(R)-Gyramide A (hydrochloride) is a bacterial DNA gyrase inhibitor that disrupts supercoiling activity with an IC50 value of 3.3 µM. (R)-Gyramide A (hydrochloride) demonstrates antibacterial activity against E. coli, P. aeruginosa, and S. enterica (MICs of 10-80 µM). (R)-Gyramide A (hydrochloride) does not affect the closely related enzyme topoisomerase IV.