1996
DOI: 10.1128/iai.64.12.5413-5416.1996
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Mutants of the Escherichia coli heat-labile enterotoxin with reduced ADP-ribosylation activity or no activity retain the immunogenic properties of the native holotoxin

Abstract: The Escherichia coli heat-labile enterotoxin (LT) is a potent inducer of mucosal immune responses. In a previous study (L. DeHaan, W. R. Verweij, M. Holtrop, E. Agsteribbe, and J. Wilschut, Vaccine 14:620-626, 1996), we have shown that efficient induction of an LTB-specific mucosal immune response by LT requires the presence of the LTA chain, suggesting a possible role of the enzymatic activity of LTA in the induction of these responses. In the present study, we generated LT mutants with altered ADP-ribosylati… Show more

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Cited by 57 publications
(40 citation statements)
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“…Among the various candidates for mucosal antigen delivery, Sendai-virus-associated fusion protein seems particularly suited to function as a molecule that of cholera toxin (nCT) and heat-labile enterotoxin (nLT) cause severe diarrhoea and so are unsuitable for use in humans. To overcome these hurdles, researchers have substituted a single amino acid to generate non-toxic mutant forms of cholera toxin (mCT) and heat-labile enterotoxin (mLT) [85][86][87][88] ; these retain the adjuvanticity of the native forms but do not induce the ribosylation of ADP that is associated with toxic activity. Our efforts to devise a safe first generation toxin-based adjuvant have focused on mCT S61F (in which phenylalanine replaces serine at position 61) and mCT E112K (in which lysine replaces glutamic acid at position 112); these mutations were created by making a single amino-acid substitution in the active centre of the ADP-ribosyltransferase in the A subunit of cholera toxin 89 .…”
Section: Nalt-targeted Vaccine Deliverymentioning
confidence: 99%
“…Among the various candidates for mucosal antigen delivery, Sendai-virus-associated fusion protein seems particularly suited to function as a molecule that of cholera toxin (nCT) and heat-labile enterotoxin (nLT) cause severe diarrhoea and so are unsuitable for use in humans. To overcome these hurdles, researchers have substituted a single amino acid to generate non-toxic mutant forms of cholera toxin (mCT) and heat-labile enterotoxin (mLT) [85][86][87][88] ; these retain the adjuvanticity of the native forms but do not induce the ribosylation of ADP that is associated with toxic activity. Our efforts to devise a safe first generation toxin-based adjuvant have focused on mCT S61F (in which phenylalanine replaces serine at position 61) and mCT E112K (in which lysine replaces glutamic acid at position 112); these mutations were created by making a single amino-acid substitution in the active centre of the ADP-ribosyltransferase in the A subunit of cholera toxin 89 .…”
Section: Nalt-targeted Vaccine Deliverymentioning
confidence: 99%
“…LT mutants LT-T50G (LTA: Thr50 1 Gly) and LT-E112K (LTA: Glu112 1 Lys) were constructed, expressed and purified as previously described [22,23]. Both mutants retained affinity for the toxin receptor G M1 (Table 1, see also [22]).…”
Section: Role Of Adp-ribosylation Activity In the Adjuvant Propertiesmentioning
confidence: 99%
“…In addition, these mutants failed to stimulate intracellular cAMP levels in a cAMP accumulation test. However, in contrast to LT-E112K, the T50G mutant retained the capacity of wild-type LT to change chinese hamster ovary (CHO) cell morphology [23,24], suggesting that LT-T50G had retained some enzymatic activity. Therefore, we now reexamined the enzymatic activity of these mutants in a refined DEABAG assay, as described by Soman et al [25].…”
Section: Role Of Adp-ribosylation Activity In the Adjuvant Propertiesmentioning
confidence: 99%
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“…The current focus of engineering vaccines against LT-I or CT is to use the complete toxin including the A subunit, preferably inactivated to eliminate its toxic activity in the gut. Recently, several groups were able to remove the toxic activity of LT or CT by site-directed mutagenesis in the A subunit while retaining the adjuvant activity of the ABS hetero-hexamer (Dickinson & Clements, 1995;Douce et al, 1995;de Haan et al, 1996;Rappuoli et al, 1996). One potential disadvantage of vaccines containing these toxins is that, as the name heat-labile enterotoxin implies, the ABs holotoxin is not thermostable.…”
mentioning
confidence: 99%