chemistry , Camptothecin/pharmacology , Mice, Inbred BALB C , Cell Line, Tumor , Bifidobacterium bifidum , Mice, Nude , FemaleABSTRACT
Establishing a physical barrier between the peritoneum and the cecum is an effective method to reduce the risk of postoperative abdominal adhesions. Meloxicam (MX), a nonsteroidal anti-inflammatory drug has also been applied to prevent postoperative adhesions. However, its poor water solubility has led to low bioavailability. Herein, we developed an injectable hydrogel as a barrier and drug carrier for simultaneous postoperative adhesion prevention and treatment. A third-generation polyamide-amine dendrimer (G3) was exploited to dynamically combine with MX to increase the solubility and the bioavailability. The formed G3@MX was further used to crosslink with poly-γ-glutamic acid (γ-PGA) to prepare a hydrogel (GP@MX hydrogel) through the amide bonding. In vitro and in vivo experiments evidenced that the hydrogel had good biosafety and biodegradability. More importantly, the prepared hydrogel could control the release of MX, and the released MX is able to inhibit inflammatory responses and balance the fibrinolytic system in the injury tissues in vivo. The tunable rheological and mechanical properties (compressive moduli: from ⼠57.31 kPa to ⼠98.68 kPa;) and high anti-oxidant capacity (total free radical scavenging rate of ⼠94.56 %), in conjunction with their syringeability and biocompatibility, indicate possible opportunities for the development of advanced hydrogels for postoperative tissue adhesions management.
Subject(s)
Dendrimers , Hydrogels , Meloxicam , Nylons , Polyglutamic Acid , Hydrogels/chemistry , Hydrogels/pharmacology , Animals , Polyglutamic Acid/chemistry , Polyglutamic Acid/pharmacology , Polyglutamic Acid/analogs & derivatives , Nylons/chemistry , Tissue Adhesions/prevention & control , Dendrimers/chemistry , Dendrimers/pharmacology , Meloxicam/chemistry , Meloxicam/pharmacology , Meloxicam/administration & dosage , Mice , Inflammation/prevention & control , Inflammation/drug therapy , Anti-Inflammatory Agents, Non-Steroidal/pharmacology , Anti-Inflammatory Agents, Non-Steroidal/chemistry , Anti-Inflammatory Agents, Non-Steroidal/administration & dosage , Rats , Rats, Sprague-Dawley , Fibrinolysis/drug effects , Postoperative Complications/prevention & control , Particle Size , Injections , Drug Carriers/chemistryABSTRACT
In this study, we determined effects of an anionic siRNA delivery vector, siRNA ternary complex, which is constructed with biodegradable dendrigraft poly-L-lysine (DGL) and γ-polyglutamic acid (γ-PGA) on the melanoma cells and melanoma lung metastasis. The siRNA ternary complex showed high cellular uptake and silencing effect in melanoma cell line B16-F10/Luc cells. After intravenous administration of the siRNA ternary complex, high silencing effect was also observed in the lung of B16-F10/Luc melanoma metastasis model mice. Therefore, we applied vascular endothelial growth factor (VEGF)-siRNA on the siRNA ternary complex and determined the effect on the melanoma lung metastasis. The siRNA ternary complex containing VEGF-siRNA reduced VEGF protein levels significantly in in vitro and in vivo, and the complex successfully inhibited melanoma lung metastasis. This biodegradable and effective siRNA delivery vector, siRNA ternary complex, could be available for clinical trials.
Subject(s)
Lung Neoplasms , Melanoma, Experimental , Polyglutamic Acid , Polylysine , RNA, Small Interfering , Vascular Endothelial Growth Factor A , RNA, Small Interfering/administration & dosage , RNA, Small Interfering/pharmacology , Animals , Lung Neoplasms/secondary , Lung Neoplasms/pathology , Mice , Melanoma, Experimental/pathology , Polyglutamic Acid/chemistry , Polyglutamic Acid/analogs & derivatives , Cell Line, Tumor , Vascular Endothelial Growth Factor A/genetics , Polylysine/chemistry , Mice, Inbred C57BL , Gene Silencing , Melanoma/pathology , Melanoma/geneticsABSTRACT
Poly-γ-glutamic acid (γ-PGA) is a microbial-derived polymer with molecular weight (Mw) from 104 to 107 Da, and the high-Mw (> 7.0 × 105 Da) or ultra-high-Mw (> 5.0 × 106 Da) γ-PGA has important application value as a tissue engineering material, as a flocculant, and as a heavy metal remover. Therefore, how to produce these high-Mw γ-PGAs with low cost and high efficiency has attracted wide attention. In this study, a γ-PGA producer was isolated from the natural environment, and identified and named Bacillus subtilis GXD-20. Then, the ultra-high-Mw (> 6.0 × 106 Da) γ-PGA produced by GXD-20 was characterized. Interestingly, GXD-20 could produce γ-PGA at 42°C, and exhibited a γ-PGA titer of up to 22.29 ± 0.59 g L-1 in a 5-L fermenter after optimization of the fermentation process. Comparative genomic analysis indicated that the specific protein sequence and subcellular localization of PgdS (a γ-PGA-degrading enzyme) were closely related to the ultra-high-Mw of γ-PGA. Transcriptomic analysis revealed that the high γ-PGA titer at 42°C was mainly related to the high expression of genes encoding enzymes for sucrose transportation and utilization, nitrogen transportation, endogenous glutamate synthesis, and γ-PGA synthesis. These results provide new insights into the production of ultra-high-Mw γ-PGA by Bacillus at high temperatures.
