Skip to main content

Evaluation of Glycan-Binding Specificity by Glycoconjugate Microarray with an Evanescent-Field Fluorescence Detection System

  • Protocol
  • First Online:
Lectins

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1200))

Abstract

The glycan microarray is now an essential tool used to study lectins. With this technique, glycan-binding specificity can be easily assessed by incubation with an array immobilizing a series of glycans. Glycan microarrays have been developed by numerous research groups around the world. Among the available microarrays, our glycan microarray has two unique characteristics: one is the incorporation of an evanescent-field fluorescence detection system and the other is the use of multivalent glycopolymers. These two unique properties allow the highly sensitive detection of only nanogram quantities of lectins even in crude samples such as cell lysates and cell culture media. Thus, this system is suitable for the initial screening of lectins, lectin-like molecules, lectin candidates, and lectin mutants. Here I describe the protocols employed to analyze the glycan-binding specificity of lectins using our glycan microarray system.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Subscribe and save

Springer+ Basic
$34.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or eBook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 139.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

Similar content being viewed by others

References

  1. Boyd WC, Shapleigh E (1954) Specific precipitating activity of plant agglutinins (lectins). Science 119:419

    Article  PubMed  CAS  Google Scholar 

  2. Blixt O, Collins BE, van den Nieuwenhof IM, Crocker PR, Paulson JC (2003) Sialoside specificity of the siglec family assessed using novel multivalent probes: identification of potent inhibitors of myelin-associated glycoprotein. J Biol Chem 278:31007–31019

    Article  PubMed  CAS  Google Scholar 

  3. Mega T, Hase S (1991) Determination of lectin-sugar binding constants by microequilibrium dialysis coupled with high performance liquid chromatography. J Biochem 109:600–603

    PubMed  CAS  Google Scholar 

  4. Shinohara Y, Kim F, Shimizu M, Goto M, Tosu M, Hasegawa Y (1994) Kinetic measurement of the interaction between an oligosaccharide and lectins by a biosensor based on surface plasmon resonance. Eur J Biochem 223:189–194

    Article  PubMed  CAS  Google Scholar 

  5. Dam TK, Gerken TA, Cavada BS, Nascimento KS, Moura TR, Brewer CF (2007) Binding studies of alpha-GalNAc-specific lectins to the alpha-GalNAc (Tn-antigen) form of porcine submaxillary mucin and its smaller fragments. J Biol Chem 282:28256–28263

    Article  PubMed  CAS  Google Scholar 

  6. Tateno H, Nakamura-Tsuruta S, Hirabayashi J (2007) Frontal affinity chromatography: sugar-protein interactions. Nat Protoc 2:2529–2537

    Article  PubMed  CAS  Google Scholar 

  7. Blixt O, Head S, Mondala T, Scanlan C, Huflejt ME, Alvarez R, Bryan MC, Fazio F, Calarese D, Stevens J, Razi N, Stevens DJ, Skehel JJ, van Die I, Burton DR, Wilson IA, Cummings R, Bovin N, Wong CH, Paulson JC (2004) Printed covalent glycan array for ligand profiling of diverse glycan binding proteins. Proc Natl Acad Sci U S A 101:17033–17038

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  8. Paulson JC, Blixt O, Collins BE (2006) Sweet spots in functional glycomics. Nat Chem Biol 2:238–248

    Article  PubMed  CAS  Google Scholar 

  9. Tateno H, Mori A, Uchiyama N, Yabe R, Iwaki J, Shikanai T, Angata T, Narimatsu H, Hirabayashi J (2008) Glycoconjugate microarray based on an evanescent-field fluorescence-assisted detection principle for investigation of glycan-binding proteins. Glycobiology 18:789–798

    Article  PubMed  CAS  Google Scholar 

  10. Hu D, Tateno H, Sato T, Narimatsu H, Hirabayashi J (2013) Tailoring GalNAcalpha1-3Galbeta-specific lectins from a multi-specific fungal galectin: dramatic change of carbohydrate specificity by a single amino-acid substitution. Biochem J 453:261–270

    Article  PubMed  CAS  Google Scholar 

  11. Shimokawa M, Fukudome A, Yamashita R, Minami Y, Yagi F, Tateno H, Hirabayashi J (2012) Characterization and cloning of GNA-like lectin from the mushroom Marasmius oreades. Glycoconj J 29:457–465

    Article  PubMed  CAS  Google Scholar 

  12. Hu D, Tateno H, Kuno A, Yabe R, Hirabayashi J (2012) Directed evolution of lectins with sugar-binding specificity for 6-sulfo-galactose. J Biol Chem 287:20313–20320

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  13. Takahara K, Arita T, Tokieda S, Shibata N, Okawa Y, Tateno H, Hirabayashi J, Inaba K (2012) Difference in fine specificity to polysaccharides of C. albicans mannoprotein between mouse SIGNR1 and human DC-SIGN. Infect Immun 80:1699–1706

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  14. Tateno H, Toyoda M, Saito S, Onuma Y, Ito Y, Hiemori K, Fukumura M, Nakasu A, Nakanishi M, Ohnuma K, Akutsu H, Umezawa A, Horimoto K, Hirabayashi J, Asashima M (2011) Glycome diagnosis of human induced pluripotent stem cells using lectin microarray. J Biol Chem 286:20345–20353

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  15. Yabe R, Tateno H, Hirabayashi J (2010) Frontal affinity chromatography analysis of constructs of DC-SIGN, DC-SIGNR and LSECtin extend evidence for affinity to agalactosylated N-glycans. FEBS J 277:4010–4026

    Article  PubMed  CAS  Google Scholar 

  16. Tateno H, Ohnishi K, Yabe R, Hayatsu N, Sato T, Takeya M, Narimatsu H, Hirabayashi J (2010) Dual specificity of Langerin to sulfated and mannosylated glycans via a single C-type carbohydrate recognition domain. J Biol Chem 285:6390–6400

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  17. Mitsunaga K, Harada-Itadani J, Shikanai T, Tateno H, Ikehara Y, Hirabayashi J, Narimatsu H, Angata T (2009) Human C21orf63 is a heparin-binding protein. J Biochem 146:369–373

    Article  PubMed  CAS  Google Scholar 

  18. Yamasaki S, Matsumoto M, Takeuchi O, Matsuzawa T, Ishikawa E, Sakuma M, Tateno H, Uno J, Hirabayashi J, Mikami Y, Takeda K, Akira S, Saito T (2009) C-type lectin Mincle is an activating receptor for pathogenic fungus, Malassezia. Proc Natl Acad Sci U S A 106:1897–1902

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  19. Takeuchi T, Sennari R, Sugiura K, Tateno H, Hirabayashi J, Kasai K (2008) A C-type lectin of Caenorhabditis elegans: its sugar-binding property revealed by glycoconjugate microarray analysis. Glycobiology 377:303–306

    CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Hiroaki Tateno .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2014 Springer Science+Business Media New York

About this protocol

Cite this protocol

Tateno, H. (2014). Evaluation of Glycan-Binding Specificity by Glycoconjugate Microarray with an Evanescent-Field Fluorescence Detection System. In: Hirabayashi, J. (eds) Lectins. Methods in Molecular Biology, vol 1200. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1292-6_30

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-1292-6_30

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-1291-9

  • Online ISBN: 978-1-4939-1292-6

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics