The Effects of Amoxicillin and Vancomycin Hydrochloride Hydrate on Glutathione S-Transferase Enzyme Activity: An in vitro study

Abstract

Glutathione S-transferase (GST) enzymes detoxify a broad spectrum of xenobiotics, including

chemotherapeutic drugs, environmental carcinogens, and endogenous molecules. Glutathione transferases catalyze

the conversion of some metabolites into less toxic substances. Phase II reactions, which often result in detoxification,

are conjugation reactions of glutathione through GST enzymes that protect the organism from the attacks of highly

reactive electrophilic substances. Since the GST enzyme can metabolize toxic exogenous compounds, it has been

extensively investigated in mammals. In this study, the GST enzyme was purified in human erythrocytes with a

specific activity of 5.56 EU mg-1 protein and a yield of 2316.0 fold with 65% efficienc. SDS-polyacrylamide gel

electrophoresis was performed to check the purity of the purified enzyme and the molecular mass was determined

to be about 26 kDa. The IC50 and Ki values for amoxycillin and vancomycin hydrochloride drugs on the purified

GST enzyme were calculated and the type of inhibition was determined. As a result of the inhibition studies, we

observed that these antibiotics inhibited the GST enzyme at low doses.

Keywords

• Amoxicillin,vancomycin hydrochloride hydrate,glutathione s-transferase

Glutatyon S-Transferaz Enzim Aktivitesi Üzerine Amoksilin ve Vankomisin Hidroklorid Hidratın Etkisi: Bir in vitro çalışma

Abstract

Glutatyon S-transferaz enzimleri kemoterapik ilaçlar, çevresel karsinojenler, endojen moleküller olmak

üzere ksenobiyotiklerin geniş bir spektrumunu detoksifiye ederler. Glutatyon transferazlar, bazı metabolitleri daha

az toksik olan maddelere dönüşmesini katalize eder. Çoğunlukla detoksifikasyonla sonuçlanan faz II reaksiyonları

arasında organizmayı son derece reaktif elektrofilik maddelerin ataklarından koruyan GST enzimleri aracılığıyla

gerçekleşen glutatyonun konjugasyon reaksiyonlarıdır. GST enzimi toksik ekzojen bileşikleri metabolize

edebildiğinden dolayı ayrıntılı biçimde memelilerde araştırılmıştır. Bu çalışmada GST enzimi insan eritrositlerinde

5.56 EU mg-1 protein spesifik aktiviteyle ve 65 %verimle 2316.0 kat saflaştırıldı. Saflaştırılan enzimin saflığını

kontrol etmek için SDS-poliakrilamid jel elektroforezi yapıldı ve molekül kütlesi yaklaşık 26 kDa olarak belirlendi.

Saflaştırılan GST enzimi üzerine inhibisyon etkisi gösteren amoksisilin ve vankomisin hidroklorür için IC50

değerleri ve Ki sabitleri hesaplanarak inhibisyon tipleri belirlendi. İnhibisyon çalışmalarının sonucu olarak, bu

antibiyotiklerin düşük dozlarda GST enzimini inhibe ettiği gözlendi.

Keywords

• Amoksisilin,vankomisin hidroklorür,glutatyon s- transferaz

References

1. Alim Z, Beydemir S, 2016. Some Anticancer Agents Act on Human Serum Paraoxonase-1 to Reduce Its Activity. Chem Biol Drug Des., 88(2):188-96.
2. Alim, Z, Kilinç N, Şengül B, Beydemir Ş, Işgör MM, 2015. Some Anti-Inflammatory Agents Inhibit Esterase Activities of Human Carbonic Anhydrase Isoforms I and II: An In Vitro Study. Chemical Biology & Drug Design,. 86(4): 857-863.
3. Bradford, M.M., 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry, 72, 248-251.
4. Çiftçi M, Türkoğlu V, Çoban TA, 2007. Effects of some drugs on hepatic glucose 6-phosphate dehydrogenase activity in Lake Van Fish (Chalcalburnus Tarischii Pallas, 1811). Journal of Hazardous Materials, 143:415-418.
5. Çomaklı V, Çiftçi M., Küfrevioğlu Öİ, 2011. Purification of Glutathione S-Transferase Enzyme from Rainbow Trout Erythrocytes and Examination of the Effects of Certain Antibiotics on Enzyme Activity. Hacettepe J. Biol. & Chem.,39 (4):413–419.
6. Demirdağ R, Yerlikaya E, Kufrevioglu Öİ, Gündoğdu C, 2012. Purification of glutathione S -transferase isoenzymes from tumour and nontumour human stomach and inhibitory effects of some heavy metals on enzymes activities. Journal of Enzyme Inhibition and Medicinal Chemistry. DOI: 10.3109/14756366.2012.694878
7. Erat M, Sakiroglu H, 2013. "The effect of some antineoplastic agents on glutathione S-transferase from human erythrocytes." J Enzyme Inhib Med Chem, 28(4): 711-716.
8. Guengerich FP, 1990. Enzymatic oxidation of xenobiotic chemicals. CRC Crit. Rev. Biochem. Mol. Biol., 25: 97-153.

