2025/09/29 更新

写真a

カジ ヒロユキ
梶 裕之
KAJI Hiroyuki
所属
糖鎖生命コア研究所 糖鎖ビッグデータセンター 数理解析部門 特任教授
職名
特任教授
外部リンク

学位 1

  1. 理学博士 ( 1989年3月   青山学院大学 ) 

研究キーワード 5

  1. プロテオミクス

  2. グライコプロテオミクス

  3. 質量分析

  4. 翻訳後修飾

  5. 糖鎖修飾

経歴 2

  1. 名古屋大学   糖鎖生命コア研究所 糖鎖ビッグデータセンター 数理解析部門   特任教授

    2022年8月 - 現在

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  2. 名古屋大学   糖鎖生命コア研究所 統合生命医科学糖鎖研究センター数理解析部門   特任教授

    2022年4月 - 2022年7月

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所属学協会 3

  1. 日本糖質学会

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  2. 日本生化学会

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  3. 日本プロテオーム学会

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論文 23

  1. Analytical dissection of minor glycoforms and glycoprotein associations in rAAV preparations by multimodal glycoproteomics 国際誌

    Kuno, A; Sakaue, H; Koizumi, S; Tomioka, A; Mizukado, S; Yamaguchi, Y; Fukuhara, M; Tsunaka, Y; Kaji, H; Uchiyama, S

    ANALYTICAL AND BIOANALYTICAL CHEMISTRY   417 巻 ( 23 ) 頁: 5155 - 5170   2025年9月

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    記述言語:英語   掲載種別:研究論文(学術雑誌)   出版者・発行元:Analytical and Bioanalytical Chemistry  

    Accurate glycan analysis of viral vectors is essential for evaluating pharmaceutical quality. Recent advances in mass spectrometry–based analytical technologies have achieved glycosylation detection in adeno-associated viruses (AAVs). However, because only a minor subpopulation (< 1%) of recombinant AAV (rAAV) particles may carry glycans or associate with glycoproteins, distinguishing genuine AAV glycosylation from that of co-purified glycoproteins remains technically challenging, highlighting the need for analytical strategies that minimize glycan misassignment and reliably identify glycoprotein interactions. Here, we present a multimodal glycoproteomic approach to discriminate rare glycosylation events on rAAV capsids from glycosylated host-derived proteins associated with the particles. We employed an ultrasensitive lectin microarray coupled with a broadly reactive anti-AAV antibody to detect O-glycan-binding lectin signals in several rAAV preparations. Notably, a distinct signal was observed for Urtica dioica agglutinin (UDA). Subsequent liquid chromatography-tandem mass spectrometry, combined with UDA-based dual enrichment at both protein and peptide levels, identified a divalently high-mannose N-glycosylated peptide derived from the host AAV receptor (AAVR). Monovalent high-mannose N-glycopeptides of AAVR and Mac-2 binding protein were additionally detected using single-step protein-level enrichment, indicating an avidity-driven UDA binding mechanism. However, no N-glycosylation was detected on the rAAV capsids themselves. These findings underscore the value of integrated multimodal glycoproteomic workflows for resolving low-abundance glycosylated species and offer new insights into host-derived hitchhiker glycoproteins that may affect rAAV characterization and quality control.

    DOI: 10.1007/s00216-025-06042-4

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  2. Retention of DLK1 in the endoplasmic reticulum identifies roles for EGF-like-domain-specific O-glycans in the secretory pathway 国際誌

    Tashima, Y; Tsukamoto, Y; Tsukamoto, N; Kondo, Y; Uddin, E; Furukawa, W; Go, S; Arakawa, H; Kaji, H; Takeuchi, H; Okajima, T

    FEBS JOURNAL     2025年7月

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    記述言語:英語   掲載種別:研究論文(学術雑誌)   出版者・発行元:FEBS Journal  

