Abstract

Eighty clinical bacterial isolates were collected from Sayed Galal hospital in Cairo. Minimum inhibitory concentrations (MICs) were determined for five aminoglycoside antibiotics: netilmycin, sissomycin, spectinomycin, gentamicin and neomycin. The most potent gentamicin-resistant bacterial isolate (IS-20) was identified as Staphylococcus aureus. On the other hand, the same eighty clinical bacterial isolates were screened to produce modifying enzyme-inhibitory protein against the same five antibiotics mentioned above. Only two bacterial isolates gave activity: IS-37 and IS-46. However, IS-46 isolate higher activity than IS-37 isolate with all five antibiotics, especially with gentamicin. Bacterial isolate IS-46 was identified as Pseudomonas aeruginosa. Optimization was studied to obtain the maximum yield of gentamicin modifying-enzyme inhibitory protein. Gentamicin modifying-enzyme inhibitory protein was precipitated at 50% saturated ammonium sulfate, and then purified using ion exchange (DEAE-cellulose) and gel filtration column chromatography (Sephadex G-200). The purified inhibitory protein was electrically separated at 32 KDa. Amino acids sequence and concentration were determined by HPLC. Gentamicin modifying-enzyme inhibitory protein was combined at 128 mg.L^-1 with gentamicin antibiotic at 128 µg.ml^-1 to inhibit the growth of S. aureus. 

Key words: Antimicrobial resistance, aminoglycosides, enzyme inhibitors, protein purification.