[en] Cholera toxin (CT) is an AB5 hexameric protein responsible for the symptoms produced by Vibrio cholerae infection. In the first step of cell intoxication, the B-pentamer of the toxin binds specifically to the branched pentasaccharide moiety of ganglioside GM1 on the surface of target human intestinal epithelial cells. We present here the crystal structure of the cholera toxin B-pentamer complexed with the GM1 pentasaccharide. Each receptor binding site on the toxin is found to lie primarily within a single B-subunit, with a single solvent-mediated hydrogen bond from residue Gly 33 of an adjacent subunit. The large majority of interactions between the receptor and the toxin involve the 2 terminal sugars of GM1, galactose and sialic acid, with a smaller contribution from the N-acetyl galactosamine residue. The binding of GM1 to cholera toxin thus resembles a 2-fingered grip: the Gal(beta 1-3)GalNAc moiety representing the "forefinger" and the sialic acid representing the "thumb." The residues forming the binding site are conserved between cholera toxin and the homologous heat-labile enterotoxin from Escherichia coli, with the sole exception of His 13. Some reported differences in the binding affinity of the 2 toxins for gangliosides other than GM1 may be rationalized by sequence differences at this residue. The CTB5:GM1 pentasaccharide complex described here provides a detailed view of a protein:ganglioside specific binding interaction, and as such is of interest not only for understanding cholera pathogenesis and for the design of drugs and development of vaccines but also for modeling other protein:ganglioside interactions such as those involved in GM1-mediated signal transduction.
Disciplines :
Biochemistry, biophysics & molecular biology
Author, co-author :
Merritt, E. A.
Sarfaty, S.
van den Akker, F.
L'Hoir, C.
Martial, Joseph ; Université de Liège - ULiège > Département des sciences de la vie > GIGA-R : Biologie et génétique moléculaire
Hol, W. G.
Language :
English
Title :
Crystal structure of cholera toxin B-pentamer bound to receptor GM1 pentasaccharide
Publication date :
1994
Journal title :
Protein Science: A Publication of the Protein Society
ISSN :
0961-8368
eISSN :
1469-896X
Publisher :
Cold Spring Harbor Laboratory Press, Woodbury, United States - New York
Acquotti D, Poppe L, Dabrowski J, von der Lieth CW, Sonnino S, Tettamanti G. (1990) Three‐dimensional structure of the oligosaccharide chain of GMI ganglioside revealed by a distance‐mapping procedure: A rotating and laboratory frame nuclear Overhauser enhancement investigation of native glycolipid in dimethyl sulfoxide and in water‐dodecylphospho‐choline solutions. J Am Chem Soc 112:7112-7778.
Brünger A. X‐PLOR, version 2.1. Manual., New Haven, Connecticut, Yale University; 1990.
Cassel D, Pfeuffer T. (1978) Mechanism of cholera toxin action: Covalent modification of the guanyl nucleotide‐binding protein of the adenylate cyclase system. Proc Natl Acad Sci USA 75:2669-2673.
Chen KS, Strober W. (1990) Cholera holotoxin and its B subunit enhance Peyer's patch B cell responses induced by orally administered influenza virus: Disproportionate cholera toxin enhancement of the IgA B cell response. Eur J Immunol 20:433-436.
Clements JD, Hartzog NM, Lyon FL (1988) Adjuvant activity of Escherichia coli heat‐labile enterotoxin and effect on the induction of oral tolerance in mice to unrelated protein antigens. Vaccine 6:269-277.
Czerkinsky C, Russell MW, Lycke N, Lindblad M, Holmgren J. (1989) Oral administration of a streptococcal antigen coupled to cholera toxin B subunit evokes strong antibody responses in salivary glands and extramu‐cosal tissues. Infect Immun 57:1072-1077.
Dallas WS (1983) Conformity between heat‐labile toxin genes from human and porcine enterotoxigenic Escherichia coli. Infect Immun 40:647-652.
De Aizpurua HJ, Russell‐Jones GJ (1988) Oral vaccination. Identification of classes of proteins that provoke an immune response upon oral feeding. J Exp Med 167:440-451.
Dertzbaugh MT, Peterson DL, Macrina FL (1990) Cholera toxin B‐subunit gene fusion: Structural and functional analysis of the chimeric protein. Infect Immun 58:70-79.
De Wolf MJS, Fridkin M, Kohn LD (1981) Tryptophan residues of cholera toxin and its A and B protomers; intrinsic fluorescence and solute quenching upon interacting with the ganglioside GM1, oligo‐GM1 or dansylated oligo‐GM1. J Biol Chem 256:5489-5496.
Eidels L, Proia RL, Hart DA (1983) Membrane receptors for bacterial toxins. Microbiol Rev 47:596-620.
Elson CO, Ealding W. (1984) Generalized systemic and mucosal immunity in mice after mucosal stimulation with cholera toxin. J Immunol 132:2736-2741.
Elson CO, Ealding W. (1984) Cholera toxin feeding did not induce oral tolerance in mice and abrogated oral tolerance to an unrelated protein antigen. J Immunol 133:2892-2897.
Fukuta S, Magnani JL, Twiddy EM, Holmes RK, Ginsburg V. (1988) Comparison of the carbohydrate‐binding specificities of cholera toxin and Escherichia coli heat‐labile enterotoxins LTh‐I, LT‐IIa and LT‐IIb. Infect lmmun 56:1748-1753.
Hilbush BS, Levine JM (1992) Modulation of a Ca2+ signaling pathway by GM1 ganglioside in PC12 cells. J Biol Chem 267:24789-24795.