    In the endoplasmic reticulum (ER), O-glycosylation by O-fucose, O-glucose, and O-N-acetylglucosamine (O-GlcNAc) occur in the epidermal growth factor-like (EGF) domains of secreted or transmembrane glycoproteins. Previous studies on Notch receptors have revealed the pivotal role of these O-glycans in the cell surface expression of Notch and secretion of truncated Notch fragments. Although it has been demonstrated that O-fucose, O-glucose and O-GlcNAc stabilize individual EGF domains, their role in the secretory pathway following the completion of the folding process remains unexplored. In this study, we used delta-like homolog 1 (DLK1) containing six consecutive EGF domains as a model glycoprotein to investigate the role of EGF-domain-specific O-glycans in the secretory pathway. Semi-quantitative site-specific glycoproteomics of recombinantly expressed DLK1 revealed multiple O-fucose and O-glucose modifications, along with an unusual EGF-domain-specific O-linked N-acetylglucosamine transferase (EOGT)-dependent O-hexose modification. Consistent with the results of the secretion assay, inactivation of the glycosyltransferases modifying O-fucose and O-glucose, but not the newly identified O-hexose, perturbed the transport of DLK1 from the ER during retention, as assessed using the selective hooks (RUSH) system. Importantly, the absence of O-fucose did not affect O-glucose modification within the same EGF domain, and vice versa. Since protein O-fucosyltransferase 1 and protein O-glucosyltransferase 1 activities depend on the folded state of the EGF domains, O-glycans affected DLK1 transport independently of the folding process required for O-glycosylation in the ER. These findings highlight the distinct roles of O-glycans in facilitating the transport of DLK1 from the ER to the cell surface.

    DOI: 10.1111/febs.70168

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  3. Deglycosylation and truncation in the neuraminidase stalk are functionally equivalent in enhancing the pathogenicity of a high pathogenicity avian influenza virus in chickens 国際誌 Open Access

    Kobayashi, D; Hiono, T; Arakawa, H; Kaji, H; Ohkawara, A; Ichikawa, T; Ban, H; Isoda, N; Sakoda, Y

    JOURNAL OF VIROLOGY   99 巻 ( 3 ) 頁: e0147824   2025年3月

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    記述言語:英語   掲載種別:研究論文(学術雑誌)   出版者・発行元:Journal of Virology  

    Influenza A viruses with fewer amino acids in the neuraminidase (NA) stalk domain are primarily isolated from chickens rather than wild ducks, indicating that a shortened NA stalk is considered an adaptation marker of avian influenza viruses (AIVs) to chickens. Experimental passages of an H7N7 nonpathogenic AIV (rgVac2-P0) in chickens resulted in a highly pathogenic variant (Vac2-P3L4) with a 34-amino-acid deletion in the NA stalk, encompassing five potential N-glycosylation sites. To investigate how amino acid truncation and deglycosylation in the NA stalk contribute to increased pathogenicity, a virus with glycosylation-deficient mutations at these sites (rgVac2-P3L4/ P0NAΔGlyco) was constructed. Contrary to expectations, chickens inoculated with rgVac2-P3L4/P0NAΔGlyco exhibited variable clinical outcomes, attributed to the genetic instability of the virus. A single mutation stabilized the virus, and the mutant (rgVac2-P3L4/P0NAΔGlyco-Y65H) resulted in higher pathogenicity compared with a virus with restored glycosylation (rgVac2-P3L4/P0NA-Y65H). Glycan occupancy analysis revealed 3–4 glycans at the five potential sites. In functional analysis, glycosylation-deficient mutants, similar to the short-stalk NA virus, showed significantly reduced erythrocyte elution activity. Additionally, mutational analysis indicated variable contributions of N-glycans to elution activity across the sites. Moreover, the functionally most contributing sites of the five potential N-glycosylation motifs were consistently included in the amino acid deletions of the stalk-truncated NA in N7-subtyped field isolates, despite the varying truncation position or length. These findings suggest that the loss of glycosylation is functionally equivalent to a reduction in amino acids, and it plays a crucial role in enhancing pathogenicity in chickens and affecting NA function.

