Isolation of extracytoplasmic proteins from Serpulina hyodysenteriae B204 and molecular cloning of the flaB1 gene encoding a 38-kilodalton flagellar protein.
Extracytoplasmic proteins were released from Serpulina (Treponema) hyodysenteriae (strain B204) by treatment of whole cells with a nonionic detergent (Tween 20). Centrifugation of the Tween 20-released proteins at 100,000 x g sedimented 10 major extracytoplasmic proteins with approximate molecular masses of 44, 43.5, 42, 39, 38, 34, 33.5, 33, 31, and 29 kDa. Treatment of the sedimented fraction with 6 M urea solubilized all of the proteins except the 39-kDa protein. Peptide sequences were obtained for the purified 42-, 39-, 38-, 34-, 31-, and 29-kDa proteins. The peptide sequences of the 42-, 38-, and 31-kDa proteins indicate that they likely are components of the periplasmic flagella. The amino-terminal peptide sequence of the 38-kDa protein was used to design an oligonucleotide probe and to clone an S. hyodysenteriae DNA fragment containing the gene encoding this protein. The predicted 290-amino-acid protein sequence derived from the cloned gene was highly homologous to those of several other bacterial flagellar proteins and is preceded by consensus sigma D nucleotide sequences found upstream of other flagellar genes. On the basis of its similarity to the FlaB proteins of other spirochetes, we propose to designate the cloned S. hyodysenteriae gene flaB1 and its encoded protein FlaB1. Vaccination of pigs with FlaB1 or its recombinant counterpart did not protect them from an experimental challenge.
Gabe
J D
JD
Berlex Biosciences, Brisbane, California 94005.
Chang
R J
RJ
Slomiany
R
R
Andrews
W H
WH
McCaman
M T
MT
D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification / Q000627:therapeutic use; D001483:Base Sequence; D017401:Brachyspira hyodysenteriae / Q000145:classification / Q000235:genetics* / Q000276:immunology; D003001:Cloning, Molecular; D016384:Consensus Sequence; D004403:Dysentery / Q000517:prevention & control / Q000662:veterinary; D004926:Escherichia coli / Q000235:genetics; D005407:Flagella / Q000235:genetics* / Q000276:immunology; D005408:Flagellin; D005798:Genes, Bacterial / Q000235:genetics*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000276:immunology; D017421:Sequence Analysis; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid; D013552:Swine; D013553:Swine Diseases / Q000517:prevention & control; D014611:Vaccination
D001426:Bacterial Proteins; D010446:Peptide Fragments; D011994:Recombinant Proteins; C064192:flaB1 protein, bacteria; D005408:Flagellin