Jobling MG, Holmes RK (1991) Analysis of structure and function of the B subunit of cholera toxin by the use of site‐directed mutagenesis. Mol Microbiol 5:1755-1767.
Kraulis P. (1991) MOLSCRIPT: A program to produce both detailed and schematic plots of proteins. J Appl Crystallogr 24:946-950.
Leong J, Vinal AC, Dallas WS (1985) Nucleotide sequence comparison between heat‐labile toxin B‐subunit cistrons from Escherichra coli of human and porcine origin. Infect Immun 48:73-77.
L'hoir C, Renard A, Martial JA (1990) Expression in Escherichia coli of two mutated genes encoding the cholera toxin B subunit. Gene 89:47-52.
Liang X, Lamm ME, Nedrud JG (1988) Oral administration of cholera toxin‐Sendai virus conjugate potentiates gut and respiratory immunity against Sendai virus. J Immunol 141:1495-1501.
Lycke N, Tsuji T, Holmgren J. (1992) The adjuvant effect of Vibrio cholerae and Escherichia coli heat‐labile enterotoxin is linked to their ADP‐ribosyltransferase activity. Eur J Immunol 22:2277-2281.
Morrison WJ, Offner J, Vandenbark AA (1990) Transmembrane signalling associated with ganglioside‐induced CD4 modulation. Immunopharma‐cology 20:135-141.
Morrison WJ, Offner J, Vandenbark AA (1991) Ganglioside (GM1)‐treated T cells shed CD4. Immunopharmacology 22:77-84.
Poppe L, von der Lieth CW, Dabrowski J. (1990) Conformation of the glycolipid globoside head group in various solvents and in the micelle‐bound state. J Am Chem Soc 112:7762-7771.
Quiocho FA (1988) Molecular features and basic understanding of protein‐carbohydrate interactions: The arabinose‐binding protein‐sugar complex. Curr Topics Microbiol Immunol 139:135-148.
Schödel F, Enders G, Jung MC, Will H. (1990) Recognition of a hepatitis B virus nucleocapsid T‐cell epitope expressed as a fusion protein with the subunit B of Escherichia coli heat labile enterotoxin in attenuated salmonellae. Vaccine 8:569-572.
Schön A, Freire E. (1989) Thermodynamics of intersubunit interactions in cholera toxin upon binding to the oligosaccharide portion of its cell surface receptor, ganglioside GM1. Biochemistry 28:5019-5024.
Sixma TK, Kalk KH, van Zanten BAM, Dauter Z, Kingma J, Witholt B, Hol WGJ (1993) Refined structure of Escherichia coli heat‐labile enterotoxin, a close relative of cholera toxin. J Mol Biol 230:890-918.
Sixma TK, Pronk SE, Kalk KH, van Zanten BAM, Berghuis AM, Hol WGJ (1992) Lactose binding to heat‐labile enterotoxin revealed by X‐ray crystallography. Nature 355:561-564.
Sixma TK, Pronk SE, Kalk KH, Wartna ES, van Zanten BAM, Witholt B, Hol WGJ (1991) Crystal structure of a cholera toxin‐related heat‐labile enterotoxin from E. coli. Nature 351:371-378.
Sixma TK, Stein PE, Hol WGJ, Read RJ (1993) Comparison of the B‐pentamers of heat‐labile enterotoxin and verotoxin‐I: Two structures with remarkable similarity and dissimilarity. Biochemistry 32:191-198.
Spiegel S. (1989) Inhibition of protein kinase‐C‐dependent cellular proliferation by interaction of endogenous ganglioside GM1 with the B subunit of cholera toxin. J Biol Chem 264:16512-16517.
Stein PE, Boodhoo A, Tyrrell GJ, Brunton JL, Read RJ (1992) Crystal structure of the cell‐binding B oligomer of verotoxin‐1 from E. coli. Nature 355:748-750.
Tsuji T, Iida T, Honda T, Miwatani T, Nagahama M, Sakurai J, Matsubara H. (1987) A unique amino acid sequence of the B subunit of a heat‐labile enterotoxin isolated from a human enterotoxigenic Escherichia coli. Microb Pathol 2:381-390.
Van Heyningen S. (1983) The interaction of cholera toxin with gangliosides and the cell membrane. Curr Topics Membranes Transport 18:445-471.
Varghese JN, McKimm‐Breschkin JL, Caldwell JB, Kortt AA, Colman PM (1992) The structure of the complex between influenza virus neuraminidase and sialic acid, the viral receptor. Proteins 14:327-332.
Vyas NK (1991) Atomic features of protein‐carbohydrate interactions. Curr Opin Struct Biol 1:732-740.
Weis FMB, Davis RJ (1990) Regulation of epidermal growth factor receptor signal transduction. J Biol Chem 265:12059-12066.
Weis W, Brown JH, Cusack S, Paulson JC, Skehel JJ, Wiley DC (1988) Structure of the influenza virus haemagglutinin complexed with its receptor, sialic acid. Nature 333:426-431.
Wright CS (1990) 2.2 A resolution structure analysis of two refined N‐acetylneuraminyl‐lactose‐wheat germ agglutinin isolectin complexes. J Mol Biol 275:635-651.
Wu HY, Russell MW (1993) Induction of mucosal immunity by intranasal application of a streptococcal surface protein antigen with the cholera toxin B subunit. Infecr Immun 61:314-322.
Yamamoto T, Yokota T. (1983) Sequence of heat‐labile enterotoxin of Escherichia coli pathogenic for humans. J Bacteriol 155:728-733.