CELL RESEARCH
·
2023
·
IF 44.1
Ai Yuan-li,Wang Wei-jia,Liu Fan-jian,Fang Wei,Chen Hang-zi,Wu Liu-zheng,Hong Xuehui,Zhu Yuekun,Zhang Ci-xiong,Liu Long-yu,Hong Wen-bin,Zhou Bo,Chen Qi-tao,Wu Qiao
Pyroptosis is a type of regulated cell death executed by gasdermin family members. However, how gasdermin-mediated pyroptosis is negatively regulated remains unclear. Here, we demonstrate that mannose, a hexose, inhibits GSDME-mediated pyroptosis by activating AMP-activated protein kinase (AMPK). Mechanistically, mannose metabolism in the hexosamine biosynthetic pathway increases levels of the metabolite N -acetylglucosamine-6-phosphate (GlcNAc-6P), which binds AMPK to facilitate AMPK phosphorylation by LKB1. Activated AMPK then phosphorylates GSDME at Thr6, which leads to blockade of caspase-3-induced GSDME cleavage, thereby repressing pyroptosis. The regulatory role of AMPK-mediated GSDME phosphorylation was further confirmed in AMPK knockout and GSDME T6E or GSDME T6A knock-in mice. In mouse primary cancer models, mannose administration suppressed pyroptosis in small intestine and kidney to alleviate cisplatin- or oxaliplatin-induced tissue toxicity without impairing antitumor effects. The protective effect of mannose was also verified in a small group of patients with gastrointestinal cancer who received normal chemotherapy. Our study reveals a novel mechanism whereby mannose antagonizes GSDME-mediated pyroptosis through GlcNAc-6P-mediated activation of AMPK, and suggests the utility of mannose supplementation in alleviating chemotherapy-induced side effects in clinic applications.
研究领域:
癌症生物学
·
细胞生物学
·
药理学
·
分子生物学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
BIOTECHNOLOGY LETTERS
·
2022
·Vol. 44
·期 4
·623-633
·
IF 2.716
Hao Zhi-kui,Li Jian-song,Wang Dan-hua,He Fei,Xue Jing-shi,Yin Liang-hong,Zheng Hua-bao
Objectives Shellfish waste is a primary source for making N -acetyl- d -glucosamine. Thus, establishing a high-efficiency and low-cost bioconversion method to produce N -acetyl- d -glucosamine directly from shellfish waste was promising.
Results A mutant C81 was obtained from Chitinolyticbacter meiyuanensis SYBC-H1 via 60 Co-γ irradiation. This mutant C81 showed the highest chitinase activity of 9.8 U/mL that was 85% higher than the parent strain. The mutant C81 exhibted improved antioxidant activities, including total antioxidant capacity, superoxide radical ability, and hydroxyl radical scavenging ability, compared to that of the parent strain. Four out of nine organic solvents increased the chitinase activity by 1.9%, 6.8%, 11.7%, and 15.8%, corresponding to methylbenzene, n-heptane, petroleum ether, and n-hexane, respectively. The biphase system composed of aqueous and hexane presented a five-fold reduction of cell viability compared to the control. Using a continuous fermentation bioconversion process, 4.2 g/L GlcNAc was produced from crayfish shell powder with a yield of 80% of the chitin content. Conclusions This study demonstrated that the mutant C81 is suitable for converting crayfish shell powder into GlcNAc in an aqueous-organic system.
