Yazar "Erdogan, Orhan" seçeneğine göre listele

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    Öğe
    Expression of hCA IX isoenzyme by using sumo fusion partner and examining the effects of antitumor drugs
    (2015) Yerlikaya, Emrah; Erdogan, Orhan; Demirdag, Ramazan; Senturk, Murat; Kufrevioglu, Omer Irfan
    Objective: In this study, investigating the effects of inhibition of the enzyme activity of some antitumor drugs and the Cancer-Related Human Carbonic Anhydrase IX (hCA IX) isoenzyme expressing as a SUMO fusion protein in an Escherichia coli expression system were aimed. Methods: hCA IX isoenzyme was expressed using SUMO fusion technology. The fusion protein was expressed in a totally soluble form and the expression was verified by SDS-PAGE analysis. Affinity chromatography was used in the purification processes. The effects of certain antitumor drugs on enzyme activity were investigated in vitro conditions by using esterase activity. IC50 values of drugs showing the inhibitory effect were calculated. Inhibition types and Ki values for antitumor drugs, which inhibit the enzyme, were determined by separately plotting Lineweaver- Burk plots. Results: The molecular weight of the fusion protein was approximately 85kDa. The optimal induction concentration of IPTG and the growth temperature were found to be 1.0mM and 30oC. The fusion protein was purified at approximately 3.07-fold with a yield of 92.58%, and a specific activity of 43707EU/mg proteins by nickel nitrilo-triacetic acid resin chromatography.
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    Inhibition Effects of Some Sulfonamides on Carbonic Anhydrase from European seabass (Dicentrachus labrax)
    (2011) Erdem, Deryanur; Yerlikaya, Emrah; Ceyhun, Saltuk Bugrahan; Demirdag, Ramazan; Senturk, Murat; Erdogan, Orhan; Kufrevioglu, Omer Irfan
    Alpha-carbonic anhydrase (EC: 4.2.1.1; CA) was purified from European seabass gill and liver. The purification procedure was composed of preparation of homogenate (or hemolysate) and affinity chromatography on Sepharose 4B-tyrosine-sulfanilamide. Some sulfonamides exhibited in vitro inhibitory effects on the enzyme activity. Ki constants and IC50 values for these drugs were determined by Lineweaver-Burk graphs and plotting activity % vs. [I], respectively. IC50 values of sulfanilamide, mafenide, acetazolamide, 2-amino-1,3,5-tiyadiazol-5 sulfonamide were 980 ??M, 142 ??M, 20 ??M and 34 ??M for gill carbonic anhydrase (GCA), 126 ??M, 23 ??M, 14 ??M and 2.58 ??M for liver carbonic anhydrase (LCA), respectively. Some sulfonamides exhibited much stronger inhibitory activity against GCA and LCA at low micromolar concentrations with the Ki values ranging from 0.21 to 76.0 ??M as compared with other CAs.
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    Öğe
    Purification and characterization of carbonic anhydrase from the teleost fish Dicentrarchus labrax (European seabass) liver and toxicological effects of metals on enzyme activity
    (2011) Ceyhun, Saltuk Bugrahan; Senturk, Murat; Yerlikaya, Emrah; Erdogan, Orhan; Kufrevioglu, Omer Irfan; Ekinci, Deniz
    Carbonic anhydrase (EC 4.2.1.1; CA) was purified and characterized from the liver of the teleost fish Dicentrarchus labrax (European seabass) for the first time. The purification procedure consisted of a single step affinity chromatography on Sepharose 4B-tyrosinesulfanilamide. The enzyme was purified 78.8-fold with a yield of 46%, and a specific activity of 751.72U/mg proteins. It has an optimum pH at 7.5; an optimum temperature at 25 ?C; an optimum ionic strength at 10mM and a stable pH at 8.5. The kinetic parameters of this enzyme were determined for its esterase activity, with 4-nitrophenyl acetate (NPA) as substrate and the purified enzyme had an apparent KM and Vmax values of 0.44mM and 0.249 mol x min-1, respectively. The following metals, Al+3, Cu+2, Pb+2, Co+3, Ag+1, Zn+2 and Hg+2 showed inhibitory effects on the enzyme. Al+3 and Cu+2 exhibited the strongest inhibitory action. Pb+2 was moderate inhibitor, whereas other metals showed weaker actions. All tested metals inhibited the enzyme in a competitive manner. Our findings indicate that these metals inhibit the fish enzyme in a similar manner to other -CAs from mammals investigated earlier, but the susceptibility to various metals differ between the fish and mammalian enzymes. Our results also demonstrate that these metals might be dangerous at low micromolar concentrations for fish CA enzymes.