Journal
BIOORGANIC & MEDICINAL CHEMISTRY
Volume 20, Issue 11, Pages 3540-3550Publisher
PERGAMON-ELSEVIER SCIENCE LTD
DOI: 10.1016/j.bmc.2012.04.004
Keywords
Nitroreductase; Cold active enzyme; Prodrug activation; CB1954; Staphylococcus saprophyticus; Nitrofurazone
Funding
- Scientific and Technological Research Council of Turkey (TUBITAK) [110T754]
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A set of PCR primers based on the genome sequence were used to clone a gene encoding a hypothetical nitroreductases (named as Ssap-NtrB) from uropathogenic staphylococcus, Staphylococcus saprophyticus strain ATCC 15305, an oxygen insensitive flavoenzyme. Activity studies of the translation product revealed that the nitroreductase catalyses two electron reduction of a nitroaromatic drug of nitrofurazone (NFZ), cancer prodrugs of CB1954 and SN23862 at optimum temperature of 20 degrees C together with retaining its maximum activity considerably at 3 degrees C. The required electrons for such reduction could be supplied by either NADH or NADPH with a small preference for the latter. The gene was engineered for heterologous expression in Escherichia coli, and conditions were found in which the enzyme was produced in a mostly soluble form. The recombinant enzyme was purified to homogeneity and physical, spectral and catalytical properties were determined. The findings lead us to propose that Ssap-NtrB represents a novel nitro reductase with an unusual cold active property, which has not been described previously for prodrug activating enzymes of nitroreductases. (C) 2012 Elsevier Ltd. All rights reserved.
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