Triple Helix Structure Research Articles

Structural characterization and immune-enhancing effects of a novel polysaccharide extracted from Sargassum fusiforme

To alleviate the adverse effects of chemotherapy and bolster immune function, a novel polysaccharide derived from Sargassum fusiforme named as SFP-αII. The structural composition of SFP-αII predominantly consisted of guluronic and mannuronic acids in a molar ratio of 33.8:66.2, with an average molecular weight of 16.5 kDa. Its structure was primarily characterized by →4)-α-GulA-(1 → and →4)-β-ManA-(1 → linkages confirmed by FT-IR, methylation, and NMR analyses. The absence of a triple-helix structure was in SFP-αII was confirmed using circular dichroism and Congo red dye assays. The dimensions varied with lengths ranging from 20 nm up to 3 μm revealed by atomic force microscopy (AFM). SFP-αII has been found to enhance immunomodulatory activity in cyclophosphamide (CTX)-induced immunosuppressed mice. This was evidenced by improvements in immune organ indices, cytokine levels, and the release of nitric oxide (NO). Specifically, SFP-αII mitigated immunosuppression by upregulating the secretion of IL-1β (167.3 %) and TNF-α (227.1 %) at a dose of 400 mg/kg, compared with the CTX group in macrophages. Ultimately, SFP-αII may serve as a mechanism for immune enhancement through modulation of TLR4-mediated NF-κB and MAPK signaling pathways. This integration of traditional Chinese and Western medicine, leveraging SFP-αII as a potential functional food could be pivotal in alleviating immunosuppressive side effects in CTX treatment.

Tibetan kefir grains fermentation alters physicochemical properties and improves antioxidant activities of Lycium barbarum pulp polysaccharides

There is a lack of research on how Tibetan kefir grains fermentation alters the physicochemical properties and biological activity of Lycium barbarum pulp polysaccharides, despite some reports that fermentation can affect the structure and activity of plant polysaccharides. This study demonstrated that, through fermentation, the molecular weight of polysaccharides decreased from 25.33 to 15.11 kg/mol while the contents of total sugar and uronic acid increased by 19.11% and 40.38%, respectively. Furthermore, after fermentation, the polysaccharides exhibited an uneven and rough surface along with a reduced number of branched chains and triple helix structures. Tibetan kefir grains fermentation enhanced the antioxidant activity of polysaccharides, which may be attributed to an increase in arabinose, galactose, and uronic acid content and a decrease in polysaccharide molecular weight. This research offers an alternative viewpoint on the potential application of Tibetan kefir grains-fermented Lycium barbarum pulp polysaccharides in functional foods.

Effect of core–sheath bi‐polymeric scaffolds fabricated from acid‐soluble collagen and poly(lactic acid) derivatives on wound healing

AbstractThe core–sheath bi‐polymeric scaffold has been proven as an encouraging material based on the requirement of scaffolds. This study aims to prepare electrospun core–sheath scaffolds by using acid‐soluble collagen (ASC) as core material and poly(lactic acid) (PLA) or PLA‐g‐VAc as sheath material to get the most in combination from a hydrophilic and a hydrophobic polymer. ASC is extracted from waste Tilapia fish skin conserving the triple helix structure of the α1 (130 kDa) chain, and a α2 (120 kDa) chain cross‐linked with the β (280 kDa) chain confirmed by amino acid profile, sodium dodecyl sulphate‐polyacrylamide gel electrophoresis. PLA‐g‐VAc is prepared by grafting vinyl acetate (VAc) onto the PLA chain using benzoyl peroxide as the initiator. FT‐IR, 1H NMR, and 13C NMR of PLA‐g‐VAc reveal that grafting occurs between the double bond of VAc and the methine group of PLA. The morphology of the scaffolds is determined by the field emission scanning electron microscope. FT‐IR, thermogravimetric analysis, differential scanning calorimetry, XRD, and water contact angle measurements are used for further characterization of scaffolds. In vivo, cytotoxicity analysis on the Vero cell line exposes that scaffolds are biocompatible. Application of scaffolds to the surgically produced wounding of skin in a rat model followed by histological assay indicates the enhanced properties of core–sheath scaffolds rather than the single polymeric scaffolds.

