| Budget Amount *help |
¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 1997: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 1996: ¥900,000 (Direct Cost: ¥900,000)
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| Research Abstract |
Se, an essential trace element, is known to constitute the active site of glutathione peroxidase (GPx) which catalyzes the reduction of hydroperoxides. Guinea pigs have extremely low activity of cellular GPx (cGPx) in the liver and kidney. To examine the expression of other selenoproteins, we investigated the incorporation of Se into proteins or peptides in the liver of guinea pigs after the intraperitoneal injection of radiolabeled selenite. SDS-PAGE analysis of the liver homogenate revealed that the major band of radioactive Se was found at 18 kDa, which coincided with the molecular weight of phospholipid hydroperoxide GPx (PHGPx). Upon the chromatographic separation by BSP-GSH affinity column and the subsequent DEAB Sephacel column, the 18 kDa Se-containing protein co-eluted with the activity of PHGPx. RT-PCR analysis of mRNA obtained from the liver of guinea pigs using primers constructed based on the rat PHGPx resulted in an amplification of PHGPx cDNA.The sequence of the amplified cDNA showed 98% homology to the rat PHGPx. Thus, in the liver of guinea pigs, in which the expression of cGPx is extremely suppressed, PHGPx gene is actively expressed to form the protein with the activity comparable to that of mice and rats. On the other hand, SDS-PAGE analysis of kidney proteins revealed an existence of a 23 kDa Se-containing protein, This size was similar to that of the monomer of cGPx, but this protein had no enzyme activity. 2-D gel analysis of kidney proteins demonstrated that the isoelectric points of the 23 kDa Se-containing protein were different from that of cGPx. The properties of this Se-containing protein with no GPx activity should be further clarified.
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