研究领域:
微生物学
·
生物化学
·
生物工程
✓ 已验证
DOI
PubMed
高影响因子
TALANTA
·
2020
·
IF 5.339
Nan Yin,Ruihuan Zhao,Jie Zhang,Dingding Yang,Zhimin Guo,Rui Liu,Xin Yao
Abnormal O-linked N-acetylglucosamine (O-GlcNAc) concentrations have been associated with many diseases, but the lack of accurate detection method limited O-GlcNAc to be used as a biomarker in clinical diagnosis. Then O-GlcNAc transferase (OGT) has drawn researchers’ attention as it closed related to the level of O-GlcNAc and be considered to be a promising new target for diseases diagnosis. Nevertheless, the existing OGT detection methods are either need labeling or the sensitity can not meet the needs of clinic testing. Herein, a label-free and sensitive SPR biosensor was developed for accurate detection of OGT based on a multi-functional peptide. The designed peptide contains three recognition sites, one is the cleavage site of protease K, one is the O-GlcNAcylated site by OGT, and another is six histidine which be used as the signal report probe to recognize Ni 2+ . The immobilized peptide would be cleavaged by proteinase K, then the His-tag residue part will leave the surface of Au film, resulting less His-tag could bind to Ni 2+ and a small SPR signal would be record. If the peptide is O-GlcNAcylated by OGT, the cleaving reaction would be limited due to the adjacent site of O-GlcNAcylation. Then more His-tag can be left on the Au film and a bigger SPR signal could be record, this signal is associated with the concentration of OGT. Utilizing the change of the peptide configuration as a signal report probe for OGT detection not only avoids labeling of peptide, but also makes the method more sensitive. The determination linear range of OGT is from 2.00 × 10 −13 to 5.00 × 10 −8 M with a detection limit of 1.19 × 10 −13 M, and the separation of two enzyme reactions ensured the high selectivity of the method. Finally, the sensing system was successfully used for OGT detection in blood samples with satisfied recovery. In summary, the label-free SPR platform for accurate detection of OGT in real samples is helpful to promote OGT serve as a biomarker for early clinical diagnosis of O-GlcNAc related diseases.
研究领域:
生物化学
·
分析化学
·
生物传感器
·
临床诊断
✓ 已验证
产品关联
DOI
PubMed
高影响因子
BIOCHEMICAL ENGINEERING JOURNAL
·
2015
·
IF 2.467
Cong Gao,Alei Zhang,Kequan Chen,Zhikui Hao,Junmao Tong,Pingkai Ouyang
In this study, a novel chitinase-producing bacterium Chitinibacter sp. GC72 was isolated and investigated for N -acetyl-D-glucosamine production from crayfish shell enzymatic degradation. A dimeric chitinase was purified, which had an optimal activity at a pH of 6.8 and 40 °C. The metal ions Al 3+ , Cu 2+ , and Zn 2+ inhibited the chitinase activity, whereas Ca 2+ , Mn 2+ , and Mg 2+ promoted the activity. The chitinase was active on p -NP-GlcNAc with apparent K m value of 152.83 μmol/L and V m value of 49.12 μmol/L min at 37 °C. Based on the hydrolysate formed, the chitinase was characterized as an exo-hydrolytic N -acetyl glucosaminidase. Furthermore, combined treatment of ultra-micro grinding and ultrasonic with direct enzymatic hydrolysis of crayfish shell for GlcNAc production was investigated, which resulted in 15.2 g of GlcNAc production from 100 g of crayfish shell following chitinase degradation for 36 h.
研究领域:
微生物学
·
酶学
·
生物技术
✓ 已验证
产品关联
DOI
高影响因子
PROCESS BIOCHEMISTRY
·
2024
·
IF 3.7
Yang Xu,Bo OuYang,Linye Deng,Mei Liao,Tianhui Tang,Dongming Lan,Yonghua Wang
Chitin represents the most abundant source of renewable biomass in marine ecosystems, and its degradation by chitinase holds potential for numerous industrial applications. In this study, a novel hyperthermophilic chitinase, TbChi52, derived from a deep-sea bacterium of the Thermotogae phylum was introduced. TbChi52 exhibits optimal enzymatic activity at 90 ℃ and pH 5.5. Furthermore, it shows exceptional thermal stability characterized by a prolonged thermal denaturation half-life and a significant melting temperature. Functionally, TbChi52 displays dual activities, acting as both an exo-chitinase and a β-N-acetylaminoglucosidase (NAGase). It yields N-acetyl-D-glucosamine (GlcNAc) as the primary product during the hydrolysis of colloidal chitin. Leveraging the robust thermo-stability of TbChi52, a streamlined, one-step purification protocol was developed. A mere 10-minute thermal treatment at 75 ℃ suffices to yield a highly pure TbChi52 preparation with minimal enzyme activity loss in chitin hydrolysis, achieving an 87.4 % hydrolysis rate of colloidal chitin and a 92.7 % purity of GlcNAc. Collectively, the outstanding thermal properties and the simple preparation of TbChi52 highlight its potential for industrial applications. It provides a sustainable and eco-friendly approach to harnessing the world's most abundant marine biomass, along with serving as a potential enzyme-engineered template tailored for specific applications.
