Kısmen saflaştırılan sığır karaciğer arginazının kinetik özellikleri ve bazı kimyasal bileşiklerin enzim aktivitesine etkilerinin araştırılması

Bu çalışmada, olası tıbbi uygulamalara muhtemel bir hedef olarak sığır karaciğerinden arginaz enzimi kısmen saflaştırılarak, bazı kinetik özellikleri incelenmiştir. Saflaştırılan enzimin önce arginin substratı için Michaelis-Menten ve Lineweaver-Burk grafikleri çizilerek Km değeri 4.5 mM olarak hesaplanmıştır. Daha sonra sodyum siyanid, prolin, çinko klorür, sodyum dietilditiyokarbamat, histidin ve gentamisin' in enzim aktivitesi üzerindeki etkileri araştırılmıştır. Sodyum siyanidin enzim üzerinde karışık, prolin ve ZnCl2' nin yarışmalı inhibisyon meydana getirdiği, sodyum dietilditiyokarbamat, histidin ve gentamisinin enzim aktivitesine denenen konsantrasyonlarda gözlenebilen bir etkisi olmadığı belirlenmiştir.

Investigation of the kinetic properties of partially purified bovine liver arginase and the effect of some chemical compounds on enzyme activity

In this study, arginase was partially purified from bovine liver and some of its kinetic properties were investigated for evaluation of possible medical applications. ,The Km value for its natural substrate arginine was determined as 4.5 mM by plotting the Michaelis-Menten and Lineweaver-Burk graphics. Then, effect of sodium cyanide, proline, zinc chloride, sodium diethyldithiocarbamate, histidine and gentamycine on enzyme activities were investigated. Sodium cyanide was found to be a mixed type inhibitor of the enzyme whereas proline and ZnCl2 were competitive inhibitors. Addition of sodium-diethyldithiocarbamate, histidine and gentamycine caused no observable effect on the enzyme activity within the experimental concentrations.

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1. Aminlari M. Novel colorimetric method for assaying argina-se activity. Clin Biochem. 25:431-436, 1992.
2. Halifeoğlu I. insan karaciğer, eritrosit ve uterus doku argina-zının kinetik özellikleri. Doktara Tezi. Fırat Üniversitesi Sağlık Bilimleri Enstitüsü Müdürlüğü.1993
3. Crody Ö., Dizikes Gj., Cederbaun SD. Human arginase iso-enzymes. Isoenzymes Curr Top Biol Med Res,13:181-214,1987
4. Bauscur L., Cabello J., Veliz M., Gonzales A. Molecular Forms of human liver arginase Bochem. Biophys. Aç-ta,! 28, 149-54, 1966.
5. VVİlliams-Ashman HG., Canellakis ZN. Polyamines in mam-malian Biology and Medicine. The University of Chica-go Press. Perspective in Biology and Medicine. 421-451, 1979.
6. Gotoh T, Sonoki T, Nagasaki A, Terada K, Takiguchi M, Mo-ri M. Molecular cloning for cDNA for non-hepatic mi-tochondrial arginase (arginase II) and comparison of its induction vvith nitric oxide synthase in a murine mac-rophage-like celi line. Febbs Lett. 1996;395:119-22
7. Leu SY., Wang SR.,Clinical significance of arginase in colo-rectal cancer. Cancer, 70:5, 733-736, 1992.
8. Koza RA., Megosh LC., Palmieri M., O'Brien TG. Constitu-tively elevated levels ofornithine and polyamines in mo-use epidermal papillomas. Carcinogenesis, 12:9, 1619-1625, 1991.
9. Trapeznikova SS., Navasardiants DG., Levchenko Ll., Com-parative study of the activity of arginase isoenzymes in brain tumors of humans and experimental animals. Vopr Med Khim. 32:6, 29-34. 1986.
10. Harris BE., Pretlovv TP., Bradley EL., Arginase activity in prosthatic fluid of patients vvith benign prothatic hyperplasia and prosthatic carcinoma. Cancer Res. 43:6, 3008-3012, 1983.
11. Wu CW., Chi CW., Tsay SH., The effects of argkıase on ne-oplasm I. The role of arginase in the immunesuppressi-ve effects of extract from gastric cancer. Chung Hua Min Kuo VVei Sheng Wu Chi Mien I Hsueh Cha Chih. 20:4, 279-289, 1987.
12. Tietz NW. Clinical Guide to Laboratory Tests. Ed. NVV. Ti-etz. WB Saunders Company, Philadelphia. 1990.
13. Porembska Z., Kedra M., Early diagnosis of myocardial in-farction by arginase activity determination. Clin. Chem. Açta, 60:3, 355-361, 1975.
14. Terheggen HG, Lavinha F, Colombo JP. Familial hyperargi-ninemia. J. Genet. Hum. 1972;20:69-84
15. Harrell D., Sokolovvsky M., Beef liver arginase. Isolation and Molecular Properties. Eur J. Biochem. 25, 102-108, 1972.
16. Lovvry O, Rosenbraugh N, Farr L, Rondall R. Protein meau-surement vvith the folin-phenol reagent. J.Bİ-ol.Chem.1951;183:265-275
17. Chinard FP. Photometric estimation of proline and ornithi-ne. J.Bİol Chem. 199:91-95,1952.
18. Spector EB., Jenkinson CP., Grigor., MR., Kern RM., Ceder-baum SD., Subcellular location and differential anti-body specificityof arginase in tissue culture and vvhole animals. Int J Dev Neiroscience. 12:4, 337-42, 1994.
19. Akbay A. Sığır karaciğer arginazının kısmen saflaştırılması ve bazı kinetik özelliklerinin araştırılması. Ankara Üni-versitesi Tıp Fakültesi Biyokimya Anabilim Dalı, Uz-manlık Tezi. Ankara,1997
20. Musznka G, Severina OL, Lobyreva LW. Characteristics of arginases from plant, ureotelic and uricotelic organisms. Açta Biochem Pol.1972;19:235-242
21. Ikemoto M., Tabata M.,Murachi T. Purification and proper-ties of human erythrocyte arginase. Ann. Clin Biochem. 26:4, 547-553 1989.
22. ikemoto M, Tabata M, Miyake T, Kono T, Mori M, Totani M, Murachi T. Expression of human liver arginase in Esche-rischia coli. Purification and properties of the product. Biochem J. 1990 ; 270 : 697-703
23. Savoca KV., Abuchovvski A., van Es T., Davis FF.,Palczak NC. Preparation of a non immunogenic arginase by the covalent attachment of polyethylene glycol. Biochim Biophys Açta, 578:1, 47-53, 1979.
24. Segel I. Enzyme Kinetics. A VVİley Interscience Publication, Nevv York, 1975.
25. Green SM., Ginsburg A., Levvis MS., Hensley P. Roles of metal ions in the maintenance of the tertiary and quar-ter narystructure of arginase from Saccharomyces cere-visiae. J.Bİol. Chem. 266:32, 21474-81,1991