• BMB Reports
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• v.45 no.6
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• pp.323-330
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• 2012

The glycome consists of all glycans (or carbohydrates) within a biological system, and modulates a wide range of important biological activities, from protein folding to cellular communications. The mining of the glycome for disease markers represents a new paradigm for biomarker discovery; however, this effort is severely complicated by the vast complexity and structural diversity of glycans. This review summarizes recent developments in analytical technology and methodology as applied to the fields of glycomics and glycoproteomics. Mass spectrometric strategies for glycan compositional profiling are described, as are potential refinements which allow structure-specific profiling. Analytical methods that can discern protein glycosylation at a specific site of modification are also discussed in detail. Biomarker discovery applications are shown at each level of analysis, highlighting the key role that glycoscience can play in helping scientists understand disease biology.

• Genomics & Informatics
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• v.8 no.2
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• pp.63-69
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• 2010

Monofunctional biosynthetic peptidoglycan transglycosylase (MBPT) catalyzes the formation of the glycan chain in bacterial cell walls from peptidoglycan subunits: N-acetylglucosamine (NAG) and acetylmuramic acid (NAM). Bifunctional glycosyltransferases such as the penicillin binding protein (PBP) have peptidoglycan glycosyltransferase (PGT) on their C terminal end which links together the peptidoglycan subunits while transpeptidase (TP) on the N terminal end cross-links the peptide moieties on the NAM monosaccharide of the peptide subunits to create the bacterial cell wall. The singular function of MBPT resembles the C terminal end of PBP as it too contains and utilizes a similar PGT domain. In this article we analyzed the infectious and non infectious protein sequences of MBPT from 31 different strains of bacteria using a variety of bioinformatic tools. Motif analysis, dot-plot comparison, and phylogenetic analysis identified a number of significant differences between infectious and non-infectious protein sequences. In this paper we have made an attempt to explain, analyze and discuss these differences from an evolutionary perspective. The results of our sequence analysis may open the door for utilizing MBPT as a new target to fight a variety of infectious bacteria.

• Mass Spectrometry Letters
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• v.13 no.4
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• pp.139-145
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• 2022

Protein glycosylation is a common post-translational modification by non-template-based biosynthesis. In fungal biotechnology, which has great applications in pharmaceuticals and industries, the importance of research on fungal glycoproteins and glycans is accelerating. In particular, the importance of quantitative analysis of fungal glycans is emerging in research on the production of filamentous fungal proteins by genetic modification. Reliable mass spectrometry-based techniques for quantitative glycomics have evolved into chemical, enzymatic, and metabolic stable isotope labeling methods. In this study, we intend to expand quantitative glycomics by metabolic isotope labeling of glycans in Aspergillus niger, a filamentous fungus model, by the MILPIG method. We demonstrate that incubation of filamentous fungi in a culture medium with carbon-13 labeled glucose (1-¹³C₁) efficiently incorporates carbon-13 into N-linked glycans. In addition, for quantitative validation of this method, light and heavy glycans are mixed 1:1 to show the performance of quantitative analysis of various N-linked glycans simultaneously. We have successfully quantified fungal glycans by MILPIG and expect it to be widely applicable to glycan expression levels under various biological conditions in fungi.

• Mass Spectrometry Letters
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• v.6 no.3
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• pp.53-58
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• 2015

Recombinant erythropoietins (EPOs) are an important class of biotherapeutics that stimulate red blood cell production. The quality, safety, and potency of EPO variants are determined largely by their glycosylation, which makes up nearly half their mass. Thus, detailed glycomic analyses are important to assess biotherapeutic quality and establish the equivalency of biosimilar EPOs now coming to market. High-resolution mass spectrometry (MS) has recently emerged as the premier tool for glycan analysis in EPOs. Using the accurate mass measurements provided by high-resolution MS, the compositions of even large, complex glycans can easily be determined. When combined with a nano-LC separation, differentiation of structural isomers also becomes a possibility. These components, together, provide a comprehensive picture of biotherapeutic glycosylation. In this review, we provide an overview of MS-based analytical platform for glycomic characterization of EPO biotherapeutics and biosimilars.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.14
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• pp.5601-5605
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• 2014

