Isolation of immunogenic and lethal peptides of alpha-toxin from Clostridium novyi type B.
The lethal alpha-toxin was isolated from the culture filtrate of Clostridium novyi type B using ammonium sulfate precipitation and ion exchange chromatography. The alpha-toxin has a mol. wt of 190,000 and does not contain any disulfide cross-linkages. It consists of a single polypeptide chain. The peptide fragments resulting from the cyanogen-bromide cleavage were isolated using reversed phase and gel filtration HPLC. The immunogenic actions of these peptides and peptide mixtures were studied in Balb/c mice. Three polyclonal antisera recognizing the uncleaved native toxin could be found using an ELISA test (Br3, Bro2, Bro5). One peptide mixture (Tx5), which was proved lethal in shell-less quail eggs (in vitro), was rechromatographed with gel filtration HPLC that resulted in one peptide with mol. wt 3000 (Txleth), which again proved lethal in the shell-less quail egg lethality test. The immunogenic peptides differ from the lethal one, therefore we assumed different locations on the polypeptide chain. The separation of the immunogenic, non-toxic fragment from the lethal one may allow the production of a highly specific non-toxic vaccine. By using synthetically produced immunogenic peptides, time-consuming purification methods and working with the whole toxin will become unnecessary.
Schranner
I
I
Institut für Physiologie, Physiologische Chemie und Ernährungsphysiologie, Tierärztliche Fakultät, Universität München, F.R.G.
Erhard
M H
MH
Kaltner
H
H
Lösch
U
U
D000595:Amino Acid Sequence; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D003013:Clostridium / Q000737:chemistry*; D003370:Coturnix / Q000196:embryology; D046508:Culture Techniques; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D007114:Immunization; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D010455:Peptides / Q000276:immunology / Q000302:isolation & purification* / Q000633:toxicity; D010738:Type C Phospholipases / Q000737:chemistry* / Q000302:isolation & purification
D010455:Peptides; D010738:Type C Phospholipases