| Project/Area Number |
01570236
|
|---|
| Research Category |
Grant-in-Aid for General Scientific Research (C)
|
| Allocation Type | Single-year Grants |
|---|
| Research Field |
細菌学
|
|---|
| Research Institution | Research Institute for Microbial Diseases, Osaka University |
|---|
Principal Investigator |
YAMAMOTO Koichiro Biken, Osaka Univ., Research Instructor, 微生物病研究所, 助手 (30158274)
|
|---|
| Co-Investigator(Kenkyū-buntansha) |
YOH Myonsun Biken, Osaka Univ., Research Instructor, 微生物病研究所, 助手 (70093482)
|
|---|
| Project Period (FY) |
1989 – 1990
|
| Project Status |
Completed (Fiscal Year 1990)
|
| Budget Amount *help |
¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 1990: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 1989: ¥1,200,000 (Direct Cost: ¥1,200,000)
|
|---|
| Keywords | mutation / Amino Acid Sequence / hemolysin / Pathogenic Factor / cloning / domain / vibrio / Nucleotide Sequence / 毒素 / コレラ菌 / サイトディレクティド変異 / バルニフィカス菌 / タンパク毒素 |
|---|
| Research Abstract |
Site-directed mutagenesis of hlyA of Vibrio cholerae at common domain found in Vibrio vulnificus cytolysin was carried out.
1. Oligonucleotide primers were synthesized as one of the amino acids of the domain in substituted to another amino acid.
The mutations were designed to substitute to another amino acid of which
(A) hydropathy and
(B) charge was greatly changed and
(C) the third nucleotide of the codons was changed
where it gives amino acid substitution.
2. Site-directed mutations were carried out using these oligonucleotide primers and the mutation was confirmed by DNA sequencing and ten mutants were obtained.
3. The mutant precursors were over-produced in E. Coli. The mutant cytolysins were extracted and partially purified. The quantities of the cytolysin antigens and the protein contents were determined.
4. The specific activities of the mutant cytolysins were 1/10 - 1/100 of the native cytolysin. This result suggest that the common domain found in V. Cholerae and V. Vulnificus play an important role(s) in cytolytic activities of the cytolysins.
|