    DOI: 10.1128/jvi.01478-24

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  4. Inter-tissue glycan heterogeneity: site-specific glycoform analysis of mouse tissue <i>N</i>-glycoproteomes using MS1-based glycopeptide detection method assisted by lectin microarray 国際誌

    Nagai-Okatani, C; Tomioka, A; Tominaga, D; Sakaue, H; Kuno, A; Kaji, H

    ANALYTICAL AND BIOANALYTICAL CHEMISTRY   417 巻 ( 5 ) 頁: 973 - 988   2025年2月

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    記述言語:英語   掲載種別:研究論文(学術雑誌)   出版者・発行元:Analytical and Bioanalytical Chemistry  

    To understand the biological and pathological functions of protein glycosylation, the glycan heterogeneities for each glycosite in a single glycoprotein need to be elucidated depending on the type and status of cells. For this aim, a reliable strategy is needed to compare site-specific glycoforms of multiple glycoprotein samples in a comprehensive manner. To analyze this “inter-heterogeneity” of samples, we previously introduced an MS1-based glycopeptide detection method, “Glyco-RIDGE.” Here, to elucidate inter-tissue glycan heterogeneities, this estimation method was applied to site-specific N-glycoforms of glycoproteins from six normal mouse tissues (liver, kidney, spleen, pancreas, stomach, and testis). The analyses of desialylated glycopeptides estimated 11,325 site-specific N-glycoforms with 239 glycan compositions at 1260 sites (1122 non-redundant core peptides) in 800 glycoproteins, revealing inter-tissue differences in macro-, micro-, and meta-glycan heterogeneities. To obtain detailed information on their glycan features and tissue distribution, the Glyco-RIDGE results were compared with laser microdissection-assisted lectin microarray (LMD-LMA)–based mouse tissue glycome mapping data deposited on LM-GlycomeAtlas, as well as MS-based mouse tissue glycome data deposited on GlycomeAtlas. This integrated approach supported the certainty of Glyco-RIDGE results and suggested that inter-tissue differences exist in glycan motifs, such as alpha-galactose and bisecting N-acetylglucosamine, in both whole tissue glycomes and specific glycoproteins, Anpep and Lamc1. In addition, the comparison with LMD-LMA-based tissue glycome mapping data suggested the possibility of estimating the tissue distribution of the assigned glycans and glycopeptides. Taken together, these findings demonstrate the utility of an integrated approach using MS assisted by LMA for large-scale analyses.

    DOI: 10.1007/s00216-024-05686-y

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  5. Current issues of tandem mass spectrum (MS2)-based glycoproteomics and efforts to complement them 国際誌 Open Access

    Kaji, H

    BBA ADVANCES   7 巻   頁: 100158 - 100158   2025年

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    記述言語:英語   掲載種別:研究論文(学術雑誌)   出版者・発行元:Bba Advances  

    With the development of liquid chromatography/mass spectrometers that support proteomics and the associated development of analytical methods and data analysis software, the number of proteins that can be identified and quantified in a single analysis is approaching the level of transcriptomics. However, many problems remain to be solved in protein glycosylation analysis. This mini-review discusses the technical issues of MS2-based glycoprotein identification and efforts to complement them by the Human Glycome Atlas (HGA) Project in Japan.

    DOI: 10.1016/j.bbadva.2025.100158

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  6. GRable Version 1.0: A Software Tool for Site-Specific fi c Glycoform Analysis With Improved MS1-Based Glycopeptide Detection With Parallel Clustering and Confidence fi dence Evaluation With MS2 Information 国際誌 Open Access

    Nagai-Okatani, C; Tominaga, D; Tomioka, A; Sakaue, H; Goda, N; Ko, SGR; Kuno, A; Kaji, H

