| Budget Amount *help |
¥6,800,000 (Direct Cost: ¥6,800,000)
Fiscal Year 1990: ¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 1989: ¥4,900,000 (Direct Cost: ¥4,900,000)
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| Research Abstract |
In this study, I aimed at (1) elucidating the mechanism of oxalogenesis in mammals, and (2) cloning and sequencing the serine : pyruvate aminotransferase (SPT) cDNA and gene from human liver for the gene analysis in the future of primary hyperozaluria type 1 (PH1). Since a patient with PH1 died during the period of this study, SPT-cDNA was also cloned from the patient's liver and the nucleotide sequence was determined.
With respect to the oxalate production in vivo, none of the three enzymes known to catalyze in vitro the oxidation of glyoxylate to oxalate appeared to play substantial role. Instead, oral administration to rat of thiazolidine-2, 4-dicarboxylate, an adduct of glyoxylate with cysteine, was shown to increase slightly the urinary excretion of oxalate. Studies on the possible involvement of thiazolidine-2, 4-dicarboxylate in the oxalate formation from glyoxylate are currently under way. With respect to the genetic defect in the case of PH1, nucleotide sequence analysis of SPT-cDNA from the patient's liver revealed a point mutation of T to C at position 613 (relative to the first nucleotide, A, of the ATG triplet for the initiation Met) encoding a Ser to Pro substitution at residue 205. The T to C conversion created a new SmaI site, which enabled us to demonstrate that the point mutation had occurred in the patient's SPT gene. SmaI digestion of genomic DNA may be useful for the diagnostic gene analysis of this type of PH1. In the liver of this patient, SPT was very low with respect to not only the activity but also the protein detectable on Western blot and immunocytochemical analyses, but the level of SPT-mRNA was found on RNA blot analysis to be even higher than normal, suggesting that the degradation of SPT is accelerated in the patient's liver due to the mutation.
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