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A novel method for screening the glutathione transferase inhibitors.

Wang Z, Jin L, Wegrzyn G, Wegrzyn A - BMC Biochem. (2009)

Bottom Line: It has been found that the resistance to many anticancer chemotherapeutics is directly correlated with the over-expression of GSTs.Our results showed that two fragments of GST, named F1 peptide (GYWKIKGLV) and F2 peptide (KWRNKKFELGLEFPNL), can significantly inhibit the GST activity.When these two fragments were compared with several known potent GST inhibitors, the order of inhibition efficiency (measured in reactions with 2,4-dinitrochlorobenzene (CDNB) and glutathione as substrates) was determined as follows: tannic acid > cibacron blue > F2 peptide > hematin > F1 peptide > ethacrynic acid.

View Article: PubMed Central - HTML - PubMed

Affiliation: CAS-MPG Partner Institute for Computational Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 200031, Shanghai, PR China. zjwang@shmu.edu.cn

ABSTRACT

Background: Glutathione transferases (GSTs) belong to the family of Phase II detoxification enzymes. GSTs catalyze the conjugation of glutathione to different endogenous and exogenous electrophilic compounds. Over-expression of GSTs was demonstrated in a number of different human cancer cells. It has been found that the resistance to many anticancer chemotherapeutics is directly correlated with the over-expression of GSTs. Therefore, it appears to be important to find new GST inhibitors to prevent the resistance of cells to anticancer drugs. In order to search for glutathione transferase (GST) inhibitors, a novel method was designed.

Results: Our results showed that two fragments of GST, named F1 peptide (GYWKIKGLV) and F2 peptide (KWRNKKFELGLEFPNL), can significantly inhibit the GST activity. When these two fragments were compared with several known potent GST inhibitors, the order of inhibition efficiency (measured in reactions with 2,4-dinitrochlorobenzene (CDNB) and glutathione as substrates) was determined as follows: tannic acid > cibacron blue > F2 peptide > hematin > F1 peptide > ethacrynic acid. Moreover, the F1 peptide appeared to be a noncompetitive inhibitor of the GST-catalyzed reaction, while the F2 peptide was determined as a competitive inhibitor of this reaction.

Conclusion: It appears that the F2 peptide can be used as a new potent specific GST inhibitor. It is proposed that the novel method, described in this report, might be useful for screening the inhibitors of not only GST but also other enzymes.

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Lineweaver-Burk plot of the GST activity with varying glutathione (GSH) concentrations. A): The activity was measured with 1 mM CDNB, and different concentrations of GSH, and the F1 peptide. B): The activity was measured with 1 mM CDNB, and different concentrations of GSH, and the F2 peptide.
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Figure 5: Lineweaver-Burk plot of the GST activity with varying glutathione (GSH) concentrations. A): The activity was measured with 1 mM CDNB, and different concentrations of GSH, and the F1 peptide. B): The activity was measured with 1 mM CDNB, and different concentrations of GSH, and the F2 peptide.

Mentions: The Lineweaver-Burk plot for glutathione as the variable peptide concentration was applied to determine the types of inhibition caused by F1 and F2 peptides. The plot provides a useful graphical method for analysis of the Michaelis-Menten equation. The effects of the peptide on GST-catalyzed reaction kinetics were determined by analysis of apparent Vmax, inhibitor constant Ki and [I]/Ki values. Our results indicated that the F1 peptide exerted a noncompetitive inhibition in the GST-catalyzed reaction with the changing glutathione and F1 peptide concentrations (Vmax decreased while Km remained unchanged) (Fig. 5A). However, the F2 peptide exerted a competitive inhibition in this reaction, with the changing glutathione and F2 peptide concentrations (Km decreased while Vmax remained unchanged) (Fig. 5B).


A novel method for screening the glutathione transferase inhibitors.

Wang Z, Jin L, Wegrzyn G, Wegrzyn A - BMC Biochem. (2009)

Lineweaver-Burk plot of the GST activity with varying glutathione (GSH) concentrations. A): The activity was measured with 1 mM CDNB, and different concentrations of GSH, and the F1 peptide. B): The activity was measured with 1 mM CDNB, and different concentrations of GSH, and the F2 peptide.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2662877&req=5

Figure 5: Lineweaver-Burk plot of the GST activity with varying glutathione (GSH) concentrations. A): The activity was measured with 1 mM CDNB, and different concentrations of GSH, and the F1 peptide. B): The activity was measured with 1 mM CDNB, and different concentrations of GSH, and the F2 peptide.
Mentions: The Lineweaver-Burk plot for glutathione as the variable peptide concentration was applied to determine the types of inhibition caused by F1 and F2 peptides. The plot provides a useful graphical method for analysis of the Michaelis-Menten equation. The effects of the peptide on GST-catalyzed reaction kinetics were determined by analysis of apparent Vmax, inhibitor constant Ki and [I]/Ki values. Our results indicated that the F1 peptide exerted a noncompetitive inhibition in the GST-catalyzed reaction with the changing glutathione and F1 peptide concentrations (Vmax decreased while Km remained unchanged) (Fig. 5A). However, the F2 peptide exerted a competitive inhibition in this reaction, with the changing glutathione and F2 peptide concentrations (Km decreased while Vmax remained unchanged) (Fig. 5B).

Bottom Line: It has been found that the resistance to many anticancer chemotherapeutics is directly correlated with the over-expression of GSTs.Our results showed that two fragments of GST, named F1 peptide (GYWKIKGLV) and F2 peptide (KWRNKKFELGLEFPNL), can significantly inhibit the GST activity.When these two fragments were compared with several known potent GST inhibitors, the order of inhibition efficiency (measured in reactions with 2,4-dinitrochlorobenzene (CDNB) and glutathione as substrates) was determined as follows: tannic acid > cibacron blue > F2 peptide > hematin > F1 peptide > ethacrynic acid.

View Article: PubMed Central - HTML - PubMed

Affiliation: CAS-MPG Partner Institute for Computational Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 200031, Shanghai, PR China. zjwang@shmu.edu.cn

ABSTRACT

Background: Glutathione transferases (GSTs) belong to the family of Phase II detoxification enzymes. GSTs catalyze the conjugation of glutathione to different endogenous and exogenous electrophilic compounds. Over-expression of GSTs was demonstrated in a number of different human cancer cells. It has been found that the resistance to many anticancer chemotherapeutics is directly correlated with the over-expression of GSTs. Therefore, it appears to be important to find new GST inhibitors to prevent the resistance of cells to anticancer drugs. In order to search for glutathione transferase (GST) inhibitors, a novel method was designed.

Results: Our results showed that two fragments of GST, named F1 peptide (GYWKIKGLV) and F2 peptide (KWRNKKFELGLEFPNL), can significantly inhibit the GST activity. When these two fragments were compared with several known potent GST inhibitors, the order of inhibition efficiency (measured in reactions with 2,4-dinitrochlorobenzene (CDNB) and glutathione as substrates) was determined as follows: tannic acid > cibacron blue > F2 peptide > hematin > F1 peptide > ethacrynic acid. Moreover, the F1 peptide appeared to be a noncompetitive inhibitor of the GST-catalyzed reaction, while the F2 peptide was determined as a competitive inhibitor of this reaction.

Conclusion: It appears that the F2 peptide can be used as a new potent specific GST inhibitor. It is proposed that the novel method, described in this report, might be useful for screening the inhibitors of not only GST but also other enzymes.

Show MeSH
Related in: MedlinePlus