9. Gülçin İ, Scozzafava A, Claudiu TS, Akıncıoğlu H, Koksal Z, Turkan F, Alwasel S, 2016. The effect of caffeic acid phenethyl ester (CAPE) on metabolic enzymes including acetylcholinesterase, butyrylcholinesterase, glutathione S-transferase, lactoperoxidase, and carbonic anhydrase isoenzymes I, II, IX, and XII. Journal of Enzyme Inhibition and Medicinal Chemistry, 31(6):1095-1101.
10. Gülçin İ, Scozzafava A, Supuran CT, Koksal Z, Turkan F, Çetinkaya S, Bingöl Z, Huyut Z, Alwasel SH, 2016. Rosmarinic acid inhibits some metabolic enzymes including glutathione S-transferase, lactoperoxidase, acetylcholinesterase, butyrylcholinesterase and carbonic anhydrase isoenzymes. J Enzyme Inhib Med Chem. Dec;31(6):1698-702.
11. Güvercin, S., Erat, M., Şakiroğlu, H., 2008. Determination of Some Kinetic and Characteristic Properties of Glutathione S-transferase from Bovine Erythrocytes. Protein & Peptide Letters, 15, 6-12.
12. Halliwell B, 2009. The wanderings of a free radical. Free Radic Biol Med, 46(5):531-42.
13. Hardman JG, Limbird LE, 2001. Goodman & Gilman’s The Pharmacological Basis of Therapeutics. 10th Edition, New York: McGraw-Hill Professional Publishing.
14. Hatey F, Tosser-Klopp G, Clouscard-Martinato C, Mulsant P, Gasser F, 1998. Expressed sequence tags for genes : A review. Genet. Sel. Evol. 30:521-541. Hayes JD, Flanagan JU, Jowsey IR, 2005. Glutathione transferases. Annu Rev Pharmacol Toxicol, 45: 51-88.
15. Hee-Joong P, Hyun-Young C, Kwang-Hoon K, 2005. Purification and Biochemical Properties of Glutathione S-Transferase from Lactuca sativa. Journal of Biochemistry and Molecular Biology, 38( 2):232-237.
16. Hinchman CA, Rebbeor JF, Ballatori N, 1998. Efficient hepatic uptake and concentrative biliary excretion of a mercapturic acid. Am J Physiol, 275:612– 619.
17. Laemmli UK, 1970. Cleavage of Structural Proteins during Assembly of Head of Bacteriophage-T4. Nature, 227(5259): 680-691.
18. Liu J, Wang Z, Li F, Gao J, Wang L, Huang G, 2015. Liposomes for systematic delivery of vancomycin hydrochloride to decrease nephrotoxicity: Characterization and evaluation. Asian Journal of Pharmaceutical Sciences, 10:212-222.
19. Lizuka M, Inoue Y, Murata K, Kımura A, 1989. Purification and Some Properties of Glutathione S-Transferase from Escherichia coli B. Journal Of Bacterıology, 171(11): 6039-6042
20. Mann, M. A. (1996) A shortcut to interesting human genes : Peptide sequence tags, expressed-sequence tags and computers. Trends Biochem. Sci. 21, 494±495
21. Mazzetti AP, Fiorile MC, Primavera A, Lo Bello M, 2015. Glutathione transferases and neurodegenerative diseases. Neurochem Int,. 82:10-8.
22. Novoa – Valinas M.C, Melgar – Riol MJ, Perez - Lopez M, 2004. Hepatic glutathione S transferases from lamprey (Petromyzom marinus): purification and characterization Biochemical Systematics and Ecology, 32:169–178.
23. Riol MJM, Valiñas MCN, Fernández MAG, López MP, 2001. Glutathione S-transferases from rainbow trout liver and freshly isolated hepatocytes: purification and characterization. Comparative Biochemistry and Physiology Part C: Toxicology & Pharmacology, 128(2) : 227-235.
24. Rybak MJ. The pharmacokinetic and pharmacodynamic properties of vancomycin. Clin Infect Dis 2006;42:S35e9.
25. Sheehan D, Meade G, Vivienne MF, Catriona AD, 2001. Structure, function and evolution of glutathione transferases implications for classification of non-mammalian members of an ancient enzyme superfamily. Biochem. J., 360:1-16.
26. Sojib Bin Zaman, Muhammed Awlad Hussain, Rachel Nye, Varshil Mehta, Kazi Taib Mamun, Naznin Hossain, 2017. A Review on Antibiotic Resistance: Alarm Bells are Ringing, 9(6):1403.
27. Stevens JL, Jones DP, 1989. The mercapturic acid pathway: biosynthesis, intermediary metabolism, and physiological disposition. In: Glutathione: Chemical, Biochemical, and Medical Aspects. Part B (Dolphin D, Avramovic O, and Poulson R eds.) John Wiley and Sons, New York. 45–84.
28. Sutherland R, Croydon EAP, Rolinson GN, 1972. Amoxycillin: a new semisynthetic penicillin. BMJ, 3 : 13 -6.
29. Türkan F, Balcı N, Şakiroğlu H, 2014. Purification and characterization of glutathione s-transferase from laurel fruit (laurocerasus officinalis roem.) And inhibition the effects of some pesticides on enzyme activity. Muş Alparslan Üniversitesi fen bilimleri dergisi, 2(2):280-288.
30. Young PR, Brıedıs AV, 1989. Purification and kinetic mechanism of the majör glutathione S-transferase from bovine brain. Biochem. J., 257:541-548.
31. Zaman SB, Hussain MA , Nye R , Mehta V , Mamun KT , Hossain N , 2017. A Review on Antibiotic Resistance: Alarm Bells are Ringing, 9(6):1403.