Jun Hirabayashi

Kagawa University, Takamatu, Kagawa, Japan

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Publications (262)980.79 Total impact

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    ABSTRACT: Leguminous lectins have a conserved carbohydrate recognition site comprising four loops (A-D). Here, we randomly mutated the sequence and length of loops C and D of peanut agglutinin (PNA) and expressed the proteins on the surface of mouse green fluorescent protein (GFP)-reporter cells. Flow cytometry, limiting dilution, and cDNA cloning were used to screen for several mutated PNAs with distinct properties. The mutated PNA clones obtained using NeuAcα2-6(Galβ1-3)GalNAc as a ligand showed preference for NeuAcα2-6(Galβ1-3)GalNAc rather than non-sialylated Galβ1-3GlcNAc, whereas wild-type PNA binds to Galβ1-3GlcNAc but not sialylated Galβ1-3GalNAc. Sequence analyses revealed that for all of the glycan-reactive mutated PNA clones, (i) loop C was eight amino acids in length, (ii) loop D was identical to that of wild-type PNA, (iii) residue 127 was asparagine, (iv) residue 125 was tryptophan, and (v) residue 130 was hydrophobic tyrosine, phenylalanine, or histidine. The sugar-binding ability of wild-type PNA was increased nine-fold when Tyr125 was mutated to tryptophan, and that of mutated clone C was increased more than 30-fold after His130 was changed to tyrosine. These results provide an insight into the relationship between the amino acid sequences of the carbohydrate recognition site and sugar-binding abilities of leguminous lectins.
    08/2015; 5(3):1540-62. DOI:10.3390/biom5031540
  • Dan Hu · Hiroaki Tateno · Jun Hirabayashi ·
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    ABSTRACT: In the post genomic era, glycomics - the systematic study of all glycan structures of a given cell or organism - has emerged as an indispensable technology in various fields of biology and medicine. Lectins are regarded as "decipherers of glycans", being useful reagents for their structural analysis, and have been widely used in glycomic studies. However, the inconsistent activity and availability associated with the plant-derived lectins that comprise most of the commercially available lectins, and the limit in the range of glycan structures covered, have necessitated the development of innovative tools via engineering of lectins on existing scaffolds. This review will summarize the current state of the art of lectin engineering and highlight recent technological advances in this field. The key issues associated with the strategy of lectin engineering including selection of template lectin, construction of a mutagenesis library, and high-throughput screening methods are discussed.
    Molecules 05/2015; 20(5):7637-7656. DOI:10.3390/molecules20057637 · 2.42 Impact Factor
  • Jun Hirabayashi · Hiroaki Tateno · Yasuko Onuma · Yuzuru Ito ·
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    ABSTRACT: Human pluripotent stem cells (hPSCs), represented by embryonic stem (hESCs) and induced pluripotent stem cells (hiPSCs), are attracting increasing attention in various research fields. However, their application in a clinical scenario must overcome an important hurdle given that these cells are potentially tumorigenic. This inherent problem becomes more significant as the number of transplanted cells becomes larger. In this Progress Report, recent findings concerning a novel glycan marker for hPSCs are described, as well as attempts made in relation to its practical application to regenerative medicine. In line with current thinking in the glycoscience field, it is assumed that cellular glycomes are closely related to cell functions. Based on this premise, hESCs and hiPSCs are analyzed by an advanced glycan profiling technology-the high-density lectin microarray. It is found that all human iPSCs derived from different tissular origins show essentially the same glycan profiles, which are typified by several characteristic structural features. In addition, a recombinant lectin probe, rBC2LCN, which shows rigorous specificity to H type 1 and 3 glycan structures, is found to serve as an excellent probe for hPSCs. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
    Advanced Healthcare Materials 04/2015; DOI:10.1002/adhm.201400837 · 5.80 Impact Factor