    MOLECULAR & CELLULAR PROTEOMICS   23 巻 ( 9 ) 頁: 100833 - 100833   2024年9月

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    記述言語:英語   掲載種別:研究論文(学術雑誌)   出版者・発行元:Molecular and Cellular Proteomics  

    High-throughput intact glycopeptide analysis is crucial for elucidating the physiological and pathological status of the glycans attached to each glycoprotein. Mass spectrometry–based glycoproteomic methods are challenging because of the diversity and heterogeneity of glycan structures. Therefore, we developed an MS1-based site-specific glycoform analysis method named "Glycan heterogeneity-based Relational IDentification of Glycopeptide signals on Elution profile (Glyco-RIDGE)" for a more comprehensive analysis. This method detects glycopeptide signals as a cluster based on the mass and chromatographic properties of glycopeptides and then searches for each combination of core peptides and glycan compositions by matching their mass and retention time differences. Here, we developed a novel browser-based software named GRable for semi-automated Glyco-RIDGE analysis with significant improvements in glycopeptide detection algorithms, including "parallel clustering." This unique function improved the comprehensiveness of glycopeptide detection and allowed the analysis to focus on specific glycan structures, such as pauci-mannose. The other notable improvement is evaluating the "confidence level" of the GRable results, especially using MS2 information. This function facilitated reduced misassignment of the core peptide and glycan composition and improved the interpretation of the results. Additional improved points of the algorithms are "correction function" for accurate monoisotopic peak picking; one-to-one correspondence of clusters and core peptides even for multiply sialylated glycopeptides; and "inter-cluster analysis" function for understanding the reason for detected but unmatched clusters. The significance of these improvements was demonstrated using purified and crude glycoprotein samples, showing that GRable allowed site-specific glycoform analysis of intact sialylated glycoproteins on a large-scale and in-depth. Therefore, this software will help us analyze the status and changes in glycans to obtain biological and clinical insights into protein glycosylation by complementing the comprehensiveness of MS2-based glycoproteomics. GRable can be freely run online using a web browser via the GlyCosmos Portal (https://glycosmos.org/ grable).

    DOI: 10.1016/j.mcpro.2024.100833

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  7. The Human Glycome Atlas Project for cataloging all glycan-related omics data in human. Open Access

    Aoki-Kinoshita KF, Ando H, Angata K, Fujita M, Furukawa JI, Kaji H, Kato K, Kitajima K, Kizuka Y, Matsui Y, Nakajima K, Nishihara S, Okajima T, Sakamoto K, Sato C, Thaysen-Andersen M, Togayachi A, Yagi H, Kadomatsu K

    Glycobiology     2024年7月

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    記述言語:英語  

    DOI: 10.1093/glycob/cwae052

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  8. O-glycosylation of SARS-CoV-2 spike protein by host O-glycosyltransferase strengthens its trimeric structure Open Access

    Zhijue Xu, Han Zhang, Jiaqi Tian, Xin Ku, Rumeng Wei, Jingli Hou, Can Zhang, Fang Yang, Xia Zou, Yang Li, Hiroyuki Kaji, Sheng Ce Tao, Atsushi Kuno, Wei Yan, Lin Tai Da, Yan Zhang

    Acta biochimica et biophysica Sinica   56 巻 ( 8 ) 頁: 1118 - 1129   2024年7月

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    掲載種別:研究論文(学術雑誌)  

    DOI: 10.3724/abbs.2024127

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  9. Glycosylation of recombinant adeno-associated virus 6 国際誌 Open Access

    Yamaguchi, Y; Ishii, K; Koizumi, S; Sakaue, H; Maruno, T; Fukuhara, M; Shibuya, R; Tsunaka, Y; Matsushita, A; Bandoh, K; Torisu, T; Murata-Kishimoto, C; Tomioka, A; Mizukado, S; Kaji, H; Kashiwakura, Y; Ohmori, T; Kuno, A; Uchiyama, S

