Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.577812
Title: Structural comparisons of the E. coli prodrug-activating nitroreductase enzymes, NfsA and NfsB
Author: Day, Martin Alan
Awarding Body: University of Birmingham
Current Institution: University of Birmingham
Date of Award: 2013
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Abstract:
The major and minor nitroreductase enzymes of Escherichia coli, NfsA and NfsB, have both been identified as potential prodrug activating enzymes in Gene-Directed Enzyme Prodrug Therapy (GDEPT) of cancer with 5-(aziridin-1-yl)-2,4-dinitrobenzamide (CB1954). NfsB mutants have been selected for their ability to sensitize cells to successively lower concentrations of CB1954. In this study the crystal structures of most improved double (T41L/N71S) and triple (T41Q/N71S/F124T) mutants have been determined using X-ray crystallography. The structures help explain the improvements seen with the mutants, in particular, the structure of the triple mutant reveals a hydrogen bond between Gln41 and Thr124, explaining why this mutation has greater activity than expected from the single mutants. Crystal structures of NfsB mutants where Phe124 is replaced with either of the unnatural amino acids para-nitrophenylalanine or para-aminophenylalanine show a shift in the side chain rotamer allowing these polar groups to interact with substrates, suggesting this leads to their increased activity. For the first time, the crystal structure of NfsA is presented bound to a series of ligands, including the inhibitors succinate and fumarate, and the substrate nitrofurantoin. Another structure bound to a second FMN shows how the flexible loop may move to form a phosphate binding pocket for NADPH.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.577812  DOI: Not available
Keywords: QH426 Genetics ; RC0254 Neoplasms. Tumors. Oncology (including Cancer)
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