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    ABSTRACT: CEL-I is a galactose/N-acetyl-galactosamine-specific C-type lectin isolated from the sea cucumber Cucumaria echinata. Its carbohydrate-binding site contains a QPD (Gln-Pro-Asp) motif, which is generally recognized as the galactose specificity-determining motif in the C-type lectins. In our previous study, replacement of the QPD motif by an EPN (Glu-Pro-Asn) motif led to a weak binding affinity for mannose. Therefore, we examined the effects of an additional mutation in the carbohydrate-binding site on the specificity of the lectin. Trp105 of EPN-CEL-I was replaced by a histidine residue using site-directed mutagenesis, and the binding affinity of the resulting mutant, EPNH-CEL-I, was examined by sugar-polyamidoamine dendrimer assay, isothermal titration calorimetry, and glycoconjugate microarray analysis. Tertiary structure of the EPNH-CEL-I/mannose complex was determined by X-ray crystallographic analysis. Sugar-polyamidoamine dendrimer assay and glycoconjugate microarray analysis revealed a drastic change in the specificity of EPNH-CEL-I from galactose/N-acetyl-galactosamine to mannose. The association constant of EPNH-CEL-I for mannose was determined to be 3.17 × 10(3) M(-1) at 25°C. Mannose specificity of EPNH-CEL-I was achieved by stabilization of the binding of mannose in a correct orientation, in which the EPN motif can form proper hydrogen bonds with 3- and 4-hydroxy groups of the bound mannose. Specificity of CEL-I can be engineered by mutating a limited number of amino acid residues in addition to the QPD/EPN motifs. Versatility of the C-type carbohydrate-recognition domain structure in the recognition of various carbohydrate chains could become a promising platform to develop novel molecular recognition proteins. Copyright © 2015. Published by Elsevier B.V.
    Biochimica et Biophysica Acta 04/2015; 1850(7). DOI:10.1016/j.bbagen.2015.04.004 · 4.66 Impact Factor
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    ABSTRACT: The application of stem-cell-based therapies in regenerative medicine is hindered by the tumorigenic potential of residual human pluripotent stem cells. Previously, we identified a human pluripotent stem-cell-specific lectin probe, called rBC2LCN, by comprehensive glycome analysis using high-density lectin microarrays. Here we developed a recombinant lectin-toxin fusion protein of rBC2LCN with a catalytic domain of Pseudomonas aeruginosa exotoxin A, termed rBC2LCN-PE23, which could be expressed as a soluble form from the cytoplasm of Escherichia coli and purified to homogeneity by one-step affinity chromatography. rBC2LCN-PE23 bound to human pluripotent stem cells, followed by its internalization, allowing intracellular delivery of a cargo of cytotoxic protein. The addition of rBC2LCN-PE23 to the culture medium was sufficient to completely eliminate human pluripotent stem cells. Thus, rBC2LCN-PE23 has the potential to contribute to the safety of stem-cell-based therapies. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.
    Stem Cell Reports 04/2015; 11(5). DOI:10.1016/j.stemcr.2015.02.016 · 5.37 Impact Factor
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    ABSTRACT: Although various molecular profiling technologies have the potential to predict specific tumor phenotypes, the comprehensive profiling of lectin-bound glycans in human cancer tissues has not yet been achieved. We examined 242 advanced gastric cancer (AGC) patients without or with lymph node metastasis-N0 (n = 62) or N+ (n = 180)-by lectin microarray, and identified the specific lectins highly associated with AGC phenotypes. In seven gastric cancer cell lines, in contrast to expressed-in-cancer lectins, not-expressed-in-cancer (NEC) lectins were tentatively designated by lectin microarray. Binding signals of the specific lectins were robustly reduced in AGC patients with N+ status as compared with those with N0 status. The receiver operating characteristic curve determined the optimal cutoff value to differentiate N0 status from N+ status, and subsequent profiling of NEC lectins identified Vicia villosa agglutinin (VVA) association with the significant other lectins involved in lymph node metastasis. VVA reaction was clearly found on cancer cells, suggesting that it may result from carcinoma-stroma interaction in primary AGC, because VVA is an NEC lectin. Most intriguingly, VVA reaction was remarkably attenuated in the tumor cells of the metastatic lymph nodes, even if it was recognized in primary AGC. In AGC, histological type was strongly associated with soybean agglutinin and Bauhinia purpurea lectin, whereas p53 mutation was the best correlated with Griffonia simplicifolia lectin II. Lectin microarrays can be used to very accurately quantify the reaction of glycans with tumor tissues, and such profiles may represent the specific phenotypes, including N+ status, histological type, or p53 mutation of AGC.