研究领域:
海洋生物学
·
酶学
·
工业生物技术
✓ 已验证
产品关联
DOI
高影响因子
MOLECULES
·
2018
·Vol. 23
·期 7
·1649
·
IF 3.098
Qi Cheng,Yalu Cui,Nao Xiao,Jishun Lu,Chen-Jie Fang
TheO-linked β-N-acetylglucosamine (O-GlcNAc) modification is an abundant post-translational modification in eukaryotic cells, which plays a fundamental role in the activity of many cells and is associated with pathologies like type II diabetes, Alzheimer’s disease or some cancers. However, the precise connexion betweenO-GlcNAc-modified proteins and their function in cells is largely undefined for most cases. Confocal microscopy is a powerful and effective tool for in-cell elucidation of the function of biological molecules. Chemical labeling of non-ultraviolet or non-fluorescent carbohydrates with fluorescent tag is an essential step that makes intra-cellular microscopic inspection possible. Here we report a strategy based on the 1,3-dipolar cycloaddition, called click chemistry, between unnaturalN-acetylglucosamine (GlcNAc) analogues Ac4GlcNAc (substituted with an azido group) and the corresponding fluorescent tag Ru(bpy)2(Phen-alkyne)Cl2(4) to synthesize the fluorescent dye Ru(bpy)2(Phen-Ac4GlcNAc)Cl2(5) under mild and neutral reaction conditions. Moreover,5showed good stability, desirable fluorescence characteristics, and exhibited rather low levels of cytotoxicity against sensitive MCF-7 cells. Additionally, we have achieved successful fluorescent imaging of5transported in living MCF-7 cells. Cell images displayed that proteins are potentially labelled with5in the cytoplasm.Keywords:O-GlcNAc;click chemistry;cell imaging;fluorescent labeling;living cells
研究领域:
生物化学
·
分子生物学
·
细胞生物学
·
化学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
·
2024
·
IF 5.7
Jiao Li,Ruiyang Li,Na Shang,Yan Men,Yi Cai,Yan Zeng,Weidong Liu,Jiangang Yang,Yuanxia Sun
The addition of the O-linked N-acetylglucosamine (O-GlcNAc) is a significant modification for active molecules, such as proteins, carbohydrates, and natural products. However, the synthesis of terpenoid glycoside derivatives decorated with GlcNAc remains a challenging task due to the absence of glycosyltransferases, key enzymes for catalyzing the transfer of GlcNAc to terpenoids. In this study, we demonstrated that the enzyme mutant UGT74AC1T79Y/L48M/R28H/L109I/S15A/M76L/H47R efficiently transferred GlcNAc from uridine diphosphate (UDP)-GlcNAc to a variety of terpenoids. This powerful enzyme was employed to synthesize GlcNAc-decorated derivatives of terpenoids, including mogrol, steviol, andrographolide, protopanaxadiol, glycyrrhetinic acid, ursolic acid, and betulinic acid for the first time. To unravel the mechanism of UDP-GlcNAc recognition, we determined the X-ray crystal structure of the inactivated mutant UGT74AC1His18A/Asp111A in complex with UDP-GlcNAc at a resolution of 1.66 Å. Through molecular dynamic simulation and activity analysis, we revealed the molecular mechanism and catalytically important amino acids directly involved in the recognition of UDP-GlcNAc. Overall, this study not only provided a potent biocatalyst capable of glycodiversifying natural products but also elucidated the structural basis for UDP-GlcNAc recognition by glycosyltransferases.
