1992 Fiscal Year Final Research Report Summary
Experimental device designed for assay and purification of oxygen-sensitive enzymes and its application.
| Project/Area Number |
02557070
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| Research Category |
Grant-in-Aid for Developmental Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
Functional basic dentistry
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| Research Institution | TOHOKU UNIVERSITY |
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Principal Investigator |
YAMADA Tadashi Tohoku Univ., School of Dentistry, Prof., 歯学部, 教授 (50005021)
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| Co-Investigator(Kenkyū-buntansha) |
TAKAHASHI Nobuhiro Tohoku Univ., School of Dentistry, Assistant Prof., 歯学部, 助手 (60183852)
IWAMI Yoshimiti Tohoku Univ., School of Dentistry, Assistant Prof., 歯学部, 助手 (60005030)
ABBE Kazuhiko Tohoku Univ., School of Dentistry, Associate Prof., 歯学部, 助教授 (40151089)
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| Project Period (FY) |
1990 – 1992
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| Keywords | Oxygen sensitive enzyme / Pyruvate formate-lyase / Strictly anaerobic conditions / Oral streptococci / Sugar metabolism / Anaerobic glove box / Enzyme assay / Actinomyces |
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| Research Abstract |
An anaerobic glove box, equipped with a purification apparatus and a spectorophotometer, was manufactured as a trial and employed. Several interesting finding were turned out as follows.
Pyruvate formate-lyase (PFL) plays an important role in anaerobic sugar metabolism in oral streptococci. PFL was extremely sensitive to oxygen, and assumed to contain a protein radical near a catalytic site. The reaction of molecular oxygen with the radical seemed to generate further harmful active oxygen and bring about irreversible inactivation of the enzyme rapidly. Reversible inactive form of the enzyme was oxygen torelant since it did not contain the radical, and could be activated by PFL-activating enzyme. Partially purified PFL activating enzyme was quite unstable, but incubation of the enzyme with ferrous ion, selenium ion, pterin, restored a part of the activity. The enzyme seemed to contain "a pterin like cofactor" which was assumed to be exist in xanthine oxidase. The activating enzyme of Streptococcus mutans was triggered by pyruvate, but not oxamate, and quite unstable. The properties of the streptococcal enzyme is considered different from that of Escherichia coli.
It is essential for plaque microorganisms to keep intracellular NADH/NAD balance under anaerobic conditions. Oral streptococci employed "the PFL system" to keep the balance. Lactobacilli metabolized acetate into ethanol to oxidize NADH to keep it. Actinomyces could regenerate NAD via succinate pathway when bicarbonate was available, and increased the rate of acid production.
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