    MOLECULAR THERAPY METHODS & CLINICAL DEVELOPMENT   32 巻 ( 2 ) 頁: 101256 - 101256   2024年6月

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    記述言語:英語   掲載種別:研究論文(学術雑誌)   出版者・発行元:Molecular Therapy Methods and Clinical Development  

    Glycosylation of biopharmaceuticals can affect their safety and efficacy. Glycans can occur on recombinant adeno-associated viruses (rAAVs) that are used for gene therapy; however, the types of glycans that attach to rAAVs are controversial. Here, we conducted lectin microarray analyses on six rAAV serotype 6 (rAAV6) preparations that were produced differently. We demonstrate that O-glycans considered to be attached to rAAV6 were recognized by Agaricus bisporus agglutinin (ABA) and that N-glycans were detected in rAAV6 purified without affinity chromatography. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis showed that the N-glycans detected in rAAV6 were derived from host cell proteins. A combination of ABA-based fractionation and LC-MS/MS revealed that rAAV6 was O-glycosylated with the mucin-type glycans, O-GalNAc (Tn antigen), and mono- and di-sialylated Galβ1-3GalNAc (T antigen) at S156, T162, T194, and T201 in viral protein (VP) 2 and with O-GlcNAc at T242 in VP3. The mucin-type O-glycosylated rAAV6 particles were 0.1%–1% of total particles. Further physicochemical and biological analyses revealed that mucin-type O-glycosylated rAAV6 had a lower ratio of VP1 to VP2/VP3, resulting in a lower transduction efficiency both in vitro and in vivo compared with rAAV6 without mucin-type O-glycans. This report details conclusive evidence of rAAV glycosylation and its impact on rAAV-based therapeutics.

    DOI: 10.1016/j.omtm.2024.101256

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  10. Mucin-Type O-Glycosylation in Viral Protein 2 Indirectly Affects the Transduction Efficiency of Recombinant Adeno-Associated Virus Serotype 6

    Yamaguchi, Y; Ishii, K; Koizumi, S; Sakaue, H; Maruno, T; Fukuhara, M; Shibuya, R; Tsunaka, Y; Matsushita, A; Bandoh, K; Torisu, T; Murata-Kishimoto, C; Tomioka, A; Mizukado, S; Kaji, H; Kashiwakura, Y; Ohmori, T; Kuno, A; Uchiyama, S

    MOLECULAR THERAPY   32 巻 ( 4 ) 頁: 688 - 689   2024年4月

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  11. Combinatorial Approach with Mass Spectrometry and Lectin Microarray Dissected Site-Specific Glycostem and Glycoleaf Features of the Virion-Derived Spike Protein of Ancestral and γ Variant SARS-CoV-2 Strains. 国際誌

    Takahiro Hiono, Hiroaki Sakaue, Azusa Tomioka, Hiroyuki Kaji, Michihito Sasaki, Yasuko Orba, Hirofumi Sawa, Atsushi Kuno

    Journal of proteome research   23 巻 ( 4 ) 頁: 1408 - 1419   2024年4月

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    記述言語:英語   掲載種別:研究論文(学術雑誌)  

    The coronavirus disease (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has impacted public health globally. As the glycosylation of viral envelope glycoproteins is strongly associated with their immunogenicity, intensive studies have been conducted on the glycans of the glycoprotein of SARS-CoV-2, the spike (S) protein. Here, we conducted intensive glycoproteomic analyses of the SARS-CoV-2 S protein of ancestral and γ-variant strains using a combinatorial approach with two different technologies: mass spectrometry (MS) and lectin microarrays (LMA). Our unique MS1-based glycoproteomic technique, Glyco-RIDGE, in addition to MS2-based Byonic search, identified 1448 (ancestral strain) and 1785 (γ-variant strain) site-specific glycan compositions, respectively. Asparagine at amino acid position 20 (N20) is mainly glycosylated within two successive potential glycosylation sites, N17 and N20, of the γ-variant S protein; however, we found low-frequency glycosylation at N17. Our novel approaches, glycostem mapping and glycoleaf scoring, also illustrate the moderately branched/extended, highly fucosylated, and less sialylated natures of the glycoforms of S proteins. Subsequent LMA analysis emphasized the intensive end-capping of glycans by Lewis fucoses, which complemented the glycoproteomic features. These results illustrate the high-resolution glycoproteomic features of the SARS-CoV-2 S protein, contributing to vaccine design and understanding of viral protein synthesis.