Biophysical Investigation of RNA ⋅ DNA : DNA Triple Helix and RNA : DNA Heteroduplex Formation by the lncRNAs MEG3 and Fendrr.

Long non-coding RNAs (lncRNAs) are important regulators of gene expression and can associate with DNA as RNA : DNA heteroduplexes or RNA ⋅ DNA : DNA triple helix structures. Here, we review in vitro biochemical and biophysical experiments including electromobility shift assays (EMSA), circular dichroism (CD) spectroscopy, thermal melting analysis, microscale thermophoresis (MST), single-molecule Förster resonance energy transfer (smFRET) and nuclear magnetic resonance (NMR) spectroscopy to investigate RNA ⋅ DNA : DNA triple helix and RNA : DNA heteroduplex formation. We present the investigations of the antiparallel triplex-forming lncRNA MEG3 targeting the gene TGFB2 and the parallel triplex-forming lncRNA Fendrr with its target gene Emp2. The thermodynamic properties of these oligonucleotides lead to concentration-dependent heterogeneous mixtures, where a DNA duplex, an RNA : DNA heteroduplex and an RNA ⋅ DNA : DNA triplex coexist and their relative populations are modulated in a temperature-dependent manner. The in vitro data provide a reliable readout of triplex structures, as RNA ⋅ DNA : DNA triplexes show distinct features compared to DNA duplexes and RNA : DNA heteroduplexes. Our experimental results can be used to validate computationally predicted triple helix formation between novel disease-relevant lncRNAs and their DNA target genes.

TTSBBC: triplex target site biomarkers and barcodes in cancer.

The technology of triplex-forming oligonucleotides (TFOs) provides an approach to manipulate genes at the DNA level. TFOs bind to specific sites on genomic DNA, creating a unique intermolecular triple-helix DNA structure through Hoogsteen hydrogen bonding. This targeting by TFOs is site-specific and the locations TFOs bind are referred to as TFO target sites (TTS). Triplexes have been observed to selectively influence gene expression, homologous recombination, mutations, protein binding, and DNA damage. These sites typically feature a poly-purine sequence in duplex DNA, and the characteristics of these TTS sequences greatly influence the formation of the triplex. We introduce TTSBBC, a novel analysis and visualization platform designed to explore features of TTS sequences to enable users to design and validate TTSs. The web server can be freely accessed at https://kowalski-labapps.dellmed.utexas.edu/TTSBBC/.

Structural characterization and immunostimulant activities of polysaccharides fractionated by gradient ethanol precipitation method from Panax ginseng C. A. Meyer.

Panax ginseng C. A. Meyer is a dual-purpose plant for medicine and food, its polysaccharide is considered as an immune enhancer. Four polysaccharides, WGP-20-F, WGP-40-F, WGP-60-F and WGP-80-F were obtained from ginseng via water extraction and gradient ethanol precipitation with different molecular weights (Mw) of 1.720 × 106, 1.434 × 106, 4.225 × 104 and 1.520 × 104Da, respectively. WGP-20-F and WGP-40-F which with higher Mw and a triple-helix structure are glucans composed of 4-ɑ-Glcp, do not show remarkable immunoregulatory effects. WGP-60-F and WGP-80-F are heteropolysaccharides mainly composed of 4-ɑ-Glcp and also contain t-ɑ-Araf, 5-ɑ-Araf and 3,5-ɑ-Araf. They are spherical branched conformations without a triple-helix structure and can effectively increase the index of immune organs, lymphocyte proliferation, activate macrophages to regulate the immune system in mice and further enhance immune functions by improving delayed-type hypersensitivity reaction and antibody response. These results indicated that WGP-60-F and WGP-80-F could be used as potential immune enhancers, and gradient ethanol precipitation can be applied for the preparation of ginseng bioactive polysaccharide.

Ultrasound assisted hot water extraction of polysaccharides from Taraxacum mongolicum: Optimization, purification, structure characterization, and antioxidant activity.

