Structural and functional insights into the distinct DNA recognition mechanisms of the terminase small subunit TerS from cyanophages

• Cryo-EM structure of cyanophage Pam5 TerS reveals a nonameric ring with a spiral architecture, providing the first structural insight into TerS from cyanophages. • Pam5 TerS recognizes a specific 21-bp DNA sequence via its N-terminal HTH domain, whereas Pam1 TerS binds DNA sequence-independently, demonstrating two distinct DNA recognition strategies. • The study highlights the evolutionary plasticity of viral genome packaging mechanisms, even among phages infecting the same host. • These findings advance our understanding of TerS-mediated DNA recognition and may inform the design of phage-based therapies or biotechnological applications.