    DOI: 10.1021/acs.jproteome.3c00874

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  12. HGA Segment 1: Comprehensive acquisition of human glycan structural information

    Kaji Hiroyuki

    Glycoforum   27 巻 ( 2 ) 頁: A4   2024年4月

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    記述言語:英語   出版者・発行元:SEIKAGAKU CORPORATION  

    DOI: 10.32285/glycoforum.27a4

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  13. HGAセグメント1:ヒト糖鎖構造情報の網羅的取得

    梶 裕之

      27 巻 ( 2 ) 頁: A4J   2024年4月

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    記述言語:日本語   出版者・発行元:生化学工業株式会社  

    DOI: 10.32285/glycoforum.27a4j

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  14. Tamoxifen-resistant breast cancer cells exhibit reactivity with Wisteria floribunda agglutinin. 国際誌 Open Access

    Hlaing MT, Horimoto Y, Denda-Nagai K, Fujihira H, Noji M, Kaji H, Tomioka A, Ishizuka Y, Saeki H, Arakawa A, Saito M, Irimura T

    PloS one   17 巻 ( 8 ) 頁: e0273513 - e0273513   2022年8月

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    記述言語:英語   掲載種別:研究論文(学術雑誌)  

    DOI: 10.1371/journal.pone.0273513

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  15. Lewis glycosphingolipids as critical determinants of TRAIL sensitivity in cancer cells.

    Fukuoka T, Moriwaki K, Takamatsu S, Kondo J, Tanaka-Okamoto M, Tomioka A, Semba M, Komazawa-Sakon S, Kamada Y, Kaji H, Miyamoto Y, Inoue M, Bessho K, Miyoshi Y, Ozono K, Nakano H, Miyoshi E

    Oncogene   41 巻 ( 38 ) 頁: 4385 - 4396   2022年8月

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    記述言語:英語   掲載種別:研究論文(学術雑誌)  

    DOI: 10.1038/s41388-022-02434-3

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    その他リンク: https://www.nature.com/articles/s41388-022-02434-3

  16. A Novel Method of CD31-Combined ABO Carbohydrate Antigen Microarray Predicts Acute Antibody-Mediated Rejection in ABO-Incompatible Kidney Transplantation 国際誌 Open Access

    Tasaki M., Tateno H., Sato T., Tomioka A., Kaji H., Narimatsu H., Saito K., Nakagawa Y., Aoki T., Kamimura M., Ushiki T., Okada M., Miwa Y., Hotta K., Yoshida Y., Takahashi K., Tomita Y.

    Transplant International   35 巻   頁: 10248 - 10248   2022年3月

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    記述言語:英語   掲載種別:研究論文(学術雑誌)   出版者・発行元:Transplant International  

    Isohemagglutinin assays employing red blood cells (RBCs) are the most common assays used to measure antibody titer in ABO-incompatible kidney transplantation (ABOi KTx). However, ABO antigens expressed on RBCs are not identical to those of kidney and antibody titers do not always correlate with clinical outcome. We previously reported that CD31 was the main protein linked to ABO antigens on kidney endothelial cells (KECs), which was different from those on RBCs. We developed a new method to measure antibody titer using a microarray of recombinant CD31 (rCD31) linked to ABO antigens (CD31-ABO microarray). Mass spectrometry analysis suggested that rCD31 and native CD31 purified from human kidney had similar ABO glycan. To confirm clinical use of CD31-ABO microarray, a total of 252 plasma samples including volunteers, hemodialysis patients, and transplant recipients were examined. In transplant recipients, any initial IgG or IgM antibody intensity >30,000 against the donor blood type in the CD31-ABO microarray showed higher sensitivity, specificity, positive predictive value, and negative predictive value of AABMR, compared to isohemagglutinin assays. Use of a CD31-ABO microarray to determine antibody titer specifically against ABO antigens expressed on KECs will contribute to precisely predicting AABMR or preventing over immunosuppression following ABOi KTx.

