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Genomic Deoxyxylulose Phosphate Reductoisomerase (DXR) Mutations Conferring Resistance to the Antimalarial Drug Fosmidomycin in E. coli
journal contribution
posted on 2018-11-21, 00:00 authored by Gur Pines, Eun Joong Oh, Marcelo C. Bassalo, Alaksh Choudhury, Andrew D. Garst, Reilly G. Fankhauser, Carrie A. Eckert, Ryan T. GillSequence to activity
mapping technologies are rapidly developing,
enabling the generation and isolation of mutations conferring novel
phenotypes. Here we used the CRISPR enabled trackable genome engineering
(CREATE) technology to investigate the inhibition of the essential ispC gene in its native genomic context in Escherichia
coli. We created a full saturation library of 33 sites proximal
to the ligand binding pocket and challenged this library with the
antimalarial drug fosmidomycin, which targets the ispC gene product, DXR. This selection is especially challenging since
it is relatively weak in E. coli, with multiple
naturally occurring pathways for resistance. We identified several
previously unreported mutations that confer fosmidomycin resistance,
in highly conserved sites that also exist in pathogens including the
malaria-inducing Plasmodium falciparum. This approach
may have implications for the isolation of resistance-conferring mutations
and may affect the design of future generations of fosmidomycin-based
drugs.
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Antimalarial Drug FosmidomycinE . coli SequenceE . colisitetechnologyispC gene productCREATEMutations Conferring ResistancegenerationGenomic Deoxyxylulose Phosphate Reductoisomerasemutationresistanceligand binding pocketactivity mapping technologiestrackable genome engineeringCRISPR. coliantimalarial drug fosmidomycinisolationDXRmalaria-inducing Plasmodium falciparum
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