bims-supasi 2024-06-02 papers

bims-supasi

Biomed News

on Sulfation pathways and signalling

Issue of 2024‒06‒02
nine papers selected by
Jonathan Wolf Mueller, University of Birmingham

Chondroitin sulfate from heads of corb: Recovery, structural analysis and assessment of anticoagulant activity.
Proline Isomerization and Molten Globular Property of TgPDCD5 Secreted from Toxoplasma gondii Confers Its Regulation of Heparin Sulfate Binding.
Development of an Analytical Method for Unsaturated Disaccharides from Heparan Sulfate Using Dansylhydrazine Derivatization and High-Performance Liquid Chromatography with Fluorescence Detection.
Characterization and Structural Analysis of a Novel Carbohydrate-Binding Module from Family 96 with Chondroitin Sulfate-Specific Binding Capacity.
Extraction, Physicochemical Properties, and In Vitro Antioxidant Activities of Chondroitin Sulfate from Bovine Nose Cartilage.
Neuritogenic glycosaminoglycan hydrogels promote functional recovery after severe traumatic brain injury.
Pleiotropic Effects of Heparin and its Monitoring in the Clinical Practice.
Chondrocyte autophagy mediated by T-2 toxin via AKT/TSC/Rheb/mTOR signaling pathway and protective effect of CSA-SeNP.
Structural investigation of Fun168A unraveling the recognition mechanism of endo-1,3-fucanase towards sulfated fucan.

Carbohydr Res. 2024 May 23. pii: S0008-6215(24)00142-3. [Epub ahead of print]541 109163

Chondroitin sulfate from heads of corb: Recovery, structural analysis and assessment of anticoagulant activity.

Hajer Bougatef, Nicola Volpi, Ikram Ben Amor, Federica Capitani, Francesca Maccari, Jalel Gargouri, Assaad Sila, Ali Bougatef.

  In this study, glycosaminoglycans (GAGs) were extracted from corb (Sciaena umbra) heads and thoroughly examined for their structure. Through cellulose acetate electrophoresis, the GAGs were identified as chondroitin sulfate (CS), with a recovery yield of 10.35 %. The CS exhibited notable characteristics including a high sulfate content (12.4 %) and an average molecular weight of 38.32 kDa. Further analysis via 1H NMR spectroscopy and SAX-HPLC revealed that the CS primarily consisted of alternating units predominantly composed of monosulfated disaccharides at positions 6 and 4 of GalNAc (52.6 % and 38.8 %, respectively). The ratio of sulfate groups between positions 4 and 6 of GalNAc (4/6 ratio) was approximately 0.74, resulting in an overall charge density of 0.98. Thermal properties of the CS were assessed using techniques such as differential scanning calorimetry and thermogravimetric analysis. Notably, the CS demonstrated concentration-dependent prolongation of activated partial thromboplastin time (aPTT) and thrombin time (TT) while showing no effect on platelet function. At 200 μg/mL, aPTT and TT coagulation times were 1.4 and 3.7 times faster than the control, respectively. These findings suggest that CS derived from corb heads holds promise as an anticoagulant agent for therapy, although further clinical investigations are necessary to validate its efficacy.

Keywords:  Anticoagulant effect; Antiplatelet activity; Chondroitin sulfate; Constitutive disaccharides; Corb heads; Structural characterization

DOI:  https://doi.org/10.1016/j.carres.2024.109163
JACS Au. 2024 May 27. 4(5): 1763-1774

Proline Isomerization and Molten Globular Property of TgPDCD5 Secreted from Toxoplasma gondii Confers Its Regulation of Heparin Sulfate Binding.

Gloria Meng-Hsuan Lin, Tsun-Ai Yu, Chi-Fon Chang, Chun-Hua Hsu.

Toxoplasmosis, caused by Toxoplasma gondii, poses risks to vulnerable populations. TgPDCD5, a secreted protein of T. gondii, induces apoptosis through heparan sulfate-mediated endocytosis. The entry mechanism of TgPDCD5 has remained elusive. Here, we present the solution structure of TgPDCD5 as a helical bundle with an extended N-terminal helix, exhibiting molten globule characteristics. NMR perturbation studies reveal heparin/heparan sulfate binding involving the heparan sulfate/heparin proteoglycans-binding motif and the core region, influenced by proline isomerization of P107 residue. The heterogeneous proline recruits a cyclophilin TgCyp18, accelerating interconversion between conformers and regulating heparan/heparin binding. These atomic-level insights elucidate the binary switch's functionality, expose novel heparan sulfate-binding surfaces, and illuminate the unconventional cellular entry of pathogenic TgPDCD5.