    Gastric Cancer 04/2015; DOI:10.1007/s10120-015-0491-2 · 3.72 Impact Factor
  • Jun Hirabayashi · Atsushi Kuno · Hiroaki Tateno ·
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    ABSTRACT: The lectin microarray is an emerging technology for glycomics. It has already found maximum use in diverse fields of glycobiology by providing simple procedures for differential glycan profiling in a rapid and high-throughput manner. Since its first appearance in the literature in 2005, many application methods have been developed essentially on the same platform, comprising a series of glycan-binding proteins immobilized on an appropriate substrate such as a glass slide. Because the lectin microarray strategy does not require prior liberation of glycans from the core protein in glycoprotein analysis, it should encourage researchers not familiar with glycotechnology to use glycan analysis in future work. This feasibility should provide a broader range of experimental scientists with good opportunities to investigate novel aspects of glycoscience. Applications of the technology include not only basic sciences but also the growing fields of bio-industry. This chapter describes first the essence of glycan profiling and the basic fabrication of the lectin microarray for this purpose. In the latter part the focus is on diverse applications to both structural and functional glycomics, with emphasis on the wide applicability now available with this new technology. Finally, the importance of developing advanced lectin engineering is discussed.
    Topics in current chemistry 03/2015; 367. DOI:10.1007/128_2014_612 · 4.46 Impact Factor
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    ABSTRACT: Studies of β-glucans are often hampered by their structural diversity and complexity, which is problematic because interest in their effects on animal cells has increased in recent years. Herein, we present a comprehensive strategy for structural characterization of branched β-glucans, and as a proof-of-concept study, characterized laminarin and acid-soluble β-gluco-oligosaccharides (<4,000 Da, void volume elute fraction of gel filtration on Bio-gel P-2) from the brown algae, Ecklonia stolonifera. The strategy involves quantitative fluorescence detection-high performance liquid chromatography that enables the characterization of di- and oligosaccharides after acid hydrolysis of the glucan. We found that laminarin is composed of β1–3 (72% in mol) and β1–6 (28%) anomeric bonds, whereas the E. stolonifera glucan is composed of β1–3 (57%) and β1–6 (43%) anomeric bonds. This composition is distinct from that of other brown algae β-glucans, for which the β1–6 bond content is much smaller. We also performed a detailed structural analysis of the 11 major β-gluco-oligosaccharides prepared by mild acid hydrolysis and β1–3-specific laminarinase digestion. All 11 oligosaccharides contained branches joined to the backbone by β1–6 bonds. Five of the oligosaccharides had extended branches; in this regard, the E. stolonifera glucan is unlike other characterized β-glucans. Our strategy should enable structural characterizations of β-branched glucans, for which no practical approach has been available until now.
