Molecular evolution of Theta-class glutathione transferase for enhanced activity with the anticancer drug 1,3-bis-(2-chloroethyl)-1-nitrosourea and other alkylating agents
2010 (English)In: Archives of Biochemistry and Biophysics, ISSN 0003-9861, E-ISSN 1096-0384, Vol. 497, no 1-2, 28-34 p.Article in journal (Refereed) Published
Glutathione transferase (GST) displaying enhanced activity with the cytostatic drug 1,3-bis-(2-chloroethyl)-1-nitrosourea (BCNU) and structurally related alkylating agents was obtained by molecular evolution. Mutant libraries created by recursive recombination of cDNA coding for human and rodent Theta-class GSTs were heterologously expressed in Escherichia coli and screened with the surrogate substrate 4-nitrophenethyl bromide (NPB) for enhanced alkyltransferase activity. A mutant with a 70-fold increased catalytic efficiency with NPB, compared to human GST T1-1, was isolated. The efficiency in degrading BCNU had improved 170-fold, significantly more than with the model substrate NPB. The enhanced catalytic activity of the mutant GST was also 2-fold higher with BCNU than wild-type mouse GST T1-1, which is 80-fold more efficient than wild-type human GST T1-1. We propose that GSTs catalyzing inactivation of anticancer drugs may find clinical use in protecting sensitive normal tissues to toxic side-effects in treated patients, and as selectable markers in gene therapy.
Place, publisher, year, edition, pages
2010. Vol. 497, no 1-2, 28-34 p.
Glutathione transferase (GST), 1, 3-Bis-(2-chloroethyl)-1-nitrosourea (BCNU) Chloroethylnitrosoureas (CENUs), Directed evolution, Alkyltransferase, Alkylating agents
IdentifiersURN: urn:nbn:se:uu:diva-119509DOI: 10.1016/j.abb.2010.03.001ISI: 000277537800004OAI: oai:DiVA.org:uu-119509DiVA: diva2:300350