ANALYTICAL BIOCHEMISTRY
·
2024
·
IF 2.6
Cheng Li,Qinghui Chen,Jinqiao Rong,Houde He,Yu Lu,Yuxia Liu,Zhongfu Wang
As the main active glycoprotein of egg white, the biological functions of chicken ovomucin α- and β-subunit are closely related to the structure of glycans. However, the exact composition and structure of the subunit glycans are still unknown. We obtained highly pure chicken ovomucin α-subunit and β-subunit protein bands by the strategy combined with two-step isoelectric precipitation and SDS-PAGE gel electrophoresis. The ammonia-catalyzed one-pot procedure was then used to release and capture α-and β-subunit protein glycans with 1-phenyl- 3-Methyl-5-pyrazolone (PMP). The N / O -glycans of bis-PMP derivatives were purified and analyzed by LC-MS. More importantly, an effective dual modification was performed to accurately quantify neutral and sialylated O -glycans through methylamidation of sialic acid residues and simultaneously through carbonyl condensation reactions of reducing ends with PMP. We first showed that the α-subunit protein has only N -glycosylation modification, and the β-subunit only O -glycosylation, a total of 22 N -glycans and 20 O -glycans were identified in the α- and β-subunit, respectively. In addition, the complex N -glycan (47 %) and the sialylated O -glycan (77 %) are each major types of the above subunits. Such findings in this study provide a basis for studying the functional and biological activities of chicken ovomucin glycans.
研究领域:
生物化学
·
糖生物学
·
蛋白质组学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
RSC Advances
·
2019
·Vol. 9
·期 28
·15694-15702
·
IF 3.049
Lijing Nan,Jiao Li,Wanjun Jin,Ming Wei,Mengjun Tang,Chengjian Wang,Guiping Gong,Linjuan Huang,Ying Zhang,Zhongfu Wang
Mass spectrometry (MS) analysis combined with stable isotopic labeling is of great importance for quantitatively profiling abnormal sialylated O-glycans associated with disease development, but technically hindered by the poor releasing efficiency of O-glycans from glycoprotein or the labile nature of sialic acid residues at glycans. Herein, we developed an isotopic precursor based metabolic amplification and labeling (IPMAL) technique for relative quantitative profiling of the repertoire O-glycans between normal and tumor cells by ESI-MS. Two groups of cells were incubated with peracetylated benzyl-α-N-acetylgalactosamine (Ac3GalNAc-α-Bnd0) or a heavy labeled peracetylated benzyl-α-N-acetylgalactosamine (Ac3GalNAc-α-Bnd5) precursor respectively to amplify the repertoire of O-glycans as Bnd0/d5-O-glycans which could achieve the quantitative O-glycome analysis by ESI-MS after derivatization. The established method demonstrates desirable feasibility, accuracy (relative error (RE) ≤ 4.20%), reproducibility (coefficient of variation (CV) ≤ 7.61%, n = 3) and good quantitation linearity (R2 > 0.99, n = 3) for five Bn-O-glycans with 2 orders of magnitude. Finally, the method has been successfully applied to quantitative analysis of the repertoire O-glycome changes between normal human liver cell line L02 and human hepatoma cell line SMMC-7721. Moreover, the α-2,3/2,6 sialic acid isomers of Bn-O-glycans from these two cells have been further quantitatively distinguished when involved a sialic acid specific derivatization procedure.