    Bioactive Carbohydrates and Dietary Fibre 03/2015; 5(2). DOI:10.1016/j.bcdf.2015.03.002
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    ABSTRACT: Among sulfated glycans, little is known about 3'-sulfation because of the lack of useful probes. In the course of molecular engineering of a fungal galectin from Agrocybe cylindracea, we found that a single substitution of Glu86 with alanine resulted in acquisition of specific binding for the 3'-sulfo-Galβ1-4GlcNAc structure. Extensive glyco-technological analysis revealed that this property was obtained in a "loss-of-function" manner. Though this mutant (E86A) had low total affinity, it showed substantial binding to a naturally occurring N-glycan, of which the terminal galactose is 3-sulfated. Moreover, E86A specifically bound to HeLa cells, in which galactose-3-O-sulfotransferases (Gal3ST2 or Gal3ST3) were over-expressed. © The Authors 2015. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.
    Journal of Biochemistry 03/2015; 157(4). DOI:10.1093/jb/mvv023 · 2.58 Impact Factor
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    ABSTRACT: Galectins are a group of animal lectins characterized by their specificity for β-galactosides. Galectin-2 (Gal-2) is predominantly expressed in the gastrointestinal tract. A proteomic analysis identified Gal-2 as a protein that was S-nitrosylated when mouse gastric mucosal lysates were reacted with S-nitrosoglutathione, a physiologically relevant S-nitrosylating agent. In the present study, recombinant mouse (m)Gal-2 was S-nitrosylated using nitrosocysteine (CysNO), which had no effect on the sugar-binding specificity and dimerization capacity of the protein. On the other hand, mGal-2 oxidation by H2O2 resulted in the loss of sugar-binding ability, while S-nitrosylation prevented H2O2-inducted inactivation, presumably by protecting the Cys residue(s) in the protein. These results suggest that S-nitrosylation by nitric oxides protect Gal-2 from oxidative stress in the gastrointestinal tract. Copyright © 2015. Published by Elsevier Inc.
    Biochemical and Biophysical Research Communications 01/2015; 457(4). DOI:10.1016/j.bbrc.2015.01.055 · 2.30 Impact Factor
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    ABSTRACT: Lectins are a large group of carbohydrate-binding proteins, having been shown to comprise at least 48 protein scaffolds or protein family entries. They occur ubiquitously in living organisms-from humans to microorganisms, including viruses-and while their functions are yet to be fully elucidated, their main underlying actions are thought to mediate cell-cell and cell-glycoconjugate interactions, which play important roles in an extensive range of biological processes. The basic feature of each lectin's function resides in its specific sugar-binding properties. In this regard, it is beneficial for researchers to have access to fundamental information about the detailed oligosaccharide specificities of diverse lectins. In this review, the authors describe a publicly available lectin database named "Lectin frontier DataBase (LfDB)", which undertakes the continuous publication and updating of comprehensive data for lectin-standard oligosaccharide interactions in terms of dissociation constants (Kd's). For Kd determination, an advanced system of frontal affinity chromatography (FAC) is used, with which quantitative datasets of interactions between immobilized lectins and >100 fluorescently labeled standard glycans have been generated. The FAC system is unique in its clear principle, simple procedure and high sensitivity, with an increasing number (>67) of associated publications that attest to its reliability. Thus, LfDB, is expected to play an essential role in lectin research, not only in basic but also in applied fields of glycoscience.
    Molecules 01/2015; 20(1):951-973. DOI:10.3390/molecules20010951 · 2.42 Impact Factor
  • Jun Iwaki · Jun Hirabayashi ·
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    ABSTRACT: Frontal affinity chromatography (FAC) is a simple and versatile procedure enabling quantitative determination of diverse biological interactions in terms of dissociation constants (K d), even though these interactions are relatively weak. The method is best applied to glycans and their binding proteins, with the analytical system operating on the basis of highly reproducible isocratic elution by liquid chromatography. Its application to galectins has been successfully developed to characterize their binding specificities in detail. As a result, their minimal requirements for recognition of disaccharides, i.e., β-galactosides, as well as characteristic features of individual galectins, have been elucidated. In this chapter, we describe standard procedures to determine the K d’s for interactions between a series of standard glycans and various galectins.
