Cloning and sequencing of the gene encoding Vibrio cholerae O1 fimbrial subunit (fimbrillin) |
| |
| Authors: | Masahiko Ehara Yoshio Ichinose Toshiya Hirayama Hisao Kurazano Yuji Hidaka Kouichi Morita Akira Igarashi Shouichi Shimodori |
| |
| Affiliation: | Department of Bacteriology, Institute of Tropical Medicine, Nagasaki University, Sakamoto 1-Chome 12-4, Nagasaki 852 Japan; Department of Microbiology, Faculty of Medicine, Kyoto University, Yoshida Konoe Cho Sakyou-Ku, Kyoto 606, Japan; Institute for Protein Research, Osaka University, Suita, Osaka 565, Japan; Department of Virology, Institute of Tropical Medicine, Nagasaki University, Sakamoto 1-Chome 12-4, Nagasaki 852, Japan; Department of Microbiology, School of Health Sciences, Kyushu University, Maidashi 3-1-1, Higashi-Ku, Fukuoka 812, Japan |
| |
| Abstract: | Abstract The gene encoding an 18 kDa fimbrial subunit of Vibrio cholerae O1 was identified in a fimbriate strain Bgd17. Mixed oligoprimers were prepared based on the amino acid sequence of the N-terminus and that from a cyanogen bromide-cleaved fragment of the fimbrillin. A PCR-amplified 185 bp DNA fragment was sequenced. This 185 bp fragment was further extended to 540 bp to 3' and 5' termini by RNA-PCR using a primer containing a random hexamer at its 3' end. This fragment did not contain the stop codons. It was further extended by a gene walking method using Eco RI cassette and its primers. Finally a 660 bp fragment was obtained and sequenced. This fragment contained the complete open reading frame of the structural subunit of the fimbriae, composed of 169 amino acids with a molecular mass of 17435.65 and a leader sequence of 6 or 9 amino acids. The deduced amino acid sequence of the polypeptide encoded by the gene, designated fim A, displayed a highly conserved sequence of MKXXXGFTLI EL of type 4 fimbriae. |
| |
| Keywords: | Vibro cholerae O1 Fimbriae fim A cloning Hemagglutinin |
|
|