DOI: https://doi.org/10.1021/jacsau.3c00577
Biol Pharm Bull. 2024 ;47(5): 1054-1057

Development of an Analytical Method for Unsaturated Disaccharides from Heparan Sulfate Using Dansylhydrazine Derivatization and High-Performance Liquid Chromatography with Fluorescence Detection.

Takamasa Ishii, Ayaka Aijima, Nae Yokota, Kengo Hirai, Kyohei Higashi, Yusuke Iwasaki, Rie Ito, Nobuaki Higashi, Hiroshi Akiyama.

  Glycosaminoglycans (GAGs), such as heparan sulfate (HS), play essential roles in living organisms. Understanding the functionality of HS and its involvement in disease progression necessitates the sensitive and quantitative detection of HS-derived unsaturated disaccharides. Conventionally, fluorescence derivatization precedes the HPLC analysis of these disaccharides. However, the presence of excess unreacted derivatization reagents can inhibit rapid and sensitive analysis in chromatographic determinations. In this study, we describe analytical methods that use dansylhydrazine as a derivatization agent for the detection and determination of HS-derived unsaturated disaccharides using HPLC. In addition, we have developed a straightforward method for removing excess unreacted reagent using a MonoSpin NH2 column. This method may be employed to remove excess pre-labeling reagents, thereby facilitating the analysis of HS-derived unsaturated disaccharides with satisfactory reproducibility.

Keywords:  HPLC; MonoSpin NH2 column; dansylhydrazine; glycosaminoglycan; heparan sulfate

DOI:  https://doi.org/10.1248/bpb.b24-00194
J Agric Food Chem. 2024 May 28.

Characterization and Structural Analysis of a Novel Carbohydrate-Binding Module from Family 96 with Chondroitin Sulfate-Specific Binding Capacity.

Guanchen Liu, Xuanwei Mei, Yuying Zhang, Guangning Chen, Jiajing Li, Wenwen Tao, Menghui Sun, Long Zheng, Yaoguang Chang, Changhu Xue.

  Chondroitin sulfate (CS) is the predominant glycosaminoglycan within the human body and is widely applied in various industries. Carbohydrate-binding modules (CBMs) possessing the capacity for carbohydrate recognition are verified to be important tools for polysaccharide investigation. Only one CS-specific CBM, PhCBM100, has hitherto been characterized. In the present study, two CBM96 domains present in the same putative PL8_3 chondroitin AC lyase were discovered and recombinantly expressed. The results of microtiter plate assays and affinity gel electrophoresis assays showed that the two corresponding proteins, DmCBM96-1 and DmCBM96-2, bind specifically to CSs. The crystal structure of DmCBM96-1 was determined at a 2.20 Å resolution. It adopts a β-sandwich fold comprising two antiparallel β-sheets, showing structural similarities to TM6-N4, which is the founding member of the CBM96 family. Site mutagenesis analysis revealed that the residues of Arg27, Lys45, Tyr51, Arg53, and Arg157 are critical for CS binding. The characterization of the two CBM96 proteins demonstrates the diverse ligand specificity of the CBM96 family and provides promising tools for CS investigation.

Keywords:  CBM96; carbohydrate-binding module; chondroitin sulfate; structure

DOI:  https://doi.org/10.1021/acs.jafc.4c00090
Int J Food Sci. 2024 ;2024 6328378

Extraction, Physicochemical Properties, and In Vitro Antioxidant Activities of Chondroitin Sulfate from Bovine Nose Cartilage.

Chan Shi, Yuxuan Deng, Xin An, Yuan Chen, Xingang Lv, Qian Liu.

Beef is an important high-nutrition livestock product, and several byproducts, such as bovine cartilage, are produced during slaughter. To effectively utilize these agricultural and pastoral byproducts, combined (trypsin-papain) enzymolysis and cetylpyridine chloride purification methods were used to obtain chondroitin sulfate (CS) from the nasal cartilage of Shaanxi Yellow cattle. The effects of pH, temperature, and time on the CS yield during enzymatic hydrolysis were investigated, and the CS extraction process was optimized using response surface methodology. The best yield of CS was 21.62% under the optimum conditions of pH 6.51, temperature of 64.53°C, and enzymolysis time of 19.86 h. The molecular weight of CS from Shaanxi cattle nasal cartilage was 89.21 kDa, glucuronic acid content was 31.76 ± 0.72%, protein content was 1.12 ± 0.03%, and sulfate group content was 23.34 ± 0.08%. The nasal cartilage CS of the Yellow cattle showed strong DPPH•, •OH, and ABTS+• radical scavenging abilities and ferrous reduction ability in the experimental concentration range. This study could contribute to "turn waste into treasure" and improve the comprehensive utilization of regional characteristic biological resources.