    Methods in Molecular Biology 01/2015; 1207:63-74. DOI:10.1007/978-1-4939-1396-1_4 · 1.29 Impact Factor
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    ABSTRACT: The circadian clock regulates various behavioral and physiological rhythms in mammals. Circadian changes in olfactory functions such as neuronal firing in the olfactory bulb (OB) and olfactory sensitivity have recently been identified, although the underlying molecular mechanisms remain unknown. We analyzed the temporal profiles of glycan structures in the mouse OB using a high-density microarray that includes 96 lectins, because glycoconjugates play important roles in the nervous system such as neurite outgrowth and synaptogenesis. Sixteen lectin signals significantly fluctuated in the OB, and the intensity of all three that had high affinity for α1-2-fucose (α1-2Fuc) glycan in the microarray was higher during the nighttime. Histochemical analysis revealed that α1-2Fuc glycan is located in a diurnal manner in the lateral olfactory tract that comprises axon bundles of secondary olfactory neurons. The amount of α1-2Fuc glycan associated with the major target glycoprotein neural cell adhesion molecule (NCAM) varied in a diurnal fashion, although the mRNA and protein expression of Ncam1 did not. The mRNA and protein expression of Fut1, a α1-2-specific fucosyltransferase gene, was diurnal in the OB. Daily fluctuation of the α1-2Fuc glycan was obviously damped in homozygous Clock mutant mice with disrupted diurnal Fut1 expression, suggesting that the molecular clock governs rhythmic α1-2-fucosylation in secondary olfactory neurons. These findings suggest the possibility that the molecular clock is involved in the diurnal regulation of olfaction via α1-2-fucosylation in the olfactory system. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.
    Journal of Biological Chemistry 12/2014; 289(52):36158-65. DOI:10.1074/jbc.M114.571141 · 4.57 Impact Factor
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    ABSTRACT: The aim of the present study was to determine the physiological role of skin lectins of the Japanese bullhead shark (Heterodontus japonicus). A skin extract was subjected to affinity chromatography using seven different sugars as ligands. Molecular mass and N-terminal amino acid sequence analyses indicated elution of the same protein by each of the seven respective cognate ligands from sugar affinity columns. The predicted amino acid sequence encoded by the cDNA of this protein (designated HjCL) identified it as a novel fish subgroup VII C-type lectin evolutionarily related to snake venom lectins. HjCL was predicted to bind to mannose because of the presence of a Glu-Pro-Asn (EPN) motif; however, hemagglutination inhibition assays and glycoconjugate microarray analysis demonstrated its binding to numerous structurally diverse sugars. Competitive sugar-binding assays using affinity chromatography indicated that HjCL bound multiple sugars via a common carbohydrate-recognition domain. The mRNA encoding HjCL was specifically detected in the skin, and immunohistochemical analysis detected its expression in uncharacterized large cells in the epidermis. HjCL agglutinated the bacterial pathogen Edwardsiella tarda and promoted immediate clotting of shark blood, indicating that HjCL is involved in host defense on the skin surface especially when the shark is injured and bleeds. © The Authors 2014. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.
    Journal of Biochemistry 11/2014; 157(5). DOI:10.1093/jb/mvu080 · 2.58 Impact Factor
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    ABSTRACT: Two jacalin-related lectins (JRLs) were purified by mannose-agarose and melibiose-agarose from seeds of Treculia africana. One is galactose-recognizing JRL (gJRL), named T. africana agglutinin-G (TAA-G), and another one is mannose-recognizing JRL (mJRL), TAA-M. The yields of the two lectins from the seed flour were approximately 7.0 mg/g for gJRL and 7.2 mg/g for mJRL. The primary structure of TAA-G was determined by protein sequencing of lysyl endopeptic peptides and chymotryptic peptides. The sequence identity of TAA-G to other gJRLs was around 70%. Two-residue insertion was found around the sugar-binding sites, compared with the sequences of other gJRLs. Crystallographic studies on other gJRLs have shown that the primary sugar-binding site of gJRLs can accommodate Gal, GalNAc, and GalNAc residue of T-antigen (Galβ1-3GalNAcα-). However, hemagglutination inhibition and glycan array showed that TAA-G did not recognize GalNAc itself and T-antigen. TAA-G preferred melibiose and core 3 O-glycan.
