| Budget Amount *help |
¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 1987: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 1986: ¥800,000 (Direct Cost: ¥800,000)
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| Research Abstract |
Several flavin containing oxidases, such as xanthine oxidase, D-amino acid oxidase, glucose oxidase, and other enzymes which have been assumed to contain flavin, eg. uricase or ascorbate oxidase, were hydrolyzed by proteolysis and/or acid hydrolysis. The resulting hydrolysates were andlyzed to see whether exist any signal showing the presence of pyrroloquinoline quinone (PQQ) or PQQ-chromophores. After fractionation with Sephadex G-10 column, each fraction was examined of its activity as the prosthetic group for apo-glucose dehydrogenase (EC 1.1.99.17). From the chromatogram of some reference proteins in which PQQ is never involved, such as lysozyme and NAD-dependent alcohol dehydrogenase of several recrystallized preparations, showed no PQQ activity. On the other hand, of the examine candidates, the hydrolysate from uricase, xanthine oxidase, ascorbate oxidase potently showed the presence of PQQ in their molecule. These hydrolysate were also available as the growth stimulating substance for some microorganisms to reduce the lag phase in their growth. Spectroscopic data indicated the possibility of the existence of PQQ or PQQ-related compound in individual enzyme molecule.
Alternative model experiment was carried out with PQQ and bovine serum albumin. When PQQ was mixed with bovine serum albumin at neutral pH, a spontaneous binding of PQQ to serum albumin was observed. The resulting PQQ-BSA was fairly stable for chromatographic treatments and exhibited absorption spectrum having a peak at 336 nm. PQQ was convinced to be asociated via ipsilon amino group of lysyl residue of BSA via a Schiff base.
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