PMID:20727918

Citation	Prosser, GA, Patterson, AV and Ackerley, DF (2010) uvrB gene deletion enhances SOS chromotest sensitivity for nitroreductases that preferentially generate the 4-hydroxylamine metabolite of the anti-cancer prodrug CB1954.J. Biotechnol. 150:190-4
Abstract	CB1954 is an anti-cancer prodrug that can be reduced at either of two nitro groups to form cytotoxic metabolites. We describe here two efficient and previously uncharacterized nitroreductases, YfkO from Bacillus subtilis which reduces CB1954 exclusively at the 4-NO(2) position, and NfsA from Klebsiella pneumoniae which preferentially reduces the 2-NO(2) group. Utilizing these novel enzymes, together with three previously characterized nitroreductases, we demonstrate that the Escherichia coli SOS-chromotest assay can differentially detect the 4-nitro versus 2-nitro reduction products of CB1954 following deletion of the nucleotide excision repair gene uvrB, but not mismatch repair (mutS) or methyltransferase (ada/ogt) genes. These findings may hold significance for identification and selection of nitroreductases for CB1954-mediated gene therapy, particularly when targeting tumors that are deficient in nucleotide excision repair. Moreover, we demonstrate that comparative SOS chromotest analysis in wild type and uvrB mutant strains can be used to determine whether or not nucleotide excision repair plays a significant role in processing DNA damage resulting from activation of different nitroaromatic prodrugs.
Links	PubMed Online version:10.1016/j.jbiotec.2010.08.007
Keywords	Antineoplastic Agents; Aziridines; Bacillus subtilis; Bacterial Proteins; DNA Helicases; Escherichia coli Proteins; Gene Deletion; Kinetics; Klebsiella pneumoniae; Metronidazole; Molecular Conformation; Molecular Sequence Data; Nitrofurazone; Nitroreductases; SOS Response (Genetics)
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Phenotype Annotations

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Phenotype of	Taxon Information	Genotype Information (if known)	Condition Information	OMP ID	OMP Term Name	ECO ID	ECO Term Name	Notes	Status
a mutation or genetic difference within a strain	Taxon: Escherichia coli Strain: K-12 Substrain: NCBI Taxon ID: 83333	Genotype of Reference Strain: uvrB Genotype of Experimental Strain : uvrB(del)	Reference Condition: 50 micromolar CB1954 for 3 hours	OMP:0000172	response to chemical stimulus phenotype	ECO:0000297	LacZ transcript localization evidence	Deletion of the uvrB nucleotide excision repair gene did not affect the SOS chromatest sensitivity; Reference figure 2; NfsA (Ec)= 2-NO2 nitroreductase only, from E. coli Gene deletion enhances SOS chromatest assay sensitivity for nitroreductases which can be used for CB1954-mediated gene therapy.
a mutation or genetic difference within a strain	Taxon: Escherichia coli Strain: K-12 Substrain: NCBI Taxon ID: 83333	Genotype of Reference Strain: uvrB Genotype of Experimental Strain : uvrB(del)	Reference Condition: CB1954 metabolism by NfsA (Kp)	OMP:0000172	response to chemical stimulus phenotype			Deletion of the uvrB nucleotide excision repair gene did not affect the SOS chromatest sensitivity; Reference figure 2; NfsA (Kp)= 2-NO2 nitroreductase only, from K. pneumoniae
a mutation or genetic difference within a strain	Taxon: Escherichia coli Strain: K-12 Substrain: NCBI Taxon ID: 83333	Genotype of Reference Strain: uvrB Genotype of Experimental Strain : uvrB(del)	Reference Condition: CB1954 metabolism by NfsB	OMP:0000172	response to chemical stimulus phenotype			Deletion of the uvrB nucleotide excision repair gene did not affect the SOS chromatest sensitivity; Reference figure 2; NfsB= 2-NO2 or 4-NO2 nitroreductase.
a mutation or genetic difference within a strain	Taxon: Escherichia coli Strain: K-12 Substrain: NCBI Taxon ID: 83333	Genotype of Reference Strain: uvrB Genotype of Experimental Strain : uvrB(del)	Reference Condition: CB1954 metabolism by NemA	OMP:0000172	response to chemical stimulus phenotype			Deletion of the uvrB nucleotide excision repair gene heightened the SOS chromatest sensitivity; Reference figure 2; NemA= 4-NO2 nitroreductase only.
a mutation or genetic difference within a strain	Taxon: Escherichia coli Strain: K-12 Substrain: NCBI Taxon ID: 83333	Genotype of Reference Strain: uvrB Genotype of Experimental Strain : uvrB(del)	Reference Condition: CB1954 metabolism by Yfko (Bs)	OMP:0000172	response to chemical stimulus phenotype			Deletion of the uvrB nucleotide excision repair gene heightened the SOS chromatest sensitivity; Reference figure 2; Yfko (Bs)= 4-NO2 nitroreductase only, from B. licheniformis
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