Ultrasound assisted hot water extraction (UAHWE) was applied to extraction of polysaccharides from Taraxacum mongolicum with hot water as extract solvent. Experimental factors in UAHWE process were optimized by response surface methodology. The optimal extraction parameters to achieve the highest Taraxacum mongolicum polysaccharides (TMPs) yield (12.08±0.14)% by UAHWE were obtained under the ultrasound power of 200W, extraction temperature of 62°C, solid-to-liquid ratio of 1:20g/mL, and extraction time of 40min, and then the crude TMPs were further purified by DEAE-52 and Sephadex G-100 chromatography to obtain a homogenous polysaccharide fraction (TMPs-1-SG). Subsequently, the structure of TMPs-1-SG was characterized by UV-vis, Fourier transform infrared spectroscopy (FT-IR), high performance gel permeation chromatography (HPGPC), high performance liquid chromatography (HPLC), scanning electron microscope (SEM), transmission electron microscopy (TEM), and Congo red test. The results display that TMPs-1-SG with an average molecular weight of 5.49 × 104Da was comprised of mannose (Man), galactose (Gal), xylose (Xyl), and arabinose (Ara) with a molar ratio of 39.85:52.61:27.14:6.30. Moreover, TMPs-1-SG did not contain a triple helix structure. Furthermore, TMPs-1-SG and TEM presented a sheet-like, rod-shaped, and irregular structure. Finally, the antioxidant activity of TMPs-1-SG was evaluated by in vitro experiment. The IC50 values of scavenging DPPH and OH radicals for TMPs-1-SG achieved 0.71mg/mL and 0.75mg/mL, respectively. The findings can provide an effective method for extracting polysaccharides from natural resources.

Exploring carp scales (Cyprinus carpio L.) as a novel source of collagen for dental use: Extraction and characterization

Previous studies have shown that using bovine and pork collagen in dentistry is associated with various limitations, such as bovine spongiform encephalopathy, transmissible spongiform encephalopathy, and foot and mouth disease. This study aims to examine the extraction and characterization of collagen from common carp (Cyprinus carpio L.) scales, as an alternative source in dentistry. The samples used in this study consisted of common carp (C. carpio L.) scales. In the first step, common carp scales were acid-extracted and spray-dried to isolate collagen. After the collagen had been isolated, it was tested for moisture content, ash, acidity, and functional groups, with Indonesian National Standard (SNI) 8076:2014 as the reference standard. The characterization parameters of the common carp scale collagen powder revealed an 11% yield, 3.66% moisture content, 0.32% ash content, and a pH of 7.38. Furthermore, fourier transform infrared spectroscopy spectra confirmed the presence of a triple helix structure. The extraction of collagen from common carp scales using the acetic acid method and spray drying technique successfully adhered to the composition standards specified by the SNI 8076:2014. Based on these findings, common carp scales could be considered a viable collagen replacement source for applications in dentistry.

Structural Basis of HIV-1 gp41 N-trimer for Designing Bifunctional Peptide-based Fusion Inhibitors.

Pathogenic viruses that cause large-scale global or regional outbreaks almost always contain class I fusion proteins. Although the viruses differ in morphology, they all require fusion protein-mediated virus-host cell membranes during the early stages of host cell invasion. The CHR region and NHR region of fusion proteins can form the 6-HB structure to drive the fusion pore formation between viruses and host cells through metastable interactions. Here, we obtained bifunctional N-peptides with inhibitory activities against two viruses, HIV-1 and MERS-CoV, based on the sequences in the HIV-1 NHR region by constructing N-trimer conformation interacting with the CHR region. This study demonstrates that N-peptides with the coiled triple helix structure obtained from the NHR region in 6-HB are able to target the CHR region and exhibit inhibitory activity against a variety of viruses. Moreover, this strategy can be used to investigate antivirals against unknown viruses for future outbreaks.

Lowering Entropic Barriers in Triplex DNA Switches Facilitating Biomedical Applications at Physiological pH.

Triplex DNA switches are attractive allosteric tools for engineering smart nanodevices, but their poor triplex-forming capacity at physiological conditions limited the practical applications. To address this challenge, we proposed a low-entropy barrier design to facilitate triplex formation by introducing a hairpin duplex linker into the triplex motif, and the resulting triplex switch was termed as CTNSds. Compared to the conventional clamp-like triplex switch, CTNSds increased the triplex-forming ratio from 30 % to 91 % at pH 7.4 and stabilized the triple-helix structure in FBS and cell lysate. CTNSds was also less sensitive to free-energy disturbances, such as lengthening linkers or mismatches in the triple-helix stem. The CTNSds design was utilized to reversibly isolate CTCs from whole blood, achieving high capture efficiencies (>86 %) at pH 7.4 and release efficiencies (>80 %) at pH 8.0. Our approach broadens the potential applications of DNA switches-based switchable nanodevices, showing great promise in biosensing and biomedicine.

