Many microorganisms thrive on the abundant nutrients in the rhizosphere and they produce antagonistic metabolites. The present study is an attempt to investigate the potent bacteria producing antibiotics against pathogenic microorganisms from the rhizosphere soil samples of Kanjamalai hills, Salem district, Tamilnadu, India. A total of 6 isolates of bacteria was obtained and screened for inhibitory activity against selected opportunistic pathogens. Potent bacterial isolate (A6) was identified as Pseudomonas aeruginosa by cultural, morphological and biochemical characterization. The antimicrobial activity of broth culture of Pseudomanas aeruginosa was also observed. The antimicrobial active compound was partially purified from the broth culture and its concentration was estimated to be 373.086 mu g/ml. The obtained protein was separated in SDS-PAGE and its molecular weight was determined as 66.4 kD, 44.3 kD, 29.0 kD, 20.1 kD and 14.3 kD using standard molecular markers. The results showed that the isolated strain has high potential against Bacillus megaterium and Aspergillus niger. It can be further investigated to identify the antibacterial compounds produced and their mechanisms of inhibition.