Subject(s)
Bacillus subtilis , Glutamic Acid , Polyglutamic Acid/analogs & derivatives , Bacillus subtilis/genetics , Bacillus subtilis/metabolism , Glutamic Acid/metabolism , Molecular Weight , Polyglutamic Acid/genetics , Polyglutamic Acid/metabolism , Genomics , FermentationABSTRACT
Bacterial cellulose (BC) is an ideal candidate material for drug delivery, but the disbalance between the swelling behavior and mechanical properties limits its application. In this work, covalent crosslinking of γ-polyglutamic acid (γ-PGA) with the chitosan oligosaccharide (COS) embedded in BC was designed to remove the limitation. As a result, the dosage, time, and batch of COS addition significantly affected the mechanical properties and the yield of bacterial cellulose complex film (BCCF). The addition of 2.25 % COS at the incubation time of 0.5, 1.5, and 2 d increased the Young's modulus and the yield by 5.65 and 1.42 times, respectively, but decreased the swelling behavior to 1774 %, 46 % of that of native BC. Covalent γ-PGA transformed the dendritic structure of BCCF into a spider network, decreasing the porosity and increasing the swelling behavior by 3.46 times. The strategy balanced the swelling behavior and mechanical properties through tunning hydrogen bond, electrostatic interaction, and amido bond. The modified BCCF exhibited a desired behavior of benzalkonium chlorides transport, competent for drug delivery. Thereby, the strategy will be a competent candidate to modify BC for such potential applications as wound dressing, artificial skin, scar-inhibiting patch, and so on.
Subject(s)
Cellulose , Chitosan , Oligosaccharides , Polyglutamic Acid , Polyglutamic Acid/analogs & derivatives , Chitosan/chemistry , Cellulose/chemistry , Oligosaccharides/chemistry , Polyglutamic Acid/chemistry , Mechanical Phenomena , Bacteria/drug effects , Elastic ModulusABSTRACT
Curcumin was widely designed as nanoparticles to remove application restrictions. The occurrence of flocculation is a primary factor limiting the application of the curcumin nano-delivery system. To enhance the environmental stress resistance and functional properties of shellac-curcumin nanoparticles (S-Cur-NPs), γ-polyglutamic acid (γ-PGA) was utilized as an anti-flocculant. The encapsulation efficiency and loading capacity of S-Cur-NPs were also improved with γ-PGA incorporation. FTIR and XRD analysis confirmed the presence of amorphous characteristics in S-Cur-NPs and the combination of γ-PGA and shellac was driven by hydrogen bonding. The hydrophilic, thermodynamic, and surface potential of S-Cur-NPs was improved by the incorporation of γ-PGA. This contribution of γ-PGA on S-Cur-NPs effectively mitigated the flocculation occurrence during heating, storage, and in-vitro digestive treatment. Furthermore, it was revealed that γ-PGA enhanced the antibacterial and antioxidant properties of S-Cur-NPs and effectively protected the functional activity against heating, storage, and in-vitro digestion. Release studies conducted in simulated gastrointestinal fluids revealed that S-Cur-NPs have targeted intestinal release properties. Overall, the design of shellac with γ-PGA was a promising strategy to relieve the application stress of shellac and curcumin in the food industry.
Subject(s)
Antioxidants , Curcumin , Flocculation , Nanoparticles , Polyglutamic Acid , Curcumin/chemistry , Curcumin/pharmacology , Polyglutamic Acid/chemistry , Polyglutamic Acid/analogs & derivatives , Polyglutamic Acid/pharmacology , Nanoparticles/chemistry , Antioxidants/chemistry , Antioxidants/pharmacology , Drug Carriers/chemistry , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/chemistry , Drug Delivery Systems , Drug Liberation , Hydrophobic and Hydrophilic InteractionsABSTRACT
Redox nanoparticles have been extensively developed for chemotherapy. However, the intracellular oxidative stress induced by constant aberrant glutathione (GSH), reactive oxygen species (ROS) and gamma-glutamyl transpeptidase (GGT) homeostasis remains the primary cause of evading tumor apoptosis. Herein, an oxidative stress-amplification strategy was designed using a pH-GSH-H2O2-GGT sensitive nano-prodrug for precise synergistic chemotherapy. The disulfide bond- conjugated doxorubicin prodrug (DOX-ss) was constructed as a GSH-scavenger. Then, phenylboronic acid (PBA), DOX-ss and poly (γ-glutamic acid) (γ-PGA) were successively conjugated using chitosan oligosaccharide (COS) to obtain the nano-prodrug PBA-COS-ss-DOX/γ-PGA. The PBA-COS-ss-DOX/γ-PGA prodrug could tightly attach to the polymer chain segment by atom transfer radical polymerization. Simultaneously, the drug interacted relatively weakly with the polymer by encapsulating ionic crosslinkers in DOX@PBA-COS/γ-PGA. The disulfide bond of the DOX-ss prodrug as a GSH-scavenger could be activated using overexpressed GSH to release DOX. Particularly, PBA-COS-ss-DOX/γ-PGA could prevent premature drug leakage and facilitate DOX delivery by GGT-targeting and intracellular H2O2-cleavable linker in human hepatocellular carcinoma (HepG2) cells. Concurrently, the nano-prodrug induced strong oxidative stress and tumor cell apoptosis. Collectively, the pH-GSH-H2O2-GGT responsive nano-prodrug shows potential for synergistic tumor therapy.