    Bioscience Biotechnology and Biochemistry 08/2014; 78(12):1-9. DOI:10.1080/09168451.2014.948376 · 1.06 Impact Factor
  • Zui Fujimoto · Hiroaki Tateno · Jun Hirabayashi ·
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    ABSTRACT: Recent progress in structural biology has elucidated the three-dimensional structures and carbohydrate-binding mechanisms of most lectin families. Lectins are classified into 48 families based on their three-dimensional structures. A ribbon drawing gallery of the crystal and solution structures of representative lectins or lectin-like proteins is appended and may help to convey the diversity of lectin families, the similarity and differences between lectin families, as well as the carbohydrate-binding architectures of lectins.
    Methods in Molecular Biology 08/2014; 1200:579-606. DOI:10.1007/978-1-4939-1292-6_46 · 1.29 Impact Factor
  • Yuka Kobayashi · Hiroaki Tateno · Haruko Ogawa · Kazuo Yamamoto · Jun Hirabayashi ·
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    ABSTRACT: More than 100 years have passed since the first lectin ricin was discovered. Since then, a wide variety of lectins (lect means "select" in Latin) have been isolated from plants, animals, fungi, bacteria, as well as viruses, and their structures and properties have been characterized. At present, as many as 48 protein scaffolds have been identified as functional lectins from the viewpoint of three-dimensional structures as described in this chapter. In this chapter, representative 53 lectins are selected, and their major properties that include hemagglutinating activity, mitogen activity, blood group specificity, molecular weight, metal requirement, and sugar specificities are summarized as a comprehensive table. The list will provide a practically useful, comprehensive list for not only experienced lectin users but also many other non-expert researchers, who are not familiar to lectins and, therefore, have no access to advanced lectin biotechnologies described in other chapters.
    Methods in Molecular Biology 08/2014; 1200:555-77. DOI:10.1007/978-1-4939-1292-6_45 · 1.29 Impact Factor
  • Emi Yasuda · Tomoyuki Sako · Hiroaki Tateno · Jun Hirabayashi ·
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    ABSTRACT: Since 2005, lectin microarray technology has emerged as a simple and powerful technique for comprehensive glycan analysis. By using evanescent-field fluorescence detection technique, it has been applied for analysis of not only glycoproteins and glycolipids secreted by eukaryotic cells but also glycoconjugates on the cell surface of live eukaryotic cells. Bacterial cells are known to be decorated with polysaccharides, teichoic acids, and proteins in the peptide glycans of their cell wall and lipoteichoic acids in their phospholipid bilayer. Specific glycan structures are characteristic of many highly pathogenic bacteria, while polysaccharides moiety of lactic acid bacteria are known to play a role as probiotics to modulate the host immune response. However, the method of analysis and knowledge of glycosylation structure of bacteria are limited. Here, we describe the development of a simple and sensitive method based on lectin microarray technology for direct analysis of intact bacterial cell surface glycomes. The method involves labeling bacterial cells with SYTOX Orange before incubation with the lectin microarray. After washing, bound cells are directly detected using an evanescent-field fluorescence scanner in a liquid phase. The entire procedure takes 3 h from putting labeled bacteria on the microarray to profiling its lectin binding affinity. Using this method, we compared the cell surface glycomes from 16 different strains of L. casei/paracasei. The lectin binding profile of most strains was found to be unique. Our technique provides a novel strategy for rapid profiling of bacteria and enables us to differentiate numerous bacterial strains with relevance to the biological functions of surface glycosylation.
