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KMID : 0377519820070040309
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Chung-Ang Journal of Medicine
1982 Volume.7 No. 4 p.309 ~ p.320
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Purification and Properties of Superoxide Dismutase from Rabbit Brain
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Choi Mun-Sik
Kwon Nyoun-Soo
Lee Hi-Sung
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Abstract
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Distribution and some properities of superoxide dismutase was investigated in rabbit brain. Rabbit brain was fractionated by differential centrifugation into mitochondrial and cytosolic fractions. Cytosolic superoxide dismutase was purified by ammonium sulface precioitation, treatment with ethanol-chloroform mixture, and DEAE-cellulose column chromatography. The activity of superoxide dismutase was measured by method of McCord and Fridovich. The results are summarized as follows. 1. Rabbit brain contains two types of superoxide dismutase, one in the mitochondrial and the other in the cytosol. The mitochondrial superoxidase was inactivated by treatment with mixture of ethanol-chloroform whereas the cytosolic superoxide dismutase was not. 2. The activity in homogenate was found to be 93.3 units/g of wet tissue. The distribution of superoxide dismutase in the subcellular organelle showed that 61% of the activity was in cytosolic fraction and 39% in mitochondrial fraction. 3. The molecular weight og cytosolic superoxide dismutase was estimated to be 32,000 by gel filtration. 4. The straviolet absorption spectrum of cytosolic superoxide dismutase indicated a lack of tryptophan. The spectrum of the enzyme in the altraviolet region was similar to the absorption spectrum of phenylalanine and tyrosine. 5. Purified cytosolic superoxide dismutase contained 2 Cu^2+ and 2 Zn^2+ per mole. This enzyme was found to be similar to the other cupro-zine superoxide dismutase which have been isolated from diverse eukaryotes. 6. Mitochondrial superoxide dismutase contained Mn^2+ and was strikingly similar to the superoxide dismutase previously isolated from bacteria. 7. Cyanide at 0.5mM and 5.0 mM inhibited the activity of cytosolic superoxide dismutase 13% amd 93%, repectively. 8. Superoxide dismutase activity in mitochondria was partitioned 38% in the intermenbrane space and 62% in the matrix. 9. The activity of xanthine oxidase in homogenate was fiund to be 333 units/g of wet tissue and the relative of cytosolic and mitochondrial xanthine oxidase was 77% and 23%, respectively.
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