研究领域:
生物化学
·
医学
·
分子生物学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
GLYCOBIOLOGY
·
2021
·Vol. 31
·期 8
·931-946
·
IF 4.313
Li Chunxia,Palma Angelina S,Zhang Pengtao,Zhang Yibing,Gao Chao,Silva Lisete M,Li Zhen,Trovão Filipa,Weishaupt Markus,Seeberger Peter H,Likhosherstov Leonid M,Piskarev Vladimir,Yu Jin,Westerlind Ulrika,Chai Wengang
Glycan microarrays have played important roles in detection and specificity assignment of glycan recognition by proteins. However, the size and diversity of glycan libraries in current microarray systems are small compared to estimated glycomes, and these may lead to missed detection or incomplete assignment. For microarray construction, covalent and noncovalent immobilization are the two types of methods used, but a direct comparison of results from the two platforms is required. Here we develop a chemical strategy to prepare lipid-linked probes from both naturally derived aldehyde-terminating and synthetic amino-terminating glycans that addresses the two aspects: expansion of sequence-defined glycan libraries and comparison of the two platforms. We demonstrate the specific recognition by plant and mammalian lectins, carbohydrate-binding modules and antibodies and the overall similarities from the two platforms. Our results provide new knowledge on unique glycan-binding specificities for the immune receptor Dectin-1 toward β-glucans and the interaction of rotavirus P[19] adhesive protein with mucin O-glycan cores.
研究领域:
生物化学
·
免疫学
·
分子生物学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
·
2022
·
IF 5.895
Yu Lu,Jie Liu,Zhenhua Li,Wenqing Li,Jing Liu,Linjuan Huang,Zhongfu Wang
Casein glycomacropeptide carries various O-glycan modifications, which, together with variations in the amino acid composition of the glycopeptide, may result in different biological activities. In this study, O-glycans of casein glycomacropeptide from bovine and caprine whey powder were qualitatively and quantitatively analyzed by LC-UV-ESI-MS/MS, and their immune activities and regulatory mechanisms were compared. O-Glycans’ total content was 1.54 times higher in bovine than in caprine glycomacropeptide. The glycoform H1N1S2 (H: hexose; N: N-acetylgalactosamine; and S: N-acetylneuraminic acid) accounted for nearly 50% of total glycomacropeptide O-glycans in bovine milk but less than 20% in caprine milk. Bovine glycomacropeptide glycosylation promoted the immune activity of RAW264.7 cells, which may be linked to a higher content of disialylated O-glycans. Glycomacropeptide from both milk sources significantly upregulated the mRNA expression of IL-1α, TNF-α, and IL-10 in RAW264.7 cells and activated the MAPK immunomodulatory signaling pathway. This study demonstrates the possible use of casein glycomacropeptide as an immunomodulatory agent.
研究领域:
食品科学
·
免疫学
·
生物化学
·
分子生物学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
·
2024
·
IF 7.7
Tong Zhao,Sining Liu,Yutong Shuai,Xinyi Zhang,Min Chen,Sijie Pei,Yuxi Duan,Shukai Wang,Yu Lu,Zhongfu Wang,Guiping Gong,Linjuan Huang
Pectin is a structurally complex heteropolysaccharide that affects intestinal microorganisms and mucin O -glycans. The present study employed an in vitro model to investigate dynamic changes in microbiota during pectin fermentation. Residual pectin fragments arising from its fermentation were applied to HT29-MTX cells to study the effect of pectin structure on mucin O -glycosylation. Prevotella , Bacteroides , and Parabacteroides were found to preferentially degrade galactose, arabinose, and on the rhamnogalacturonan RG-I side chain region and methyl esterification groups of pectin. Bifidobacterium , Enterococcus , Megamonas , and Dorea metabolized the galacturonic HG region on pectin to produce butyrate. All pectin fragments were found to up-regulate total O -glycans (1.55–2.73 fold) and neutral O -glycans (1.11–1.49 fold) on HT29-MTX mucins. The large HG fragment (81.04 kDa) increased significantly the amount of non-fucosylated glycans (by 2.46-fold); whereas the small HG fragment (16.02 kDa) promoted fucosylated (by 9.25 fold), and especially di-fucosylated O -glycans. Collectively, these results demonstrate that gut microorganisms degrade pectin fragments in the following order of utilization: RG-I, RG-II, and HG. The small fragment of HG improves the expression of fucosylated O -glycans in HT29-MTX cells, mainly owing to an increase in di-fucosylated O -glycans.
研究领域:
微生物学
·
胃肠病学
·
细胞生物学
·
营养科学
✓ 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）