Skip to main content
SpringerLink
Log in

Conjugation of Synthetic Oligosaccharides to Proteins by Squaric Acid Chemistry

  • Protocol
  • First Online:
Bacterial Polysaccharides

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1954))

Abstract

Oligosaccharides equipped with amine-containing linkers can be conjugated to carrier proteins using squaric acid chemistry. In a two-step process, a squarate derivative of such oligosaccharide is formed first, which is followed by its reaction with a protein carrier. Monitoring of the conjugation reaction is achieved by SELDI-TOF-MS or MALDI-TOF-MS. This experimentally simple procedure yields desired glycoconjugates in high yields and with reproducible hapten–protein ratios.

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

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 169.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Hardcover Book
USD 219.99
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

References

  1. Tietze LF, Arlt M, Beller M et al (1991) Anticancer agents, 15. Squaric acid diethyl ester: A new coupling reagent for the formation of drug biopolymer conjugates. Synthesis of squaric acid ester amides and diamides. Chem Ber 124:1215–1221

    Article  CAS  Google Scholar 

  2. Kováč P, Xu P (2017) Controlled and highly efficient preparation of carbohydrate-based vaccines: squaric acid chemistry is the way to go. In: Pilar Rautner A, Lindhorst T, Queneau Y (eds) Carbohydrate chemistry, The royal society of chemistry, vol 42, pp 83–115

    Chapter  Google Scholar 

  3. Wurm FR, Klok H-A (2013) Be squared: expanding the horizon of squaric acid-mediated conjugations. Chem Soc Rev 42:8220–8236

    Article  CAS  Google Scholar 

  4. Palitzsch B, Hartmann S, Stergiou N et al (2014) A fully synthetic four-component antitumor vaccine consisting of a mucin glycopeptide antigen combined with three different T-helper-cell epitopes. Angew Chem Int Ed 53:14245–14249

    Article  CAS  Google Scholar 

  5. Fallarini S, Buzzi B, Giovarruscio S et al (2015) A synthetic disaccharide analogue from Neisseria meningitidis A capsular polysaccharide stimulates immune cell responses and induces immunoglobulin G (IgG) production in mice when protein-conjugated. ACS Infect Dis 1:487–496

    Article  CAS  Google Scholar 

  6. Sayeed MA, Bufano MK, Xu P et al (2015) A cholera conjugate vaccine containing O-specific polysaccharide (OSP) of V. cholerae O1 Inaba and recombinant fragment of tetanus toxin Heavy Chain (OSP:rTTHc) induces serum, memory and lamina proprial responses against OSP and is protective in mice. PLoS Neglected Trop Dis 9:e0003881

    Article  Google Scholar 

  7. Xu P, Kelly M, Vann WF et al (2017) Conjugate vaccines from bacterial antigens by squaric acid chemistry: A closer look. Chembiochem 18:799–815

    Article  CAS  Google Scholar 

  8. Hou S-J, Saksena R, Kováč P (2008) Preparation of glycoconjugates by dialkyl squarate chemistry revisited. Carbohydr Res 343:196–210

    Article  CAS  Google Scholar 

  9. Xu P, Trinh MN, Kováč P (2018) Conjugation of carbohydrates to proteins using di(triethylene glycol monomethyl ether) squaric acid ester revisited. Carbohydr Res 456:24–29

    Article  CAS  Google Scholar 

  10. Saksena R, Ma X, Kováč P (2003) One-pot preparation of a series of glycoconjugates with predetermined antigen–carrier ratio from oligosaccharides that mimic the O-PS of Vibrio cholerae O:1, serotype Ogawa. Carbohydr Res 338:2591–2603

    Article  CAS  Google Scholar 

  11. Soliman SE, Kováč P (2016) Total synthesis of the complete protective antigen of Vibrio cholerae O139. Angew Chem Int Ed 55:12850–12853

    Article  CAS  Google Scholar 

  12. Knirel YA, Widmalm G, Senchenkova SN et al (1997) Structural studies on the short-chain lipopolysaccharide of Vibrio cholerae O139 Bengal. Eur J Biochem 247:402–410

    Article  CAS  Google Scholar 

  13. Knirel YA, Senchenkova SN, Jansson P-E et al (1996) Structure of the O-specific polysaccharide of an Aeromonas Trota strain cross-reactive with Vibrio cholerae O139 Bengal. Eur J Biochem 238:160–165

    Article  CAS  Google Scholar 

  14. Chernyak A, Karavanov A, Ogawa Y et al (2001) Conjugating oligosaccharides to proteins by squaric acid diester chemistry: Rapid monitoring of the progress of conjugation, and recovery of the unused ligand. Carbohydr Res 330:479–486

    Article  CAS  Google Scholar 

  15. Pluskal MG (2000) Microscale sample preparation. Nat Biotechnol 18:104

    Article  CAS  Google Scholar 

  16. Zhang Y, Sinaiko AR, Nelsestuen GL (2012) Glycoproteins and glycosylation: apolipoprotein C3 glycoforms by top-down MALDI-TOF mass spectrometry. In: Josic D, Hixson DC (eds) Liver proteomics: methods and protocols, Methods in molecular biology, vol 909. Springer Science, New York, pp 141–150

    Chapter  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. National Institutes of Health, NIDDK, LBC, Bethesda, MD, USA

    Hélène B. Pfister, Xiaowei Lu & Sameh E. Soliman

  2. Laboratory of Bioorganic Chemistry, National Institutes of Health, Bethesda, MD, USA

    Pavol Kováč

Authors
  1. Hélène B. Pfister
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Xiaowei Lu
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Sameh E. Soliman
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Pavol Kováč
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Pavol Kováč .

Editor information

Editors and Affiliations

  1. Department of Biomedical and Molecular Sciences, Queen’s University, Kingston, ON, Canada

    Inka Brockhausen

Rights and permissions

Reprints and permissions

Copyright information

© 2019 Springer Science+Business Media, LLC, part of Springer Nature

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

Pfister, H.B., Lu, X., Soliman, S.E., Kováč, P. (2019). Conjugation of Synthetic Oligosaccharides to Proteins by Squaric Acid Chemistry. In: Brockhausen, I. (eds) Bacterial Polysaccharides. Methods in Molecular Biology, vol 1954. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-9154-9_7

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-9154-9_7

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-9153-2

  • Online ISBN: 978-1-4939-9154-9

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation