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dGMP Binding to Thymidylate Kinase from Plasmodium falciparum Shows Half-Site Binding and Induces Protein Dynamics at the Dimer Interface
journal contribution
posted on 2020-01-13, 22:29 authored by Mengshen
David Chen, Ian J. Fucci, Kaustubh Sinha, Gordon S. RulePlasmodium falciparum thymidylate kinase (PfTMK)
is an essential enzyme for the growth of the organism because of its
critical role in the de novo synthesis of deoxythymidine
5′-diphosphate (TDP), a precursor for TTP that is required
for DNA replication and repair. The kinetics, thermodynamic parameters,
and substrate binding properties of PfTMK for TMP, dGMP, ADP, and
ATP were measured and characterized by steady-state kinetics and a
combination of isothermal titration calorimetry, tryptophan fluorescence
titration, and NMR. Mutational studies were performed to investigate
residues that contribute to the unique ability of PfTMK to also utilize
dGMP as a substrate. Isothermal titration calorimetry experiments
revealed that dGMP binding exhibits a unique half-site binding mechanism.
The occlusion of the empty site in the dGMP complex is supported by
molecular mechanics calculations. Relaxation dispersion experiments
show that the dGMP and enzyme complex is more dynamic at the dimer
interface than the TMP complex on the μs–ms time scale.
The unique properties of dGMP binding need to be considered in the
design of guanosine-based PfTMK-specific inhibitors.
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dGMP Bindingguanosine-based PfTMK-specific inhibitorsPlasmodium falciparum Shows Half-Site Bindinghalf-site binding mechanismdimer interfaceIsothermal titration calorimetry experimentsTTPMutational studiesTDPtitration calorimetryTMPATPDimer Interface Plasmodium falciparum thymidylate kinaseADPdGMP binding needsubstrate binding propertiesRelaxation dispersion experiments showInduces Protein DynamicsdGMP binding exhibitsNMRThymidylate Kinasetryptophan fluorescence titrationDNA replicationmechanics calculations
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