    Methods in Molecular Biology 08/2014; 1200:295-311. DOI:10.1007/978-1-4939-1292-6_25 · 1.29 Impact Factor
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    ABSTRACT: There are huge numbers of clinical specimens being stored that contain potential diagnostic marker molecules buried by the coexistence of high-abundance proteins. To utilize such valuable stocks efficiently, we must develop appropriate techniques to verify the molecules. Glycoproteins with disease-related glycosylation changes are a group of useful molecules that have long been recognized, but their application is not fully implemented. The technology for comparative analysis of such glycoproteins in biological specimens has tended to be left behind, which often leads to loss of useful information without it being recognized. In this chapter, we feature antibody-assisted lectin profiling employing antibody-overlay lectin microarray, the most suitable technology for comparative glycoanalysis of a trace amount of glycoproteins contained in biological specimens. We believe that sharing this detailed protocol will accelerate the glycoproteomics-based discovery of glyco-biomarkers that has attracted recent attention; simultaneously, it will increase the value of clinical specimens as a gold mine of information that has yet to be exploited.
    Methods in Molecular Biology 08/2014; 1200:265-85. DOI:10.1007/978-1-4939-1292-6_23 · 1.29 Impact Factor
  • Jun Hirabayashi ·
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    ABSTRACT: Lectin-based glycomics is an emerging, comprehensive technology in the post-genome sciences. The technique utilizes a panel of lectins, which is a group of biomolecules capable of deciphering "glycocodes," with a novel platform represented by a lectin microarray. The method enables multiple glycan-lectin interaction analyses to be made so that differential glycan profiling can be performed in a rapid and sensitive manner. This approach is in clear contrast to another advanced technology, mass spectrometry, which requires prior glycan liberation. Although the lectin microarray cannot provide definitive structures of carbohydrates and their attachment sites, it gives useful clues concerning the characteristic features of glycoconjugates. These include differences not only in terminal modifications (e.g., sialic acid (Sia) linkage, types of fucosylation) but also in higher ordered structures in terms of glycan density, depth, and direction composed for both N- and O-glycans. However, before this technique began to be implemented in earnest, many other low-throughput methods were utilized in the late twentieth century. In this chapter, the author describes how the current lectin microarray technique has developed based on his personal experience.
    Methods in Molecular Biology 08/2014; 1200:225-42. DOI:10.1007/978-1-4939-1292-6_20 · 1.29 Impact Factor

Publication Stats

10k Citations
980.79 Total Impact Points


  • 2005-2015
    • Kagawa University
      • • Life Science Research Center
      • • Division of Glyco-Bioindustry and Functional Glycomics
      Takamatu, Kagawa, Japan
  • 2003-2015
    • National Institute of Advanced Industrial Science and Technology
      • • Research Center for Stem Cell Engineering
      • • Research Center for Medical Glycoscience
      Tsukuba, Ibaraki, Japan
  • 2014
    • Kitasato University
      • Department of Marine Biosciences
      Edo, Tōkyō, Japan
  • 2013
    • Japan Advanced Institute of Science and Technology
      KMQ, Ishikawa, Japan
  • 2011
    • The University of Tokyo
      Tōkyō, Japan
  • 2008
    • Ludwig-Maximilians-University of Munich
      • Faculty of Veterinary Medicine
      München, Bavaria, Germany
  • 1986-2008
    • Teikyo University
      • Faculty of Pharmaceutical Sciences
      Edo, Tōkyō, Japan
  • 2006
    • Advance Institute of Science and Technology
      Dehra, Uttarakhand, India
    • Seikagaku Corporation
      Edo, Tōkyō, Japan
    • Saitama University
      • Faculty of Science
      Saitama, Saitama, Japan
  • 2001
    • Kanazawa Medical University
      • Department of Pathology
      Kanazawa, Ishikawa, Japan
  • 1998
    • Kyorin University
      • Department of Anatomy
      Edo, Tōkyō, Japan