<b>Cryo-EM structures of CCHFV polymerase reveal a stepwise</b><b> </b><b>initiation </b><b>stabilization </b><b>pathway and a</b><b> </b><b>dual-site inhibition</b><b> </b><b>mechanism</b><b></b>

<jats:p>Crimean-Congo hemorrhagic fever virus (CCHFV) is a high-priority pathogen with high case-fatality rates, yet approved therapeutics remain unavailable. The CCHFV L segment encodes a ~450-kDa RNA-dependent RNA polymerase (L protein) orchestrating viral replication, but its structural mechanisms remain elusive. Here, we present high-resolution cryo-EM structures of the CCHFV L protein in apo, 5' vRNA-bound, 5'/3' promoter-bound, and inhibitor-bound states. The catalytic core exhibits the canonical architecture of the Bunyavirales order, featuring conserved motifs and coordinated promoter recognition via a 5' vRNA "hook" and a secondary 3' vRNA-binding site. Using suramin as a probe, we identified a dual-site mechanism of polymerase inhibition. Suramin competitively occludes the 5' vRNA-binding pocket through electrostatic mimicry of the RNA backbone and concurrently traps a distal Linker–Fingers interface, restricting the conformational dynamics required for catalysis. Collectively, these findings provide structural insights into CCHFV polymerase regulation and inhibition.</jats:p>