| Project/Area Number |
61570208
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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 | Osaka university |
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Principal Investigator |
KONDO Masaomi Osaka University・ Professor, 薬学部, 教授 (90028829)
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| Co-Investigator(Kenkyū-buntansha) |
IMAGAWA Masayoshi Osaka University・ Research Associate, 薬学部, 助手 (20136823)
TAKUBO Yoshihiro Osaka University・ Research Associate, 薬学部, 助手 (40171590)
NISHIHARA Tsutom Osaka University・ Assistant Professor, 薬学部, 助教授 (50028859)
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| Project Period (FY) |
1986 – 1987
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| Project Status |
Completed (Fiscal Year 1987)
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| Budget Amount *help |
¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1987: ¥200,000 (Direct Cost: ¥200,000)
Fiscal Year 1986: ¥1,800,000 (Direct Cost: ¥1,800,000)
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| Keywords | B. megaterium / Outer coat / Gelactosamine-6-phosphate / UDP-GlcNAc-4-epimerase / UDP-GalNAc-deacetylase / outer coat protein / outermost layer deficient mutant / トランスポゾン挿入変異 / 芽胞形成機構 / 芽胞殻 / UDP-N-acetylgalactosamine-4-epimerase / 芽胞殻蛋白特異抗体 |
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| Research Abstract |
We have studied the biosynthesis and the deposition of galactosamine-6-phosphate (Gal-6-P) polymer and some proteins during morphogenesis of the outer coat, the outermost layer of the coat of Bacillus megaterium. Immunological analysis demonstrated that the initial enzyme of the pathway for polysaccharide biosynthesis, UDP-GlcNAc-4-epimerase (GEP), began to be synthesized at T6 (Tn indicates n hours after the end of exponential growth). The structure of one of the intermediates in the later sporulating cells was elucidated to be UDP-GalN from FAB-MS and ^<13>C-NMR spectra and the activity of the deacetylation enzyme, UDP-GalNAc deacetylase (GDA) was detected from T8, two hours later than GEP. The outer coat contained three major proteins, P48, P36 and P22. P48 and P36 were found to deposit mainly by ionic bond and P22 by strong hydrophobic bond on the cell, respectively. Moreover P36 was indicated to exist on the very surface of the spore from immunoblot analysis using specific anti-surface coat protein IgG. SDS-polycarylamide gel electrophoresis of the coat proteins after treatment with pronase suggested that P48 was inside of the outermost layer. Analysis of the outer coat deficient mutant, MAE05 revealed that 1) this strain can synthesize P36 and P22 but not P48, 2) P36 and P22 deposit on the forespore and 3) P36 and P22 are lost form the forespore when mother cell are lysed. Protease-like activity was found in the culture supernatant of the later sporulating cells but it could not digest these proteins on parent mature spore. These results suggest that both O48 and Gal-6-P polymer are not indispensable for deposition of O36 and P22 and Gal-6-P polymer rather acts as barrier against loss of these proteins. The property of some transposon-inserted outer coat deficient mutants may indicate the same ordered, linear expression of P48, and both GEP and GDE genes.
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