Ultrasound Assisted Aqueous Two‐Phase Extraction of Polysaccharides from Ophiopogon japonicus: Process Optimization, Structure Characterization, and Antioxidant Activity

AbstractUltrasound assisted aqueous two‐phase extraction (UATPE) is used to extract polysaccharides from Ophiopogon japonicus (OJPs). Experimental factors in UATPE process are optimized by response surface methodology. The optimal extraction conditions are obtained as follows: the ethanol concentration of 34%, ammonium sulfate concentration of 23%, solid‐to‐liquid ratio of 1:17 g/mL, and ultrasound power of 480 W, and the yield of OJPs is 9.41% ± 0.12%, and then the crude OJPs are further purified by different column chromatography to obtain a homogenous fraction (OJPs‐2‐SG), and identified its structure via different methods. OJPs‐2‐SG with a molecular weight of 1.25 × 105 Da is comprised of mannose (Man), galactose (Gal), glucose (Glc), and rhamnose (Rha) with a molar ratio of 5.15: 26.39: 68.72:8.41. Moreover, OJPs‐2‐SG contained α and β glycosidic bonds, and didn't showed a triple helix structure. Furthermore, OJPs‐2‐SG described irregular structures, and cylindrical and conical block like features. The IC50 values of scavenging 1,1‐diphenyl‐2‐picrylhydrazyl (DPPH), hydroxyl (OH), and superoxide (O2‐) radicals for OJPs‐2‐SG achieved 3.72 mg/mL, 3.89 mg/mL, and 8.71 mg/mL, respectively. These results provide important references for the extraction and application of polysaccharides.

The effect of sequential extraction on the physicochemical and rheological properties of Naematelia aurantialba polysaccharides

To overcome the difficulty of separation and low rate of extraction caused by highly viscous polysaccharides from Naematelia aurantialba (NA), four N. aurantialba polysaccharides (NAPs) were sequentially extracted using water (enzyme−/ultrasound-assisted extraction), alkali (0.1 mol/L NaOH), and acid (0.1 mol/L HCl), and named E-NAP, U-NAP, Al-NAP, and Ac-NAP. The properties of four NAPs were different. The yields of NAPs were 26.05 % (Ac-NAP) > 20.33 % (Al-NAP) > 17.99 % (U-NAP) > 12.77 % (E-NAP), respectively. The monosaccharide composition of NAPs was composed primarily of mannose, xylose, glucose, glucuronic acid, and galactose. Sequential extraction improved the purity and solubility of NAPs, but decreased the particle size, thermal stability, water retention, and crystallinity. Two polysaccharides, U-NAP and Al-NAP, had a triple helix structure. All the NAPs were pseudoplastic fluids with concentration/frequency-dependent entangled structure. Al-NAP with the highest viscosity exhibited an elastic gel, while Ac-NAP with the lowest viscosity was a viscous gel. The behavior of NAPs differed from that predicted using the Cox-Merz rule, and in particular, E-NAP and U-NAP more significantly deviated from the rule. In this study, four NAPs with different properties were extracted sequentially, which provided a theoretical basis for the down-stream processing with high added-value and utilization of NA and NAP.

Chromatographic separation by RPLC-ESI-MS of all hydroxyproline isomers for the characterization of collagens from different sources

