Evaluation of NfsA-like nitroreductases from <Emphasis Type="Italic">Neisseria meningitidis</Emphasis> and <Emphasis Type="Italic">Bartonella henselae</Emphasis> for enzyme-prodrug therapy,targeted cellular ablation,and dinitrotoluene bioremediation

Objectives

To characterize the activities of two candidate nitroreductases, Neisseria meningitidis NfsA (NfsA_Nm) and Bartonella henselae (PnbA_Bh), with the nitro-prodrugs, CB1954 and metronidazole, and the environmental pollutants 2,4- and 2,6-dinitrotoluene.

Results

NfsA_Nm and PnbA_Bh were evaluated in Escherichia coli over-expression assays and as His₆-tagged proteins in vitro. With the anti-cancer prodrug CB1954, both enzymes were more effective than the canonical O₂-insensitive nitroreductase E. coli NfsB (NfsB_Ec), NfsA_Nm exhibiting comparable levels of activity to the leading nitroreductase candidate E. coli NfsA (NfsA_Ec). NfsA_Nm is also the first NfsA-family nitroreductase shown to produce a substantial proportion of 4-hydroxylamine end-product. NfsA_Nm and PnbA_Bh were again more efficient than NfsB_Ec at aerobic activation of metronidazole to a cytotoxic form, with NfsA_Nm appearing a promising candidate for improving zebrafish-targeted cell ablation models. NfsA_Nm was also more active than either NfsA_Ec or NfsB_Ec with 2,4- or 2,6-dinitrotoluene substrates, whereas PnbA_Bh was relatively inefficient with either substrate.

Conclusions

NfsA_Nm is a promising new nitroreductase candidate for several diverse biotechnological applications.