DOI: https://doi.org/10.1155/2024/6328378
J Neural Eng. 2024 May 28.

Neuritogenic glycosaminoglycan hydrogels promote functional recovery after severe traumatic brain injury.

Nathan Gonsalves, Min Kyong Sun, Pradeep Chopra, Charles-Francois Latchoumane, Simar Bajwa, Ruiping Tang, Bianca Patel, Gert-Jan Boons, Lohitash Karumbaiah.

  OBJECTIVE: Severe traumatic brain injury (sTBI) induced neuronal loss and brain atrophy contribute significantly to long-term disabilities. Brain extracellular matrix (ECM) associated chondroitin sulfate (CS) glycosaminoglycans promote neural stem cell (NSC) maintenance, and CS hydrogel implants have demonstrated the ability to enhance neuroprotection, in preclinical sTBI studies. However, the ability of neuritogenic chimeric peptide (CP) functionalized CS hydrogels in promoting functional recovery, after controlled cortical impact (CCI) and suction ablation (SA) induced sTBI, has not been previously demonstrated. We hypothesized that neuritogenic (CS)CP hydrogels will promote neuritogenesis of human NSCs, and accelerate brain tissue repair and functional recovery in sTBI rats.APPROACH: We synthesized chondroitin 4-O sulfate (CS-A)CP, and 4,6-O-sulfate (CS-E)CP hydrogels, using strain promoted azide-alkyne cycloaddition (SPAAC), to promote cell adhesion and neuritogenesis of human NSCs, in vitro; and assessed the ability of (CS-A)CP hydrogels in promoting tissue and functional repair, in a novel CCI-SA sTBI model, in vivo.
MAIN RESULTS: Results indicated that (CS-E)CP hydrogels significantly enhanced human NSC aggregation and migration via focal adhesion kinase complexes, when compared to NSCs in (CS-A)CP hydrogels, in vitro. In contrast, NSCs encapsulated in (CS-A)CP hydrogels differentiated into neurons bearing longer neurites and showed greater spontaneous activity, when compared to those in (CS-E)CP hydrogels. The intracavitary implantation of (CS-A)CP hydrogels, acutely after CCI-SA-sTBI, prevented neuronal and axonal loss, as determined by immunohistochemical analyses. (CS-A)CP hydrogel implanted animals also demonstrated the significantly accelerated recovery of 'reach-to-grasp' function when compared to sTBI controls, over a period of 5-weeks.
SIGNIFICANCE: These findings demonstrate the neuritogenic and neuroprotective attributes of (CS)CP "click" hydrogels, and open new avenues for using modified CS biorthogonal handles to develop tissue engineered implants for sTBI repair.

Keywords:  SPAAC click chemistry; chimeric peptide; chondroitin sulfate glycosaminoglycans; reach-to-grasp function; traumatic brain injury

DOI:  https://doi.org/10.1088/1741-2552/ad5108
Semin Thromb Hemost. 2024 May 29.

Pleiotropic Effects of Heparin and its Monitoring in the Clinical Practice.

Deepa J Arachchillage, Steve Kitchen.

Unfractionated heparin (UFH) was uncovered in 1916, has been used as an anticoagulant since 1935, and has been listed in the World Health Organization's Model List of Essential Medicines. Despite the availability of many other anticoagulants, the use of heparin (either low molecular weight heparin [LMWH] or UFH) is still substantial. Heparin has pleotropic effects including anticoagulant and several nonanticoagulant properties such as antiproliferative, anti-inflammatory activity, and anticomplement effects. Although UFH has been widely replaced by LMWH, UFH is still the preferred anticoagulant of choice for patients undergoing cardiopulmonary bypass surgery, extracorporeal membrane oxygenation, and patients with high-risk mechanical cardiac valves requiring temporary bridging with a parenteral anticoagulant. UFH is a highly negatively charged molecule and binds many positively charged molecules, hence has unpredictable pharmacokinetics, and variable anticoagulant effect on an individual patient basis. Therefore, anticoagulant effects of UFH may not be proportional to the dose of UFH given to any individual patient. In this review, we discuss the anticoagulant and nonanticoagulant activities of UFH, differences between UFH and LMWH, when to use UFH, different methods of monitoring the anticoagulant effects of UFH (including activated partial thromboplastin time, heparin anti-Xa activity level, and activated clotting time), while discussing pros and cons related to each method and comparison of clinical outcomes in patients treated with UFH monitored with different methods based on available evidence.

