Reduction of enterotoxic activity of Escherichia coli heat-stable enterotoxin by substitution for an asparagine residue.
The Escherichia coli heat-stable enterotoxins (STs) are small peptide toxins consisting of 18 (STp) or 19 (STh) amino acids. STp and STh share biologically active sequences which reside in the C-terminal 13 amino acid residues, but the role of each amino acid in the active sequences is not clear. We substituted in vivo Asp, Tyr, His, Gln, Lys, and Arg for the Asn residue at position 11 of STp by oligonucleotide-directed site-specific mutagenesis and examined the biological activities of the resulting mutants. All mutant STs reacted with both monoclonal and polyclonal antibodies, demonstrating that the amino acid substitutions at position 11 did not cause a significant change in the conformation of STp. However, the substitutions invariably caused a significant decrease in enterotoxic activities. The most remarkable decrease was observed with Asn-11----Lys-11 and Asn-11----Arg-11 mutations; that is, enterotoxic activity could not be detected in the culture supernatant of either of these mutant strains. These results indicate that Asn-11 of STp plays an essential role in the enterotoxic activity. The amide group and the length of side chain of Asn-11 seem to be especially important for enterotoxic activity.
Okamoto
K
K
Department of Microbiology, School of Medicine, Fujita-Gakuen Health University, Aichi-ken, Japan.
Okamoto
K
K
Yukitake
J
J
Miyama
A
A
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology; D001216:Asparagine; D001427:Bacterial Toxins / Q000276:immunology / Q000633:toxicity*; D004252:DNA Mutational Analysis; D004768:Enterotoxins / Q000276:immunology / Q000633:toxicity*; D004926:Escherichia coli / Q000502:physiology; D029968:Escherichia coli Proteins; D008969:Molecular Sequence Data; D013329:Structure-Activity Relationship
D000911:Antibodies, Monoclonal; D001427:Bacterial Toxins; D004768:Enterotoxins; D029968:Escherichia coli Proteins; C034151:heat stable toxin (E coli); D001216:Asparagine