During collagen biosynthesis, proline is post-translationally converted to hydroxyproline by specific enzymes. This amino acid, unique to collagen, plays a crucial role in stabilizing the collagen triple helix structure and could serve as an important biomarker for collagen content and quality analysis. Hydroxyproline has four isomers, depending on whether proline is hydroxylated at position 4 or 3 and on whether the cis- or trans- conformation is formed. Moreover, as extensive hydrolysis of collagen is required for its amino acid analysis, epimerization may also occur, although to a lesser extent, giving a total of eight possible isomers.The aim of the present study was to develop a reversed-phase high-performance liquid chromatography-UV-mass spectrometry (RPLC-UV-MS) method for the separation and quantification of all eight hydroxyproline isomers. After the chiral derivatization of the hydroxyproline isomers with Nα-(2,4-dinitro-5-fluorophenyl)-L-valinamide (L-FDVA), to enable their UV detection, the derivatized diastereoisomers were separated by testing different C18 column technologies and morphologies and optimizing operative conditions such as the mobile phase composition (solvent, additives), elution mode, flow rate and temperature. Baseline resolution of all eight isomers was achieved on a HALO® ES-C18 reversed-phase column (150×1.5 mm, 2.7 μm, 160 Å) using isocratic elution and MS-compatible mobile phase.The optimized method was validated for the quantification of hydroxyproline isomers and then applied to different collagen hydrolysates to gain insight and a deeper understanding of hydroxyproline abundances in different species (human, chicken) and sources (native, recombinant).

Effects of Ultrasonic Power on the Structure and Rheological Properties of Skin Collagen from Albacore (Thunnus alalunga).

The effects of ultrasonic power (0, 150, 300, 450, and 600 W) on the extraction yield and the structure and rheological properties of pepsin-soluble collagen (PSC) from albacore skin were investigated. Compared with the conventional pepsin extraction method, ultrasonic treatment (UPSC) significantly increased the extraction yield of collagen from albacore skin, with a maximum increase of 8.56%. The sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis revealed that peptides of low molecular weight were produced when the ultrasonic power exceeded 300 W. Meanwhile, secondary structure, tertiary structure, and X-ray diffraction analyses showed that the original triple helix structure of collagen was intact after the ultrasonic treatment. The collagen solutions extracted under different ultrasonic powers had significant effects on the dynamic frequency sweep, but a steady shear test suggested that the collagen extracted at 150 W had the best viscosity. These results indicate that an ultrasonic power between 150 and 300 W can improve not only the extraction yield of natural collagen, but also the rheological properties of the collagen solution without compromising the triple helix structure.

Study on fermentation preparation, physicochemical properties and biological activity of carboxymethylpachymaran with different degrees of substitution.

Carboxymethylpachymaran (CMP) is created by carboxymethylating pachyman (PM), which increases its water solubility and enhances a number of biological activities. Traditional polysaccharides modified by carboxymethylation employ strong chemical techniques. Carboxymethylcellulose (CMC) has been used previously for liquid fermentation to carboxymethyl modify bacterial polysaccharides. This theory can be applied to fungal polysaccharides because Poria cocos has the ability to naturally utilize cellulose. CMC with different degrees of substitution (DS) (0.7, 0.9 and 1.2) were added to P. cocos fermentation medium, and CMPs with different DS (0.38, 0.56 and 0.78, respectively) were prepared by liquid fermentation. The physical and chemical properties and biological activities of the CMPs were determined. Their structures were confirmed by Fourier transform infrared (FTIR) spectroscopy and monosaccharide composition. With the increase of DS, the viscosity and viscosity-average molecular weight of CMPs decreased, whereas polysaccharide content and water solubility increased, although the triple helix structure was not affected. The results of bioactivity assay showed that the higher the DS of CMPs, the higher the 2,2-diphenyl-1-picrylhydrazyl radical scavenging ability, and the stronger the bacterial inhibition ability. The present study has developed a method for producing CMPs by P. cocos liquid fermentation. The results of the study confirm that enhancing the DS of CMP could effectively enhance its potential biological activity. The findings provide safe and reliable raw materials for creating CMP-related foods and encourage CMP application in the functional food industry. © 2024 Society of Chemical Industry.

Structural and mechanical behavior of type-I collagen fibrils in presence of induced electrostatic interactions through ionic liquids