研究领域:
生物化学
·
结构生物学
·
合成生物学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
Nature Communications
·
2024
·Vol. 15
·期 1
·1-16
·
IF 14.7
Cao Shiyang,Wang Tong,Ren Yifan,Wu Gengshan,Zhang Yuan,Tan Yafang,Zhou Yazhou,Chen Hongyan,Zhang Yu,Song Yajun,Yang Ruifu,Du Zongmin
Post-translational addition of O-linked N-acetylglucosamine (O-GlcNAc) to proteins is commonly associated with a variety of stress responses and cellular processes in eukaryotes, but its potential roles in bacteria are unclear. Here, we show that protein HmwC acts as an O-GlcNAc transferase (OGT) responsible for O-GlcNAcylation of multiple proteins in Yersinia pestis , a flea-borne pathogen responsible for plague. We identify 64 O-GlcNAcylated proteins (comprising 65 sites) with differential abundance under conditions mimicking the mammalian host (Mh) and flea vector (Fv) environments. Deletion of hmwC , encoding a putative OGT, structurally distinct from any existing member of the GT41 family, results in reduced O-GlcNAcylation, reduced growth, and alterations in virulence properties and survival under stress. Purified HmwC can modify target proteins in vitro using UDP-GlcNAc as sugar donor. One of the target proteins, OsdY, promotes Y. pestis survival under oxidative stress conditions. Thus, our results support that regulation of antioxidative responses through O-GlcNAcylation may be a conserved process shared by prokaryotes and eukaryotes.
研究领域:
微生物学
·
病理学
·
分子生物学
·
传染病学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
·
2026
·Vol. 74
·期 28
·22149-22158
·
IF 6.7
Sun Ze-Rui,Xia Yu-Xin,Li Yi-Ran,Liu Li,Voglmeir Josef
The deacetylation of N -acetylglucosamine
(GlcNAc)
to glucosamine (GlcN) is a key step in enzymatic chitin valorization.
We identified a novel GlcNAc deacetylase (AbNGD) from Acinetobacter baumannii with high substrate specificity
and catalytic activity but poor thermal stability (half-life of 5.5
h at 37 ° C). Using the FireProt platform, we generated a combinatorial
mutant, AbNGD-M3 (T93P–C153T-A244M). Its optimal temperature
increased to 37 ° C, and its half-life reached 28.1 h (5.1-fold
improvement), while retaining 97% of wild-type catalytic efficiency
( k cat / K m =
3.4 mM –1 ·s –1 ). Site-directed
mutagenesis confirmed Asp54 and His160 as key catalytic residues.
Molecular dynamics simulations revealed that T93P reduces backbone
flexibility, C153T enhances β-sheet rigidity via hydrogen bonding,
and A244M improves hydrophobic packing by filling a core cavity. These
distal mutations collectively balance stability and activity. Substrate
channel analysis identified a likely transport pathway centered on
T1. This study expands the deacetylase repertoire and provides a framework
for engineering stable industrial enzymes.
✓ 已验证
产品关联
DOI
PubMed
高影响因子
MOLECULAR PHARMACEUTICS
·
2023
·
IF 4.9
Bingyi Lin,Siyuan Chai,Qijun Zhang,Yuejie Lu,Jiahao Hu,Jie Zhang,Yong-Zhong Du,Liming Wu
O-linked-N-acetylglucosaminylation (O-GlcNAcylation) plays a key role in hepatocellular carcinoma (HCC) development, and the inhibition of O-GlcNAcylation has therapeutic potential. To decrease the systemic adverse events and increase targeting, we used sialic acid (SA)-decorated liposomes loaded with OSMI-1, an inhibitor of the O-GlcNAcylation, to further improve the anti-HCC effect. Fifty pairs of HCC tissue samples and the cancer genome atlas database were used to analyze the expression of O-GlcNAc transferase (OGT) and its effects on prognosis and immune cell infiltration. OSMI-1 cells were treated with SA and liposomes. Western blotting, immunofluorescence, cell proliferation assay, flow cytometry, enzyme-linked immunosorbent assay, immunohistochemistry, and tumorigenicity assays were used to investigate the antitumor effect of SA-modified OSMI-1 liposomes in vitro and in vivo. OGT was highly expressed in HCC tissues, negatively correlated with the degree of tumor infiltration of CD8+ and CD4+T cells and prognosis, and positively correlated with the degree of Treg cell infiltration. SA-modified OSMI-1 liposome (OSMI-1-SAL) was synthesized with stable hydrodynamic size distribution. Both in vitro and in vivo, OSMI-1-SAL exhibited satisfactory biosafety and rapid uptake by HCC cells. Compared to free OSMI-1, OSMI-1-SAL had a stronger capacity for suppressing the proliferation and promoting the apoptosis of HCC cells. Moreover, OSMI-1-SAL effectively inhibited tumor initiation and development in mice. OSMI-1-SAL also promoted the release of damage-associated molecular patterns, including anticalreticulin, high-mobility-group protein B1, and adenosine triphosphate, from HCC cells and further promoted the activation and proliferation of the CD8+ and CD4+T cells. In conclusion, the OSMI-1-SAL synthesized in this study can target HCC cells, inhibit tumor proliferation, induce tumor immunogenic cell death, enhance tumor immunogenicity, and promote antitumor immune responses, which has the potential for clinical application in the future.
