Background: The ability of Pseudouridimycin (PUM) to occupy the nucleotide addition
site of bacterial RNA Polymerase (RNAP) underlies its inhibitory potency, as previously reported.
PUM has gained high research interest as a broad-spectrum nucleoside analog that has demonstrated
exciting potentials in treating drug-resistant bacterial infections.
Objective: Herein, we identified, for the first time, a novel complementary mechanism by which
PUM elicits its inhibitory effects on bacterial RNAP.
Materials and Methods: The dynamic binding behavior of PUM to bacterial RNAP was studied
using various dynamic analysis approaches.
Results: Findings revealed that in addition to occupying the nucleotide addition site, PUM also
interrupts the unimpeded entry and exit of DNA by reducing the mechanistic extension of the
RNAP cleft and perturbing the primary conformations of the switch regions. Moreover, PUM binding
reduced the distances between key residues in the β and β’ subunits that extend to accommodate
Conclusion: This study’s findings present structural insights that would contribute to the structurebased
design of potent and selective PUM inhibitors.