Advertisement

Conjugation of Synthetic Oligosaccharides to Proteins by Squaric Acid Chemistry

  • Hélène B. Pfister
  • Xiaowei Lu
  • Sameh E. Soliman
  • Pavol KováčEmail author
Protocol
Part of the Methods in Molecular Biology book series (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.

Key words

Oligosaccharides Neoglycoconjugates Squaric acid chemistry Conjugation Vaccines 

References

  1. 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 124:1215–1221CrossRefGoogle Scholar
  2. 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 83–115CrossRefGoogle Scholar
  3. 3.
    Wurm FR, Klok H-A (2013) Be squared: expanding the horizon of squaric acid-mediated conjugations. Chem Soc Rev 42:8220–8236CrossRefGoogle Scholar
  4. 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 53:14245–14249CrossRefGoogle Scholar
  5. 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 1:487–496CrossRefGoogle Scholar
  6. 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:e0003881CrossRefGoogle Scholar
  7. 7.
    Xu P, Kelly M, Vann WF et al (2017) Conjugate vaccines from bacterial antigens by squaric acid chemistry: A closer look. Chembiochem 18:799–815CrossRefGoogle Scholar
  8. 8.
    Hou S-J, Saksena R, Kováč P (2008) Preparation of glycoconjugates by dialkyl squarate chemistry revisited. Carbohydr Res 343:196–210CrossRefGoogle Scholar
  9. 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 456:24–29CrossRefGoogle Scholar
  10. 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 338:2591–2603CrossRefGoogle Scholar
  11. 11.
    Soliman SE, Kováč P (2016) Total synthesis of the complete protective antigen of Vibrio cholerae O139. Angew Chem Int Ed 55:12850–12853CrossRefGoogle Scholar
  12. 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 247:402–410CrossRefGoogle Scholar
  13. 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 238:160–165CrossRefGoogle Scholar
  14. 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 330:479–486CrossRefGoogle Scholar
  15. 15.
    Pluskal MG (2000) Microscale sample preparation. Nat Biotechnol 18:104CrossRefGoogle Scholar
  16. 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 141–150CrossRefGoogle Scholar

Copyright information

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

Authors and Affiliations

  • Hélène B. Pfister
    • 1
  • Xiaowei Lu
    • 1
  • Sameh E. Soliman
    • 1
  • Pavol Kováč
    • 2
    Email author
  1. 1.National Institutes of HealthNIDDK, LBCBethesdaUSA
  2. 2.Laboratory of Bioorganic ChemistryNational Institutes of HealthBethesdaUSA

Personalised recommendations