Background: Agrocybe aegerita Lectin (AAL) has been identified to have high affinity for sulfated and ${\alpha}2$-3-linked sialic acid glycoconjugates, especially the sulfated and sialyl TF (Thomsen-Friedenreich) disaccharide. This study was conducted to investigate the clinicopathological and prognostic value of AAL in identifying aberrant glycosylation in colorectal cancer (CRC). Materials and Methods: Glycoconjugate expression in 59 CRC tissues were detected using AAL-histochemistry. Clinicopathological associates of expression were analyzed with chisquare test or Fisher's exact test. Relationships between expression and the various clinicopathological parameters was estimated using Kaplan-Meier analysis and Cox regression models. Results: AAL specific glycoconjugate expression was significantly higher in tumor than corresponding normal tissues (66.1% and 46.1%, respectively, p=0.037), correlating with depth of invasion (p=0.015) and TNM stage (p=0.024). Patients with lower expression levels had a significantly higher survival rate than those with higher expression (p=0.046 by log rank test and p=0.047 by Breslow test for overall survival; p=0.054 by log rank test and P=0.038 by Breslow test for progress free survival). A marginally significant association was found between AAL specific glycoconjugate expression and overall survival by univariate Cox regression analysis (p=0.059). Conclusions: Lower AAL specific glycoconjugate expression is a significant favorable prognostic factor for overall and progress free survival in CRC. This is the first report about the employment of AAL for histochemical analysis of cancer tissues. The binding characteristics of AAL means it has potential to become a powerful tool for the glycan investigation and clinical application.

• Genomics & Informatics
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• v.21 no.4
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• pp.46.1-46.10
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• 2023

Colon adenocarcinoma (COAD) is the predominant type of colorectal cancer. Early diagnosis and treatment can significantly improve the prognosis of COAD patients. Anoctamin 7 (ANO7), an anion channel protein, has been implicated in prostate cancer and other types of cancer. In this study, we analyzed the expression of ANO7 and its correlation with clinicopathological characteristics among COAD patients using the Gene Expression Profiling Interactive Analysis 2 (GEPIA2) and the University of Alabama at Birmingham CANcer (UALCAN) databases. The GEPIA2, Kaplan-Meier plotter, and the Survival Genie platform were employed for survival analysis. The co-expression network and potential function of ANO7 in COAD were analyzed using GeneFriends, the Database for Annotation, Visualization and Integrated Discovery (DAVID), GeneMANIA, and Pathway Studio. Our data analysis revealed a significant reduction in ANO7 expression levels within COAD tissues compared to normal tissues. Additionally, ANO7 expression was found to be associated with race and histological subtype. The COAD patients exhibiting low ANO7 expression had lower survival rates compared to those with high ANO7 expression. The genes correlated with ANO7 were significantly enriched in proteolysis and mucin type O-glycan biosynthesis pathway. Furthermore, ANO7 demonstrated a direct interaction and a positive co-expression correlation with mucin 2 (MUC2). In conclusion, our findings suggest that ANO7 might serve as a potential prognostic biomarker and potentially plays a role in proteolysis and mucin biosynthesis in the context of COAD.

• Molecules and Cells
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• v.24 no.3
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• pp.416-423
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• 2007

Alterations in the glycan chains of cell surface glycoconjugates are frequently involved biological processes such as cell-cell interaction, cell migration, differentiation and development. Cultured embryonic (E18) rat cortical neurons underwent apoptosis in response to camptothecin, and lectin histochemistry showed that binding to apoptotic neurons of FITC-conjugated Maackia amurensis agglutinin (MAA), which is specific for terminal ${\alpha}2,3$-sialic acid residues, increased progressively with increasing concentrations of camptothecin. Analysis of the total proteins of apoptotic neurons by SDS-PAGE, and lectin blotting using HRP-labeled MAA, revealed that the expression of terminal ${\alpha}2,3$-sialic acid residues on an unknown protein with an apparent molecular mass of 25.6 kDa also increased in apoptotic neurons. NP-HPLC analysis of the total cellular N-glycans of normal and apoptotic neurons demonstrated that the expression of structurally simpler biantennary types of N-glycans fell by 49% during apoptosis whereas the more branched triantennary types of N-glycans with terminal sialic acid residues increased by up to 59%. These results suggest that increased surface expression of ${\alpha}2,3$-sialic acid residues and hyperglycosylation of N-glycans is a common feature of cellular responses to changes in cell physiology such as tumorigenesis and apoptosis.

• Korean Journal of Microbiology
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• v.46 no.1
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• pp.96-103
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• 2010

Oligosaccharyltransferase (OTase) catalyzes the transfer of a lipid-linked oligosaccharide (LLO) to the nascent polypeptide. Most eukaryotes have an OTase composed of a multisubunit protein complex. However, the kinetoplastid Leishmania major and the bacterium Campylobacter jejuni have only a single subunit for OTase activity, Stt3p and PglB, respectively. In this study, a new in vitro assay for OTase was developed by using a fluorescent peptide containing N-glycosylation sequon, Asn-Xaa-Thr/Ser, where Xaa can be any amino acid residue except Pro. L. major Stt3p and C. jejuni PglB as a model OTase enzyme demonstrated the formation of glycopeptides from a fluorescent peptide through OTase activities. For separation and measurement of the glycopeptides produced by the OTases, Tricine-SDS-PAGE, a lectin column and fluorospectrophotometer, and HPLC were applied. Comparison of these assay methods for analyzing a fluorescent glycopeptide showed HPLC analysis is the best method for separation of glycopeptides and nonglycosylated peptides as well as for quantify the peptides than other methods.

• Journal of Life Science
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• v.26 no.3
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• pp.275-281
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• 2016

Glycan modification is important in pharmaceutical industry. Especially, sialic acid affects the bioactivity and stability of medicine. Milk of pig has been used as bioreactor to produce various pharmaceutical proteins. Therefore, it is necessary to modify the glycan chain in pig mammary grand. β-1,4-N-Acetylglucosaminyltransferase A (pMGAT4A) is one of the essential enzymes for increase of sialic acid content, but pig MGAT4A is unclear. In this study, the pMGAT4A was identified and characterized. The pMGAT4A has 1638 nucleotides encoding 535 amino acids and type II membrane topology, which is one of the common features in many glycosyltransferases. The gene was strongly expressed in liver and mammary gland, whereas was weakly expressed in small intestine, stomach and bladder. For functional test, HA-tagged MGAT4A was over-expressed in porcine kidney (PK-15) cell line. Forced expression of pMGAT4A gene was identified by qPCR, and we identified that pMGAT4A is located in Golgi complex by co- staining with HA antibody and BODIPY TR ceramide. In addition, we identified the increase of mannose-β-1,4-N-acetylglucosamine structure by ELISA and immunofluorescence using Datura stramonium agglutinin (DSA), which recognizes mannose-β-1,4-Nacetylglucosamine. Through the specific activity analysis, we showed that pMGAT4A modified bi-antennary to tri-antennary. This event affects sialic acid content. Therefore, we thought that over-expression of pMGAT4A will be necessary in pig mammary grand for improved medicine.

• Animal Bioscience
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• v.37 no.6
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• pp.982-992
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• 2024

Objective: Jining Grey goat is a local Chinese goat breed that is well known for its high fertility and excellent meat quality but shows low meat production performance. Numerous studies have focused on revealing the genetic mechanism of its high fertility, but its highlighting meat quality and muscle growth mechanism still need to be studied. Methods: In this research, an integrative analysis of the genomics and transcriptomics of Jining Grey goats compared with Boer goats was performed to identify candidate genes and pathways related to the mechanisms of meat quality and muscle development. Results: Our results overlap among five genes (ABHD2, FN1, PGM2L1, PRKAG3, RAVER2) and detected a set of candidate genes associated with fatty acid metabolism (PRKAG3, HADHB, FASN, ACADM), amino acid metabolism (KMT2C, PLOD3, NSD2, SETDB1, STT3B, MAN1A2, BCKDHB, NAT8L, P4HA3) and muscle development (MSTN, PPARGC1A, ANKRD2). Several pathways have also been detected, such as the FoxO signaling pathway and Apelin signaling pathway that play roles in lipid metabolism, lysine degradation, N-glycan biosynthesis, valine, leucine and isoleucine degradation that involving with amino acid metabolism. Conclusion: The comparative genomic and transcriptomic analysis of Jining Grey goat and Boer goat revealed the mechanisms underlying the meat quality and meat productive performance of goats. These results provide valuable information for future breeding of goats.