| Home | E-Submission | Sitemap | Contact us |  
Korean J Parasitol > Volume 29(3):1991 > Article

Original Article
Korean J Parasitol. 1991 Sep;29(3):259-266. English.
Published online Mar 20, 1994.  http://dx.doi.org/10.3347/kjp.1991.29.3.259
Copyright © 1991 by The Korean Society for Parasitology
Purification and characterization of a Cu, Zn-superoxide dismutase from adult Paragonimus westermani
Y B Chung,C Y Song,H S Lee,*Y Kong,** and S Y Cho**
Department of Biology, College of Natural Sciences, Chung-Ang University, Seoul 156-756, Korea.

In cytosolic fraction of adult Paragonimus westermani, superoxide dismutase activity was identified (4.3 units/mg of specific activity) using a xanthine-xanthine oxidase system. The enzyme was purified 150 fold in its activity using the ammonium sulfate precipitation, DEAE-Trisacryl M anion-exchange chromatography and Sephadex G-100 molecular sieve chromatography. The enzyme exhibited the enhanced activity at pH 10.0. The enzyme activity totally disappeared in 1.0mM cyanide while it remained 77.8% even in 10 mM azide. These findings indicated that the enzyme was Cu, Zn-SOD type. Molecular mass of the enzyme was estimated to be 34 kDa by gel filtration and 17 kDa on reducing SDS-polyacrylamide gel electrophoresis which indicated a dimer protein.


Fig. 1
Elution profile of SOD on the 1st DEAE-Trisacryl1 M anion-exchange chromatography. An allocation of each fraction was assayed form SOD activity (○) and protein (•). A linear gradient was indicated by dotted line. Pooled fractions containing SOD activity was shown by a bar (-).

Fig. 2
Elution profile of SOD on the 2nd DEAE-Trisacry1 M anion-exchange chromatography. Markings are the same as described in Fig. 1.

Fig. 3
Elution profile of SOD on the 3nd DEAE-Trisacry1 M anion-exchange chromatography. Markings are the same as described in Fig. 1.

Fig. 4
Eatimation of molecular mass of Cu, Zn-SOD by Sephadex G-100 gel filtration.

A : Bovine serum albumin (68 kDa)

B : Ovalbumin (43 kDa)

C : Chymotrypsinogen A (25 kDa)

D : Cytochrome c (12.4 kDa)

Fig. 5
Findings on SDS-PAGE of Cu, Zn-SOD of P. westermani. The samples were electrophoresed at 12.5% gel in reducing condition.

Mr : Molecular mass in kDa

C : Crude extract

1 : 1st DEAE-Trisacryl M anion-exchange chromatography

2 : 2nd DEAE-Trisacryl M anion-exchange chromatography

3 : 3rd DEAE-Trisacryl M anion-exchange chromatography

4 : Sephadex G-100 column chromatography


Table 1
Purification of cytosolic superoxide dismutase from P. westermani

Table 2
Effects of inhibitors on activity of cytosolic SOD of P. westermani

Table 3
Effects of pH on activity of cytosolic SOD of P. westermani

1. Bromberg Y, et al. J Biol Chem 1985;290:13539–13545.
2. Callahan HL, Crouch RK, James ER. Helminth anti-oxidant enzymes: a protective mechanism against host oxidants?. Parasitol Today 1988;4(8):218–225.
3. Choi WY, Lee OR, Jin YK, Chi JG. [Lung Findings In Experimental Paragonimiasis]. Korean J Parasitol 1979;17(2):132–146.
4. Dryer SE, Dryer RL, Autor AP. Enhancement of mitochondrial, cyanide-resistant superoxide dismutase in the livers of rats treated with 2,4-dinitrophenol. J Biol Chem 1980;255(3):1054–1057.
5. Fairfield AS, Meshnick SR, Eaton JW. Malaria parasites adopt host cell superoxide dismutase. Science 1983;221(4612):764–766.
6. Fridovich I. Superoxide dismutases. Annu Rev Biochem 1975;44:147–159.
7. Geller BL, Winge DR. Rat liver Cu,Zn-superoxide dismutase. Subcellular location in lysosomes. J Biol Chem 1982;257(15):8945–8952.
8. Halliwell B. Biochemical mechanisms accounting for the toxic action of oxygen on living organisms: the key role of superoxide dismutase. Cell Biol Int Rep 1978;2(2):113–128.
9. Hjalmarsson K, Marklund SL, Engstrom A, Edlund T. Isolation and sequence of complementary DNA encoding human extracellular superoxide dismutase. Proc Natl Acad Sci U S A 1987;84(18):6340–6344.
10. Kazura JW, Meshnick SR. Scavenger enzymes and resistance to oxygen mediated damage in Trichinella spiralis. Mol Biochem Parasitol 1984;10(1):1–10.
11. Kim SH, Kong Y, Kim SI, Kang SY, Cho SY. Immunoblot observation of antigenic protein fractions in Paragonimus westermani reacting with human patients sera. Korean J Parasitol 1988;26(4):239–243.
12. Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 1970;227(5259):680–685.
13. Lee YM, Ayala FJ, Misra HP. Purification and properties of superoxide dismutase from Drosophila melanogaster. J Biol Chem 1981;256(16):8506–8509.
14. Lowry OH, Rosebrough NJ, Farr AL, Randall RJ. Protein measurement with the Folin phenol reagent. J Biol Chem 1951;193(1):265–275.
15. McCord JM, Fridovich I. The reduction of cytochrome c by milk xanthine oxidase. J Biol Chem 1968;243(21):5753–5760.
16. McCord JM, Fridovich I. Superoxide dismutase. An enzymic function for erythrocuprein (hemocuprein). J Biol Chem 1969;244(22):6049–6055.
17. McKerrow JH. Parasite proteases. Exp Parasitol 1989;68(1):111–115.
18. Misra HP, Fridovich I. Inhibition of superoxide dismutases by azide. Arch Biochem Biophys 1978;189(2):317–322.
19. Moon JS, et al. Chung-Ang J Med 1982;7:163–172.
20. Rhoads ML. Trichinella spiralis: identification and purification of superoxide dismutase. Exp Parasitol 1983;56(1):41–54.
21. Roder JC, Helfand SL, Werkmeister J, McGarry R, Beaumont TJ, Duwe A. Oxygen intermediates are triggered early in the cytolytic pathway of human NK cells. Nature 1982;298(5874):569–572.
22. Rotilio G, Bray RC, Fielden EM. A pulse radiolysis study of superoxide dismutase. Biochim Biophys Acta 1972;268(2):605–609.
23. Salin ML, McCord JM. Superoxide dismutases in polymorphonuclear leukocytes. J Clin Invest 1974;54(4):1005–1009.
24. Yost FJ Jr, Fridovich I. An iron-containing superoxide dismutase from Escherichia coli. J Biol Chem 1973;248(14):4905–4908.
Editorial Office
Department of Molecular Parasitology, Samsung Medical Center, School of Medicine, Sungkyunkwan University,
2066 Seobu-ro, Jangan-gu, Suwon 16419, Gyeonggi-do, Korea.
Tel: +82-31-299-6251   FAX: +82-1-299-6269   E-mail: kjp.editor@gmail.com
About |  Browse Articles |  Current Issue |  For Authors and Reviewers
Copyright © 2022 by The Korean Society for Parasitology and Tropical Medicine.     Developed in M2PI