Cloning and expression of a Serpula (Treponema) hyodysenteriae hemolysin gene.
Serpula (Treponema) hyodysenteriae, the etiologic agent of swine dysentery, produces a hemolysin which is thought to be an important factor in the pathogenesis of the disease. We report the cloning, sequencing, and expression of a hemolysin gene (tly) from S. hyodysenteriae B204. A pUC19 gene bank of strain B204 was constructed in the Escherichia coli K-12 strain DH5 alpha, and hemolytic recombinants were identified by plating the library on blood agar plates. From the hemolytic recombinants, a 1.5-kb DNA fragment could be isolated that contained information necessary for the production of a hemolysin/cytotoxin in E. coli. Nucleotide sequence determination of this 1.5-kb fragment showed that it contained an open reading frame capable of encoding a 26.9-kDa protein. The recombinant hemolysin was easily released from E. coli by osmotic shock. As with the native hemolysin, the recombinant hemolysin is EDTA insensitive, thermolabile, and cytotoxic for several eukaryotic cell lines. Southern blot hybridization showed that the cloned S. hyodysenteriae hemolysin gene tly is present in all pathogenic strains of S. hyodysenteriae tested and absent in the nonpathogenic, weakly hemolytic spirochete S. innocens.
Muir
S
S
Department of Molecular Biology, Research Institute of Scripps Clinic, La Jolla, California 92037.
Koopman
M B
MB
Libby
S J
SJ
Joens
L A
LA
Heffron
F
F
Kusters
J G
JG
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002872:Chromosome Deletion; D003001:Cloning, Molecular; D006224:Cricetinae; D004285:Dogs; D015870:Gene Expression; D005798:Genes, Bacterial; D006460:Hemolysin Proteins / Q000032:analysis / Q000235:genetics* / Q000633:toxicity; D006801:Humans; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000494:pharmacology; D011994:Recombinant Proteins / Q000032:analysis / Q000633:toxicity; D014208:Treponema / Q000235:genetics* / Q000472:pathogenicity; D014774:Virulence
D006460:Hemolysin Proteins; D009842:Oligopeptides; D011994:Recombinant Proteins; C047981:arginyl-glycyl-aspartic acid