Isolation of halotolerant glutaminase producing microorganisms was carried out using medium containing different concentrations of NaCl viz. 2.9, 3.5 and 4 %. Only four fungal isolates (F1H, F2H, F3H and F4H) were able to grow at all concentrations has been selected and purified. Production of glutaminase was carried out in broth medium containing 4% NaCl. Quantitaive assay for glutaminase led to 6.43, 5.72, 6.12 and 5.86 U/ml for each isolate extracellularly, respectively. In case of intracellular production assay led to 5.2, 5.46, 5.18 and 5.77 U/ml , respectively. The isolate F1H has showed the highest productivity extracellularly and stability on subculturing so, it was chosen and selected for furthur investigations. The optimized conditions for glutaminase production by isolate F1H was found to be 6 days at 35 o C and pH 7 under static and dark incbation conditions with inoculum size of 4.38x10 6 spores/ ml. The optimization led to production of 7.66 U/ml of enzyme with great stability at 4% salt concentration which is a good sign for probability that the enzyme could have unique properties which can be confirmed through more investigatios.The isolate was identified genetically and has shown 100% similarity to Aspergillus flavus CZCU-9 and it was given the name Aspergillus flavus CZCU-9, F1H.