Abstract The function unknown gene PA2384 of Pseudomonas aeruginosa PAO1 has been previously shown dramatically responsive to iron limitation. In the present study, a bioinformatics analysis showed that PA2384 has a weak similarity to the N-terminus DNA-binding domain of Fur, the well-known ferric uptake regulator. To investigate the potential function of PA2384 in iron regulation a P. aeruginosa PAO1 recombinant (pUCP20::PA2384) over-expressing PA2384 and a PA2384 disrupted mutant PAO1*PA2384 were constructed. Physiological characterization showed that the knockout mutant had a longer lag phase. Genome-scale transcriptional profiles at different growth stages were compared between the wild type and the ΔPA2384 mutant grown under iron-limiting conditions. The expression of more than 350 genes was affected by the knockout of PA2384. Among them, 71 genes involved in iron uptake were significantly down-regulated in the absence of PA2384. One hundred two quorum sensing (QS) dependent genes displayed differential transcriptions, including genes involved in the biosynthesis of some important virulence factors such as pyocyanin, rhamnolipids and hydrogen cyanide. The transcription of genes responsible for the synthesis of Pseudomonas quinolone signal (PQS) was greatly enhanced by the knockout of PA2384. Furthermore, the knockout of PA2384 also resulted in an altered expression of genes involved in electron transfer, central metabolism, phosphorus starvation and translation. It implies that PA2384 might affect more physiological processes than iron acquisition in P. aeruginosa.