Investigation of the effects of some water-soluble vitamins on glutathione reductase enzyme purified from bovine liver

Glutathione reductase (GR, EC 1.8.1.7) is an antioxidant enzyme and is involved in the reducing reaction ofoxidized glutathione. Glutathione and reduced glutathione are very crucial for micro- and macroorganisms because of their barrier function against radicals. Therefore, the aim of this article was to purify GR and evaluate the relationship between purified GR and vitamins. GR was purified from bovine liver using 2’,5’-ADP Sepharose 4B column materials. To check the purity of each subunit of the enzyme, SDS-PAGE electrophoresis was performed and each one was 55 kDa. Afterwards, specific activity and purified ratio for the enzyme were calculated as 54 EU/mg × protein and 2700 times, respectively. Vitamins have a regulatory role in organisms and also some, like vitamin B, are coenzymes due to having cofactor effects. In this project, variable concentrations of some water-soluble vitamins, riboflavin, ascorbic acid, nicotinamide, folic acid, and thiamine, were tested. As a result of these kinetic findings, nicotinamide and folic acid increased the activity of glutathione reductase, while thiamine decreased it. Riboflavin and ascorbic acid did not show a stable effect on enzyme activity. Ki and IC50 constants were found as 50.16 ± 5.63 mM and 21.04 mM, respectively, when Lineweaver–Burk and activity % vs. inhibitor concentration graphs were drawn for thiamine. The results of this article illustrate that the behaviors of vitamins could be complicated and vary from one living organism to other; in other words, they are unpredictable.

