1. In vitro kinetics and binding studies of inhibitory compounds (PRK and SRB) MtArgJ
In vitro kinetics and binding studies of inhibitory compounds (PRK and SRB) MtArgJ A, BChemical structures of (A) Pranlukast (PRK) and (B) Sorafenib (SRB).C, DSaturation curve fit using Michaelis–Menten plot for MtArgJ activity at two different inhibitor concentrations for (C) PRK and (D) SRB, quantified by TLC‐based assay.E, FDixon plot analysis performed by plotting 1/v against varying inhibitor concentration, at three different substrate concentrations for (E) PRK and (F) SRB.G, HFluorescence‐based dye (ANS) displacement assay to determine the binding of (G) PRK and (H) SRB into the hydrophobic pocket on MtArgJ. Kd values for both were determined by plotting net change in relative fluorescence upon inhibitor binding as a function of inhibitor concentration.I, JThermal shift assay (TSA) to determine apparent Kd for inhibitor binding to MtArgJ: with increasing concentration of (I) PRK and (J) SRB to examine the shift in melting temperature (Tm) of MtArgJ. Apparent Kd values calculated by plotting net change in melting temperature upon inhibitor binding as a function of inhibitor concentration.Data information: (E–J) Mean and standard error (SE) determined from three experimental replicates (n = 3) and two technical duplicates.
Archita Mishra et al. EMBO Mol Med. 2018;emmm
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