Structural maintenance of chromosomes (SMC) complexes play pivotal roles in genome organization and maintenance across all domains of life. In prokaryotes, SMC-family Wadjet complexes structurally resemble the widespread MukBEF but serve a defensive role by inhibiting plasmid transformation. We previously showed that Wadjet specifically cleaves plasmid DNA
however, the molecular mechanism underlying plasmid recognition remains unclear. Here, we use in vitro single-molecule imaging to directly visualize DNA loop extrusion and plasmid cleavage by Wadjet. We find that Wadjet is a symmetric loop extruder that simultaneously reels in DNA from both sides of a loop and that this activity requires a dimeric JetABC supercomplex. On surface-anchored plasmid DNAs, Wadjet extrudes the full length of a 44-kb-pair plasmid, stalls, and cleaves DNA. Our findings reveal the role of loop extrusion in the specific recognition and elimination of plasmids by Wadjet and establish loop extrusion as an evolutionarily conserved mechanism among SMC complexes across all kingdoms of life.