    DOI: 10.3389/ti.2022.10248

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  17. Sensitive New Assay System for Serum <i>Wisteria floribunda</i> Agglutinin-Reactive Ceruloplasmin That Distinguishes Ovarian Clear Cell Carcinoma from Endometrioma. 国際誌

    Sogabe M, Kojima S, Kaya T, Tomioka A, Kaji H, Sato T, Chiba Y, Shimizu A, Tanaka N, Suzuki N, Hayashi I, Mikami M, Togayachi A, Narimatsu H

    Analytical chemistry   94 巻 ( 5 ) 頁: 2476 - 2484   2022年2月

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    記述言語:英語   掲載種別:研究論文(学術雑誌)  

    DOI: 10.1021/acs.analchem.1c04302

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  18. O-glycosylated HBsAg peptide can induce specific antibody neutralizing HBV infection 国際誌 Open Access

    Angata K., Wagatsuma T., Togayachi A., Sato T., Sogabe M., Tajiri K., Ozawa T., Nagashima I., Shimizu H., Iijima S., Korenaga M., Kuno A., Kaji H., Mizokami M., Narimatsu H.

    Biochimica et Biophysica Acta - General Subjects   1866 巻 ( 1 ) 頁: 130020 - 130020   2022年1月

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    記述言語:英語   掲載種別:研究論文(学術雑誌)   出版者・発行元:Biochimica et Biophysica Acta - General Subjects  

    Background: Hepatitis B virus (HBV), which causes hepatitis, liver cirrhosis, and hepatocellular carcinoma, is a global human health problem. HBV contains three envelope proteins, S-, M-, and L-hepatitis B surface antigen (HBsAg). We recently found that O-glycosylated M-HBsAg, reactive with jacalin lectin, is one of the primary components of HBV DNA-containing virus particles. Thus, we aimed to analyze and target the glycosylation of HBsAg. Methods: HBsAg prepared from the serum of Japanese patients with HBV were analyzed using mass spectrometry. The glycopeptide modified with O-glycan was generated and used for immunization. The specificity of the generated antibody and the HBV infection inhibition activity was examined. Results: Mass spectrometry analysis revealed that T37 and/or T38 on M-HBsAg of genotype C were modulated by ±NeuAc(α2,3)Gal(β1,3)GalNAc. Chemically and enzymatically synthesized O-glycosylated peptide (Glyco-PS2) induced antibodies that recognize mainly PreS2 in M-HBsAg not in L-HBsAg, whereas the non-glycosylated peptide (PS2) induced antisera recognizing L-HBsAg but not O-glycosylated M-HBsAg. The removal of O-glycan from M-HBsAg partly decreased the reactivity of the Glyco-PS2 antibody, suggesting that peptide part was also recognized by the antibody. The antibody further demonstrated the inhibition of HBV infection in human hepatic cells in vitro. Conclusions: Glycosylation of HBsAg occurs differently in different HBsAgs in a site-specific manner. The new Glyco-PS2 antibody, recognizing O-glycosylated M-HBsAg of genotype C, could inhibit HBV infection. General significance: The detailed analysis of HBsAg identified different glycosylations of HBV surface. The glycosylated peptide based on mass spectrometry analysis showed higher potential to induce functional antibody against HBV.

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  19. Host-Dependent Producibility of Recombinant <i>Cypridina noctiluca</i> Luciferase With Glycosylation Defects. 国際誌 Open Access

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  20. Identification of intact glycopeptides by liquid chromatography/tandem mass spectrometry followed by database search.

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MISC 3

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