DOI: https://doi.org/10.1055/s-0044-1786990
Osteoarthritis Cartilage. 2024 May 28. pii: S1063-4584(24)01206-8. [Epub ahead of print]

Chondrocyte autophagy mediated by T-2 toxin via AKT/TSC/Rheb/mTOR signaling pathway and protective effect of CSA-SeNP.

Xue Lin, Haobiao Liu, Lichun Qiao, Huan Deng, Miaoye Bao, Zhihao Yang, Yujie He, Rongqi Xiang, Huifang He, Jing Han.

  OBJECTIVE: Kashin-Beck disease (KBD) is an endemic, degenerative, and cartilage-damaging disease for which low selenium and T-2 toxin are considered environmental pathogenic factors. This study aimed to investigate the molecular mechanisms of autophagy in cartilage damage caused by T-2 toxin and the protective effect of chondroitin sulfate A nano-elemental selenium (CSA-SeNP) on the cartilage.METHODS: KBD chondrocytes and C28/I2 human chondrocyte cell lines were used. T-2 toxin, AKT inhibitor, and CSA-SeNP treatment experiments were conducted separately, with a treatment time of 24 hours. Autophagy was monitored using MDC staining, and mRFP-GFP-LC3 adenovirus, respectively. RT-qPCR and western blotting were used to detect the expression of the relevant genes and proteins.
RESULTS: The suppression of autophagy observed in KBD chondrocytes was replicated by applying 10 ng/mL T-2 toxin to C28/I2 chondrocytes for 24 hours. The AKT/TSCR/Rheb/mTOR signaling pathway was activated by T-2 toxin, which inhibits autophagy. The supplementation with CSA-SeNP alleviated the inhibition of autophagy by T-2 toxin through the AKT/TSCR/Rheb/mTOR signaling pathway.
CONCLUSIONS: Loss of autophagy regulated by the AKT/TSCR/Rheb/mTOR signaling pathway plays an important role in cartilage damage caused by T-2 toxin. CSA-SeNP supplementation attenuated inhibition of autophagy in chondrocyte by T-2 toxin by modulating this signaling pathway. These findings provide promising new targets for the prevention and treatment of cartilage disease.

Keywords:  AKT/TSCR/Rheb/mTOR signaling pathway; Chondrocyte autophagy; Chondroitin sulfate A nano-elemental selenium; T-2 toxin

DOI:  https://doi.org/10.1016/j.joca.2024.05.007
Int J Biol Macromol. 2024 May 23. pii: S0141-8130(24)03427-5. [Epub ahead of print]271(Pt 1): 132622

Structural investigation of Fun168A unraveling the recognition mechanism of endo-1,3-fucanase towards sulfated fucan.

Guangning Chen, Sheng Dong, Yuying Zhang, Jingjing Shen, Guanchen Liu, Fangyi Chen, Xinyu Li, Changhu Xue, Qiu Cui, Yingang Feng, Yaoguang Chang.

  BACKGROUND: Sulfated fucan has gained interest due to its various physiological activities. Endo-1,3-fucanases are valuable tools for investigating the structure and establishing structure-activity relationships of sulfated fucan. However, the substrate recognition mechanism of endo-1,3-fucanases towards sulfated fucan remains unclear, limiting the application of endo-1,3-fucanases in sulfated fucan research.SCOPE AND APPROACH: This study presented the first crystal structure of endo-1,3-fucanase (Fun168A) and its complex with the tetrasaccharide product, utilizing X-ray diffraction techniques. The novel subsite specificity of Fun168A was identified through glycomics and nuclear magnetic resonance (NMR).
KEY FINDINGS AND CONCLUSIONS: The structure of Fun168A was determined at 1.92 Å. Residues D206 and E264 acted as the nucleophile and general acid/base, respectively. Notably, Fun168A strategically positioned a series of polar residues at the subsites ranging from -2 to +3, enabling interactions with the sulfate groups of sulfated fucan through salt bridges or hydrogen bonds. Based on the structure of Fun168A and its substrate recognition mechanisms, the novel subsite specificities at the -2 and +2 subsites of Fun168A were identified. Overall, this study provided insight into the structure and substrate recognition mechanism of endo-1,3-fucanase for the first time and offered a valuable tool for further research and development of sulfated fucan.

Keywords:  Crystal structure; Fucanase; GH168

DOI:  https://doi.org/10.1016/j.ijbiomac.2024.132622