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Surprising leads for a cholera toxin receptor-binding antagonist: crystallographic studies of CTB mutants.

Merritt EA, Sarfaty S, Chang TT, Palmer LM, Jobling MG, Holmes RK, Hol WG
Structure 3 561-70 (1995)
Cited: 20 times
EuropePMC logo PMID: 8590017

Abstract

Background

Because agents which inhibit the receptor binding of cholera toxin constitute possible lead compounds for the structure-based design of anti-cholera drugs, detailed investigation of the toxin's receptor-binding site is of key importance. The substitution Gly-->Asp at residue 33 of the cholera toxin B subunit (CTB) has been reported to abolish receptor-binding ability. The substitution Arg35-->Asp has been reported to result in deficient assembly of the AB5 holotoxin. The molecular basis for these effects was not readily apparent from analysis of an earlier crystal structure of the wild-type toxin B pentamer in a complex with the receptor pentasaccharide.

Results

We now report at a resolution of 2.0 A the crystal structure of a recombinant CTB pentamer containing the Gly33-->Asp substitution. The observed conformation of the Asp33 side chain suggests that the loss in binding affinity is due to a steric clash with atoms C9 and O9 of the sialic acid moiety of the receptor, ganglioside GM1. The crystal structure also reveals an unexpected mode of pentamer-pentamer interaction in which pairs of toxin pentamers are joined by reciprocal insertion of the imidazole ring of His13 from one subunit of each pentamer into one of the receptor-binding sites on the other. The surface of interaction at each pentamer-pentamer interface is on the order of 500 A2, and primarily involves contact of residues 10-14 with the receptor-binding site on the associated pentamer. This same pentamer-pentamer interaction is also present in the crystal structure of a second recombinant CTB containing an Arg-->Asp substitution at residue 35, which we have determined at 2.1 A resolution.

Conclusion

These structures suggest that analogs to all or part of the pentapeptide Ala-Glu-Tyr-His-Asn, corresponding to residues 10-14 of CTB, may constitute lead compounds for the design of binding-site inhibitors.

Reviews citing this publication (4)

  1. Cholera toxin - a foe & a friend. Sanchez J, Holmgren J. Indian J Med Res 133 153-163 (2011)
  2. AB toxins: a paradigm switch from deadly to desirable. Odumosu O, Nicholas D, Yano H, Langridge W. Toxins (Basel) 2 1612-1645 (2010)
  3. Overview of protein structural and functional folds. Sun PD, Foster CE, Boyington JC. Curr Protoc Protein Sci Chapter 17 Unit 17.1 (2004)
  4. Structural bioinformatic analysis of DsbA proteins and their pathogenicity associated substrates. Santos-Martin C, Wang G, Subedi P, Hor L, Totsika M, Paxman JJ, Heras B. Comput Struct Biotechnol J 19 4725-4737 (2021)

Articles citing this publication (16)

  1. Structural studies of receptor binding by cholera toxin mutants. Merritt EA, Sarfaty S, Jobling MG, Chang T, Holmes RK, Hirst TR, Hol WG. Protein Sci 6 1516-1528 (1997)
  2. Designing an efficient multi-epitope peptide vaccine against Vibrio cholerae via combined immunoinformatics and protein interaction based approaches. Nezafat N, Karimi Z, Eslami M, Mohkam M, Zandian S, Ghasemi Y. Comput Biol Chem 62 82-95 (2016)
  3. Barrel structures in proteins: automatic identification and classification including a sequence analysis of TIM barrels. Nagano N, Hutchinson EG, Thornton JM. Protein Sci 8 2072-2084 (1999)
  4. Structural foundation for the design of receptor antagonists targeting Escherichia coli heat-labile enterotoxin. Merritt EA, Sarfaty S, Feil IK, Hol WG. Structure 5 1485-1499 (1997)
  5. Construction and characterization of versatile cloning vectors for efficient delivery of native foreign proteins to the periplasm of Escherichia coli. Jobling MG, Palmer LM, Erbe JL, Holmes RK. Plasmid 38 158-173 (1997)
  6. Mutational analysis of ganglioside GM(1)-binding ability, pentamer formation, and epitopes of cholera toxin B (CTB) subunits and CTB/heat-labile enterotoxin B subunit chimeras. Jobling MG, Holmes RK. Infect Immun 70 1260-1271 (2002)
  7. Modeling oligomers with Cn or Dn symmetry: application to CAPRI target 10. Berchanski A, Segal D, Eisenstein M. Proteins 60 202-206 (2005)
  8. A single native ganglioside GM1-binding site is sufficient for cholera toxin to bind to cells and complete the intoxication pathway. Jobling MG, Yang Z, Kam WR, Lencer WI, Holmes RK. mBio 3 (2012)
  9. GM1 ganglioside-independent intoxication by Cholera toxin. Cervin J, Wands AM, Casselbrant A, Wu H, Krishnamurthy S, Cvjetkovic A, Estelius J, Dedic B, Sethi A, Wallom KL, Riise R, Bäckström M, Wallenius V, Platt FM, Lebens M, Teneberg S, Fändriks L, Kohler JJ, Yrlid U. PLoS Pathog 14 e1006862 (2018)
  10. Tumor marker disaccharide D-Gal-beta 1, 3-GalNAc complexed to heat-labile enterotoxin from Escherichia coli. van den Akker F, Steensma E, Hol WG. Protein Sci 5 1184-1188 (1996)
  11. Suppression of dendritic cell activation by diabetes autoantigens linked to the cholera toxin B subunit. Odumosu O, Payne K, Baez I, Jutzy J, Wall N, Langridge W. Immunobiology 216 447-456 (2011)
  12. Unique biological activity of botulinum D/C mosaic neurotoxin in murine species. Nakamura K, Kohda T, Shibata Y, Tsukamoto K, Arimitsu H, Hayashi M, Mukamoto M, Sasakawa N, Kozaki S. Infect Immun 80 2886-2893 (2012)
  13. Structural inferences for Cholera toxin mutations in Vibrio cholerae. Shamini G, Ravichandran M, Sinnott JT, Somboonwit C, Sidhu HS, Shapshak P, Kangueane P. Bioinformation 6 1-9 (2011)
  14. Loss of biological activity due to Glu-->Arg mutation at residue 11 of the B subunit of cholera toxin. Yamaoka J, Yamasaki S, Kurazono H, Imamura S, Noda M, Miyai K, Takeda Y. Microb Pathog 23 297-302 (1997)
  15. Parametric models to compute tryptophan fluorescence wavelengths from classical protein simulations. Lopez AJ, Martínez L. J Comput Chem 39 1249-1258 (2018)
  16. Anti-Cholera toxin activity of selected polyphenols from Careya arborea, Punica granatum, and Psidium guajava. Charla R, Patil PP, Patil VS, Bhandare VV, Karoshi V, Balaganur V, Joshi RK, Harish DR, Roy S. Front Cell Infect Microbiol 13 1106293 (2023)


Related citations provided by authors (2)

  1. 2.2 Angstroms Crystal Structure of Cholera Toxin B5 Pentamer Bound to Receptor GM1 Pentasaccharide. Merritt EA, Sarfaty S, Van Den Akker F, L'Hoir C, Martial JA, Hol WGJ Protein Sci. 3 166- (1994)
  2. Analysis of structure and function of the B subunit of cholera toxin by the use of site-directed mutagenesis.. Jobling MG, Holmes RK Mol Microbiol 5 1755-67 (1991)

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