Tuning the self-assembly of collagen has broad applications in the biomedical field owing to their desired biological performance as collagenous materials with tunable functionalities can further determine cellular responses. In this work, an attempt has been made to tune the self-assembly of collagen using ionic liquids, viz., imidazolium chloride (IC) and choline dihydrogen phosphate (CDHP) at its physiological pH, followed by probing assembled systems using various characterization methods. Turbidity measurements of fibrillar networks were performed to ascertain the rate of fibril formation in addition of imidazolium chloride and choline dihydrogen phosphate to collagen at physiological pH. Morphological changes were examined using Scanning Electron Microscope (SEM), binding affinities were measured by Microscale Thermophoresis (MST), in addition to, changes in the shear viscosity, mechanical strength of collagen fibrils when interacted with imidazolium and choline based ILs were carried out using rotational rheometer and Quartz Crystal Microbalance (QCM) measurements. Experimental result depicts that CDHP imparts better crosslinking as well as mechanical strength compare to IC, which is already known for destabilizing the triple helix structure is inhibiting the fibril formation. This self-assembled, ionic-liquid treated collagen-fibrillar system would accelerate various force modulated fibrillar network study, for mimicking the ECM and tissue engineering application.

Evaluation of the Relationship Between Collagen Matrix Damage and Proteoglycan Activity Following Submaximal Fatigue Loading

Background and Hypothesis:Previous research indicated that fatigue loading of the anterior cruciate ligament (ACL) leads to disruption and unraveling of the collagen triple helix structure. Using a novel in vivo murine model, we have spectroscopically shown that collagen unraveling at the molecular level is associated with an increase in proteoglycan activity. Moreover, we found an increase in tissue compliance within these damaged tissue domains. We hypothesize that the increase in proteoglycan activity is largely responsible for this mechanical change. To investigate this, we first need to identify the proteoglycan(s) responsible for our biochemical spectra that indicatedan elevation in activity associated with unraveled collagen domains. For this pilot project, we histomorphometrically investigated the proteoglycan activity of two potential protein candidates (decorin, versican) following sub-maximal ACL fatigue loading.
 Experimental Design or Project Methods:The right knees of 20 mice underwent 5,000 cycles of moderate or strenuous fatigue loading. Left knees served as a control. Samples were collected 1 and 72 hours after loading. Immunohistochemistry was performed to detect decorin and versican activity in the ACLs.
 Results:A greater area of the ACL was stained with decorin relative to versican in ACLs that underwent moderate fatiguing after 1-hour rest (p = 0.02) and 72-hour rest (p < 0.01). Similar differences were found in ACLs after strenuous fatiguing and 72-hour rest (p < 0.01). No significant temporal differences were found in versican (p = 0.76) or decorin (p = 0.53) activity in fatigued ACLs relative to controls.
 Conclusion and Potential Impact:These preliminary findings suggest that versican nor decorin are the proteoglycans colocalizing with unraveled collagen. We are currently investigating aggrecan, lumican, and biglycan as other potential proteoglycan candidates to explain our spectroscopic data.

Laboratory measurements on the suppression of vortex-induced vibrations and rotation in long-span riserless drill strings

Riserless drilling is a leading offshore technology, but the absence of a riser causes significant vibration during outflow and rotation. However, effective methods for addressing vortex-induced vibration coupling rotation are inadequate. Experiments were conducted on riserless drill strings, and the dynamic responses were analyzed by modal decomposition theory. Results indicate that structures with higher helix numbers and screw heights exit or delay entry to the lock-in region. Triple-helix designs with screw heights of 0.2 Diameter (D) and 0.25D achieve up to 93% suppression efficiency. Increasing helix number and screw height reduces rotating drill string frequencies and delays the occurrence of the dominant frequency jump phenomenon, resulting in enhanced suppression effects. Amplitude suppression primarily occurs in medium and weak regions, while helical strakes significantly reduce amplitudes in lock-in regions. For controlling vibration and minimizing fatigue damage in rotating drill strings, it is recommended to use triple-helix structures with lower screw heights.

PBL Teaching and Evaluation Model Embedded in Triple Helix Structure —Research on Reform of Educational Evaluation and Cultivation of Practical Ability Based on Higher Vocational Students’ Education

PBL Teaching and Evaluation Model Embedded in Triple Helix Structure —Research on Reform of Educational Evaluation and Cultivation of Practical Ability Based on Higher Vocational Students’ Education

Insights into the 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC)-induced gelation of fully carboxylated collagen and the fabrication of hydrogels with enhanced mechanical properties

EDC-induced gelation of succinylated collagen with a high concentration was realized, and its triple helix structure is crucial for this process. G′ of succinylated collagen hydrogel was improved to 310 Pa compared to 20 Pa of the collagen hydrogel.