8V5B

Structure of the oxygen-insensitive NAD(P)H-dependent nitroreductase NfsB_Ec F70A/F108Y in complex with FMN

Summary for 8V5B

• Entry DOI: 10.2210/pdb8v5b/pdb
• Descriptor: Dihydropteridine reductase, FLAVIN MONONUCLEOTIDE, ACETATE ION, ... (6 entities in total)
• Functional Keywords: nitroreductase, flavoprotein, fmn, oxidoreductase
• Biological source: Escherichia coli
• Total number of polymer chains: 4
• Total formula weight: 98504.32

Authors

Sharrock, A.V.,Ackerley, D.F.,Arcus, V. (deposition date: 2023-11-30, release date: 2024-10-09)

Primary citation

Sharrock, A.V.,Mumm, J.S.,Williams, E.M.,Cenas, N.,Smaill, J.B.,Patterson, A.V.,Ackerley, D.F.,Bagdziunas, G.,Arcus, V.L.
Structural Evaluation of a Nitroreductase Engineered for Improved Activation of the 5-Nitroimidazole PET Probe SN33623.
Int J Mol Sci, 25:-, 2024

PubMed Abstract: Bacterial nitroreductase enzymes capable of activating imaging probes and prodrugs are valuable tools for gene-directed enzyme prodrug therapies and targeted cell ablation models. We recently engineered a nitroreductase ( NfsB F70A/F108Y) for the substantially enhanced reduction of the 5-nitroimidazole PET-capable probe, SN33623, which permits the theranostic imaging of vectors labeled with oxygen-insensitive bacterial nitroreductases. This mutant enzyme also shows improved activation of the DNA-alkylation prodrugs CB1954 and metronidazole. To elucidate the mechanism behind these enhancements, we resolved the crystal structure of the mutant enzyme to 1.98 Å and compared it to the wild-type enzyme. Structural analysis revealed an expanded substrate access channel and new hydrogen bonding interactions. Additionally, computational modeling of SN33623, CB1954, and metronidazole binding in the active sites of both the mutant and wild-type enzymes revealed key differences in substrate orientations and interactions, with improvements in activity being mirrored by reduced distances between the N5-H of isoalloxazine and the substrate nitro group oxygen in the mutant models. These findings deepen our understanding of nitroreductase substrate specificity and catalytic mechanisms and have potential implications for developing more effective theranostic imaging strategies in cancer treatment.

PubMed: 38928299
DOI: 10.3390/ijms25126593

Experimental method

X-RAY DIFFRACTION (1.98 Å)