| Project/Area Number |
14580651
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Functional biochemistry
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| Research Institution | Kyushu Dental College |
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Principal Investigator |
NOGUCHI Tomoo Kyushu Dental College, Dept.of dentistry, Professor, 歯学部, 教授 (30073688)
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| Co-Investigator(Kenkyū-buntansha) |
MASUDA Wataru Kyushu Dental College, Dept.of dentistry, Assistant, 歯学部, 助手 (80295865)
HAYASHI Sueko Kyushu Dental College, Dept.of dentistry, Assistant Professor, 歯学部, 講師 (30047807)
FUJIWARA Satoko Kyushu Dental College, Dept.of dentistry, Associate Professor, 歯学部, 助教授 (20047806)
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| Project Period (FY) |
2002 – 2003
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| Project Status |
Completed (Fiscal Year 2003)
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| Budget Amount *help |
¥3,000,000 (Direct Cost: ¥3,000,000)
Fiscal Year 2003: ¥1,200,000 (Direct Cost: ¥1,200,000)
Fiscal Year 2002: ¥1,800,000 (Direct Cost: ¥1,800,000)
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| Keywords | Animal evolution / Purine degradation / Urate degradation / Allantoinase / Allantoicase / Ureidoglycollate lyase / Peroxisomes / Frog allantoinase cDNA / ウレイドグリコレートリアーゼ |
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| Research Abstract |
End products of purine degradation vary from species to species. The end product of purine degrade is urate in humans, hominoid primates and new world monkeys, allantoin by uricase in mammals other than primates and old world monkeys, allantoiate by allantoinase(ALN) in some teleosts, urea by allantoicase(ALC) in fish and amphibians and ammonia by urease in many invertebrates.
The degradation of purines to urate is common to all animal species, while the degradation of urate is much less complete in higher animals. We have reported that in marine fish, the degrading enzymes of purines to urate are located in the cytosol, while the degrading enzymes of urate to urea are located in the peroxisomes. This shows that in purine degradation, peroxisomal enzymes have been lost during evolution. ALN and ALC are different proteins in fish liver, whereas the two enzymes form a complex in amphibian liver.
We examined how the mechanism to form amphibian allantoinase-allantoicase complex(ALNC) is lost
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in higher animals during animal evolution (1).The cDNA encoding ALN of the subunit of amphibian ALNC was sequenced. We got some clones from λ gt11 library using anti ALN of bullfrog (2).In saltwater fish liver, the end products of purine degradation were urea and glyoxylate. Glyoxylate may be converted to glycine by alanine : glyoxylate aminotransferase for the reutilization of purine carbons (3).It is generally accepted that all of the allantoin-degrading enzymes were lost during mammalian evolution. Surprisingly, ureidoglycollate lyase has been found in mammalian tissue. The apparent Km(17mM) of the rat enzyme for ureidoglycollate was much higher than that (0.33mM)of fish-liver ureidoglycollate lyase. Mammals have lost the function in vivo by elevating the Km for ureidoglycollate during evolution (4).We reported the isolation of cDNA encoding frog catalase from a liver library cloned in λgt11. The 528 amino acid sequence deduced from the nucleotid sequence has a calculated Mr of 59871. We found free ALN only with ALN activity which is differ from ALN subunit composed of ALNC. The enzyme shows the middle type of fish ALN and amphibian ALNC (5).We separated the fish with three kinds of habitat : salt water, brackish water and fresh water and with three kinds of food habit : carnivorous,herbivorous and omnivovous. We determined the subcellular distribution of ALN, peroxisomes matrix or cytosoles. We determined ALC type, peroxisomal matrix or membrane-bound of outer surface of peroxisomes. The cDNA of each fish, was synthesized with RT-PCR using mRNA and The full-length was determined by the DNA sequencer. We compared the results with subunit of frog ALN and Xenopus liver ALC of C-end amino acid sequence and determined the signal of transference to and from distribution in peroxisomes Less
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