Development of two novel nontoxic mutants of Escherichia coli heat-labile enterotoxin.
- Author:
Eun Jeong PARK
1
;
Ji Hoon CHANG
;
Jang Seong KIM
;
Soo Il CHUNG
;
Jung Sun YUM
Author Information
1. Mogam Biotechnology Research Institute, Kyonggido, Korea.
- Publication Type:Original Article ; Research Support, Non-U.S. Gov't
- Keywords:
mutant LT (mLT);
ADP-ribosyltransferase activity;
cAMP;
immunogenicity;
secretory IgA (sIgA)
- MeSH:
Amino Acid Substitution;
Animal;
Bacterial Toxins/toxicity*;
Bacterial Toxins/metabolism;
Bacterial Toxins/immunology*;
Bacterial Toxins/genetics;
CHO Cells;
Cyclic AMP/metabolism;
Enterotoxins/toxicity*;
Enterotoxins/metabolism;
Enterotoxins/immunology*;
Enterotoxins/genetics;
Enzyme-Linked Immunosorbent Assay;
Escherichia coli*/metabolism;
Escherichia coli*/genetics;
Female;
Hamsters;
IgA, Secretory/blood;
Ileum/metabolism;
Immunity, Mucosal;
Mice;
Mice, Inbred BALB C;
Mutagenesis, Site-Directed;
NAD+ ADP-Ribosyltransferase/metabolism;
Recombinant Proteins/toxicity;
Recombinant Proteins/metabolism;
Recombinant Proteins/immunology;
Recombinant Proteins/chemistry
- From:Experimental & Molecular Medicine
1999;31(2):101-107
- CountryRepublic of Korea
- Language:English
-
Abstract:
Escherichia coli heat-labile enterotoxin (LT) is composed of catalytic A and non-catalytic homo-pentameric B subunits and causes diarrheal disease in human and animals. In order to produce a nontoxic LT for vaccine and adjuvant development, two novel derivatives of LT were constructed by a site-directed mutagenesis of A subunit; Ser63 to Tyr63 in LTS63Y and Glu110, Glu112 were deleted in LT delta 110/112. The purified mutant LTs (mLTs) showed a similar molecular structural complex as AB5 to that of wild LT. In contrast to wild-type LT, mLTs failed to induce either elongation activity, ADP-ribosyltransferase activity, cAMP synthesis in CHO cells or fluid accumulation in mouse small intestine in vivo. Mice immunized with mLTs either intragastrically or intranasally elicited high titers of LT-specific serum and mucosal antibodies comparable to those induced by wild-type LT. These results indicate that substitution of Ser63 to Tyr63 or deletion of Glu110 and Glu112 eliminate the toxicity of LT without a change of AB5 conformation, and both mutants are immunogenic to LT itself. Therefore, both mLTs may be used to develop novel anti-diarrheal vaccines against enterotoxigenic E. coli.
