| Project/Area Number |
62570193
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
細菌学
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| Research Institution | Tokusima University |
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Principal Investigator |
TAKUMI Kenji University of Tokushima School of Medicine Assistant Professor, 医学部, 助教授 (90035428)
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| Co-Investigator(Kenkyū-buntansha) |
TAKEOKA Aya University of Tokushima School of medicine Assistant, 医学部, 教務員 (00116831)
KOGA Teturo University of Tokushima School of Medicine Assistant, 医学部, 助手 (20093859)
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| Project Period (FY) |
1987 – 1988
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| Project Status |
Completed (Fiscal Year 1988)
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| Budget Amount *help |
¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 1988: ¥400,000 (Direct Cost: ¥400,000)
Fiscal Year 1987: ¥1,800,000 (Direct Cost: ¥1,800,000)
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| Keywords | Self-assembly and reatachement of structural protein / Immunoelectron microscopy / Pathogenic clostridia / Clostridium boltulinum type E / cell surface structure / 蛋白非分解性ボツリヌス菌 / 共通抗原 / 免疫電顕法 / A型 / 細胞壁 / 蛋白抗原 / 菌体表層蛋白抗原 / 細胞付着 / 免疫電顕法(フェリチン抗体法 / プロティンA-金コロイド法) / A型, E型ボツリヌス菌 / 免疫電顕法(フェリチン抗体法, プロティンA-金コロイド法) |
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| Research Abstract |
Surface layers of regularly arranged (RA) protein subunits are now recognized as a common feature of both eubacterial and archaebacterial cell walls. These RAs are noncovalently attached to the surface of the outer membrane or the peptidoglycan sacculus in linear, tetragonal, or hexahonal arragements, formimg in some bases, the primary barrier between the cell and the environment. Clostridium botulinum type E, which is one of the most important bacterial species for the toxigenicbacterial food-borne poisoning has also such RA layered structure. This study atempted to isolate the RA of the organism and to analyse its immunochemical properties by immunobloting together with its ultramicrostructure by using negatively stained thin sections or immunoelectron microscopy.
The RA was extracted selectively from the cell wall by the tr.eatment with 4 M guanidin hydrochloride (GHCL) or 6 M urea. The extract was then submitted to SDS-PAGE, by which that two protein subunits with molecular weights of 900,000 (90K) and 600,000(6oK) are major constituents of the RA is firstly ascertained. Immunoblot with rabbit antiserum to the whole cells indicated that these proteins are surface-exposed components of the cell wall. This surface localization of the protein subunits also confermed by immunoelectraon microscopically by Con-A-gold particle conjugate method. On the other hand, the subunits of linearly ordered RA layers isolated by 4M GHCL were tested for their ability to reform the regular paterns in the absence of cell wall fragments. In negatively stained preparations the majority of the self-arrenged products of the linearly ordered subunits were observed:the diameter of 40-280nm; the length of about 1 um.
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