PDF

___

1. Haldane JBS, William PJ. Enzymes. In: Nelson DL, Cox MM (editors). Lehninger Principles of Biochemistry. New York, NY, USA: W.H. Freeman and Company, 2008, pp. 183-233.
2. Akkuş İ. Serbest Radikaller ve Fizyopatolojik Etkileri. Konya, Turkey: Mimoza Yayınları, 1995 (in Turkish).
3. Çiftci M, Kufrevioglu Öİ, Gundogdu M, Ozmen I. Effects of some antibiotics on enzyme activity of glucose-6- phosphate dehydrogenase from human erythrocytes. Pharmacological Research 2000; (41): 109-113. doi: 10.1006/ phrs.1999.0566
4. Keha E, Küfrevioğlu Öİ. Biyokimya. 3rd ed. Erzurum, Turkey: Aktif Yayınevi, 2004 (in Turkish).
5. Reed DJ, Introduction to Biochemical Toxicology. New York, NY, USA: Wiley-Interscience, 2001.
6. de Lamotte F, Vianey-Liuaud N, Duviau MP, Korehel KJ. Glutathione reductase in wheat grain. 1. Isolation and characterization. Journal of Agricultural and Food Chemistry 2000; (48): 4978-4983. doi: 10.1021/jf0003808
7. Mullineaux P, Enard C, Hellens R, Creissen G. Characterisation of a glutathione reductase gene and its genetic locus from pea (Pisum sativum L.). Planta 1996; (200): 186-194. doi: 10.1007/BF00208308
8. Bülbül M, Erat M. Investigation of the effects of some sulfonamide derivatives on the activities of glucose-6- phosphate dehydrogenase, 6-phosphogluconate dehydrogenase and glutathione reductase from human Erythrocytes. Journal of Enzyme Inhibition and Medicinal Chemistry 2008; (23): 418-423. doi: 10.1080/14756360701546413
9. Fatemi F, Kazemnejad S, Dadkhah A, Rahmati M, Allameh A. Differential effects of acetaminophen and aflatoxin B1 on expression of liver class-p glutathione s-transferase in growing rats. Iranian Journal of Science & Technology, Transaction A 2007; (31): 43-52. doi: 10.22099/ijsts.2007.2315
10. Anthea M, Hopkins J, McLaughlin CW, Johnson S, Warner MQ et al. Human Biology and Health. Englewood Cliffs, NJ, USA: Prentice Hall, 1993.
11. Gözükara EM. Biyokimya. Ankara, Turkey: Nobel Tıp Kitabevleri, 2011 (in Turkish).
12. Bender DA. Nutritional Biochemistry of the Vitamins, 2nd ed. Cambridge, UK: Cambridge University Press, 2003.
13. Altun M, Türkoğlu V, Çelik İ. The effect of some antibiotics on glutathione reductase enzyme purified from liver and erythrocyte of Lake Van pearl mullet. Pharmaceutical Biology 2015; (53): 1647-1652. doi: 10.3109/13880209.2014.997830
14. Sikanyika M, Araga D, McDevitt CA, Maher MJ. The structure and activity of the glutathione reductase from Streptococcus pneumoniae. Acta Crystallographica Section F 2019; (F75): 54-61. doi: 10.1107/S2053230X18016527
15. Kıvanç MR, Türkoğlu V. Investigation of the effects of natural compounds isolated from Arum rupicola var. rupicola on glutathione reductase enzyme purified from bovine liver. Biomedical Chromatography 2019; 2019: e4560. doi: 10.1002/bmc.4560
16. Chanson A, Rock E, Martin JF, Liotard A, Brachet P. Preferential response of glutathione-related enzymes to folate-dependent changes in the redox state of rat liver. European Journal of Nutrition 2007; 46(4): 204-212. doi: 10.1007/s00394-007-0651-1
17. Takechi R, Pallebage-Gamarallage MM, Lam V, Giles C, Mamo JC. Nutraceutical agents with anti-inflammatory properties prevent dietary saturated-fat induced disturbances in blood-brain barrier function in wild-type mice. Journal of Neuroinflammation 2013; (10): 73. doi: 10.1186/1742-2094-10-73
18. Carlberg I, Mannervik B. Purification and characterization of glutathione reductase from calf liver. An improved procedure for affinity chromatography on 2′,5′-ADP-Sepharose 4B. Analytical Biochemistry 1981; (116): 531-536. doi: 10.1016/0003-2697(81)90398-5
19. Bradford MM. A rapid and sensitive method for the quantitation of microgram quantities utilizing the principle of protein dye binding. Analytical Biochemistry 1976; (72): 248-54. doi: 10.1006/abio.1976.9999
20. Arora K, Ahmad R, Srivastava AK. Purification and characterization of glutathione reductase (E.C.1.8.1.7) from bovine filarial worms Setaria cervi. Journal of Parasitic Diseases 2013; (37): 94-104. doi: 10.1007/s12639-012-0138-8
21. Tekman B, Ozdemir H, Senturk M, Ciftci M. Purification and characterization of glutathione reductase from rainbow trout (Oncorhynchus mykiss) liver and inhibition effects of metal ions on enzyme activity. Comparative Biochemistry and Physiology Part C 2008; (148): 117-121. doi: 10.1016/j.cbpc.2008.04.005
22. Estrela JM, Ortega A, Obrador E. Glutathione in cancer biology and therapy. Critical Reviews in Clinical Laboratory Sciences 2006; 43 (2): 143-181. doi: 10.1080/10408360500523878
23. Jones DP. Redox potential of GSH/GSSG couple: assay and biological significance. Methods in Enzymology 2002; (348): 93-112. doi: 10.1016/s0076-6879(02)48630-2
24. Gardiner CS, Reed DJ. Status of glutathione during oxidant-induced oxidative stress in the preimplantation mouse embryo. Biology of Reproduction 1994; (51): 1307-1314. doi: 10.1095/biolreprod51.6.1307
25. Le Trang N, Bhargava KK, Cerami A. Purification of glutathione reductase from gerbil liver in two steps. Analytical Biochemistry 1983; (133): 94-99. doi: 10.1016/0003-2697(83)90226-9
26. Erat M, Sakiroglu H, Ciftci M. Effects of some antibiotics on glutathione reductase from bovine erythrocytes. Veterinarni Medicina 2003; (48): 305-312. doi: 10.17221/5784-VETMED
27. Erat M, Ciftci M. In vitro effects of some antibiotics on glutathione reductase from sheep liver. Journal of Enzyme Inhibition and Medicinal Chemistry 2003; (18): 545–550. doi: 10.1080/14756360310001624957
28. Kapoor G, Bagai U, Banyal HS. Purification and characterization of Plasmodium berghei glutathione reductase. Journal of Parasitic Diseases 2008; (32): 22-29.
29. Ulusu NN, Tandogan B. Purification and kinetic properties of glutathione reductase from bovine liver. Molecular and Cellular Biochemistry 2007; (303): 45-51. doi: 10.1007/s11010-007-9454-1
30. Ulusu G, Erat M, Çiftçi M, Şakiroğlu H, Bakan E. Purification and characterization of glutathione reductase from sheep liver. Turkish Journal of Veterinary and Animal Sciences 2005; (29): 1109-1117.
31. Taşer P, Çiftçi M. Purification and characterization of glutathione reductase from turkey liver. Turkish Journal of Veterinary and Animal Sciences 2012; (36): 546-553. doi: 10.3906/vet-1103-5
32. Wen LM, Jiang WD, Liu Y, Wu P, Zhao J et al. Evaluation the effect of thiamin deficiency on intestinal immunity of young grass carp (Ctenopharyngodon idella). Fish & Shellfish Immunology 2015; 46 (2): 501-515. doi: 10.1016/j.fsi.2015.07.001
33. Li SQ, Feng L, Jiang WD, Liu Y, Jiang J et al. Deficiency of dietary niacin impaired gill immunity and antioxidant capacity, and changes its tight junction proteins via regulating NF-κB, TOR, Nrf2 and MLCK signaling pathways in young grass carp (Ctenopharyngodon idella). Fish & Shellfish Immunology 2016; (55): 212-222. doi: 10.1016/j.fsi.2016.05.005
34. Adelekan DA, Thurnham DI. Glutathione peroxidase (EC 1.11.1.9) and superoxide dismutase (EC 1.15.1.1) activities in riboflavin-deficient rats infected with Plasmodium berghei malaria. British Journal of Nutrition 1998; (79): 305-309. doi: 10.1079/BJN19980048
35. Bates J. Glutathione and related indices in rat lenses, liver and red cells during riboflavin deficiency and its correction. Experimental Eye Research 1991; (53): 123-130. doi: 10.1016/0014-4835(91)90154-7
36. Zhao XL, Li YK, Cao SJ, Hu JH, Wang WH et al. Protective effects of ascorbic acid and vitamin E on antioxidant enzyme activity of freeze-thawed semen of Qinchuan bulls. Genetics and Molecular Research 2015; 14 (1): 2572- 2581. doi: 10.4238/2015.March.30.16
37. Shokrzadeh M, Shobi S, Attar H, Shayegan S, Payam SS et al. Effect of vitamins A, E and C on liver enzyme activity in rats exposed to organophosphate pesticide diazinon. Pakistan Journal of Biological Sciences 2012; (19): 936-941. doi: 10.3923/pjbs.2012.936.941
38. Dwivedi VK, Chandra M, Misra PC, Misra MK. Effect of vitamin E on platelet enzymatic anti-oxidants in the patients of myocardial infarction. Indian Journal of Clinical Biochemistry 2005; 20 (1): 21-25. doi: 10.1007/BF02893036
39. Jeya Sheela P, Haniffa MA, Vigneshwaran R, Kavitha K. Effect of vitamins on digestive enzyme activities and growth performance of striped murrel Channa striatus. Journal of Research in Biology 2017; 7 (8): 2377-2385.