Purification of the STB enterotoxin of Escherichia coli and the role of selected amino acids on its secretion, stability and toxicity.
The methanol-insoluble heat-stable enterotoxin of Escherichia coli (STB) was purified and characterized by automated Edman degradation and tryptic peptide analysis. The amino-terminal residue, Ser-24, confirmed that the first 23 amino acids inferred from the gene sequence were removed during translocation through the E. coli inner membrane. Tryptic peptide analysis coupled with automated Edman degradation revealed that disulphide bonds are formed between residues Cys-33 and Cys-71 and between Cys-44 and Cys-59. Oligonucleotide-directed mutagenesis performed on the STB gene demonstrated that disulphide bond formation does not precede translocation of the polypeptide through the inner membrane and that disulphide bridge formation is a periplasmic event; apparently, elimination of either of two disulphides of STB renders the molecule susceptible to periplasmic proteolysis. In addition, a loop defined by the Cys-44-Cys-59 bond contains at least two amino acids (Arg-52 and Asp-53) required for STB toxic activity.
Dreyfus
L A
LA
Division of Cell Biology and Biophysics, School of Biological Sciences, University of Missouri-Kansas City 64110.
Urban
R G
RG
Whipp
S C
SC
Slaughter
C
C
Tachias
K
K
Kupersztoch
Y M
YM
Drefus
L A
LA
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000502:physiology*; D001427:Bacterial Toxins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism / Q000633:toxicity; D001483:Base Sequence; D004269:DNA, Bacterial; D004220:Disulfides / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D004768:Enterotoxins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism / Q000633:toxicity; D004926:Escherichia coli / Q000737:chemistry* / Q000378:metabolism; D029968:Escherichia coli Proteins; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D013329:Structure-Activity Relationship
D000596:Amino Acids; D001427:Bacterial Toxins; D004269:DNA, Bacterial; D004220:Disulfides; D004768:Enterotoxins; D029968:Escherichia coli Proteins; C034151:heat stable toxin (E coli)