✓ 已验证
产品关联
DOI
PubMed
高影响因子
POLYMER DEGRADATION AND STABILITY
·
2021
·
IF 5.03
Ning Zhou,Pengfan Yang,Jie Chen,Guoguang Wei,Chengyong Wang,Alei Zhang,Kequan Chen,Pingkai Ouyang
In this study, the effect of pretreatment with three organic solvent systems on enzymatic hydrolysis of chitin was investigated. Firstly, genes coding three chitinolytic enzymes Cm Chi1, Cm Chi4, and Cm NAGase from Chitinolyticbacter meiyuanensis SYBC-H1 were cloned and functionally expressed. Then, a cocktail capable of producing GlcNAc from chitin was formulated with 71.4% recombinant Cm Chi1, 23.8% recombinant Cm Chi4, and 4.8% recombinant Cm NAGase. Next, three organic solvent systems, including LiCl/dimethylacetamide, CaCl 2 /Methanol, and hexafluoroisopropanol were applied to treat chitin, resulting in 4.90–8.21-fold improvement of hydrolysis efficiency compared to raw chitin. The decreased crystallinity and nanofiber size structure without deacetylation, confirmed by SEM, FTIR, XRD, and EA, aided in the increased efficiency. Finally, yields of 20.97 mM, 34.50 mM, and 32.53 mM of GlcNAc with a purity of 96% were respectively obtained from 10 g/L of CLD, CHP, and CCM in 24 h.
研究领域:
生物技术
·
生物化学
·
材料科学
✓ 已验证
产品关联
DOI
高影响因子
MOLECULES
·
2022
·Vol. 27
·期 15
·4720
·
IF 4.927
Xinjun Yu,Zengchao Jiang,Xiaodan Xu,Changyi Huang,Zheyi Yao,Xiao Yang,Yinjun Zhang,Dongsheng Wang,Chun Wei,Xuwei Zhuang
Chitin, the second richest polymer in nature, is composed of the monomer N-acetylglucosamine (GlcNAc), which has numerous functions and is widely applied in the medical, food, and chemical industries. However, due to the highly crystalline configuration and low accessibility in water of the chitin resources, such as shrimp and crab shells, the chitin is difficult utilize, and the traditional chemical method causes serious environment pollution and a waste of resources. In the present study, three genes encoding chitinolytic enzymes, including the N-acetylglucosaminidase fromOstrinia furnacalis(OfHex1), endo-chitinase fromTrichoderma viride(TvChi1), and multifunctional chitinase fromChitinolyticbacter meiyuanensis(CmChi1), were expressed in thePichia pastorissystem, and the positive transformants with multiple copies were isolated by the PTVA (post-transformational vector amplification) method, respectively. The three recombinants OfHex1, TvChi1, and CmChi1 were induced by methanol and purified by the chitin affinity adsorption method. The purified recombinants OfHex1 and TvChi1 were characterized, and they were further used together for degrading chitin from shrimp and crab shells to produce GlcNAc through liquid-assisted grinding (LAG) under a water-less condition. The substrate chitin concentration reached up to 300 g/L, and the highest yield of the product GlcNAc reached up to 61.3 g/L using the mechano-enzymatic method. A yield rate of up to 102.2 g GlcNAc per 1 g enzyme was obtained.Keywords:chitinase;shrimp and crab shell;N-acetylglucosamine;mechanoenzymology;heterologous expression
研究领域:
生物化学
·
生物技术
·
环境科学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）