Identification of catechol dioxygenases of acinetobacter species, with improved ability to degrade crude oil

Abstract

Background: Various microbes involved in alkanes degradation or reducing organics oil products have been identified among. The goal of this work was to screen the soil bacteria with potential ability to degrade alkanes and characterized using a combination of genetic and physiological methods.Methods: The microbial species of soil sample was isolated and the loss of hydrocarbon was calculated periodically by gravimetric analysis. Isolated bacterial strain with potential ability to degrade the engine oil in Baghdad city was further tested for hydrocarbon-degrading abilities using 2, 6-DCPIP. Colonies of bacteria were counted using serial dilution methods of the soil sample. One of the highly bioactive degrading strains ISN53 was selected and a different fraction of crude extract were analyzed for their hydrocarbon degrading activity. The purified active fraction characterised in LC/MS and SDS PAGE. Presence of chlorocatechol dioxygenases was checked by PCR and expression was analyzed in RT PCR. The envelope of ISn21 isolate was checked for any changes under different conditions using microbial adhesion to the hydrocarbon test (MATH). Results: Serratia marcesens (Sm53) and Acinetobacter venetianus (ISn21) were isolated and ISn21 showed maximum growth on the third day and up to 72% degradation on the 7th day of incubation. LC-MS/MS spectrometry analysis identified the bioactive enzyme catechol dioxygenases in crude extract. The purified catechol dioxygenases enzyme had 35 kDa weight which showed the maximum activity between 48 to 72 hour of incubation. The expression of its gene was 2 fold more than control the maximum expression was observed at 60 to 72 hours of incubation. ISn21 was extremely hydrophobic both in MMV and in LB medium.Conclusion: Using a naturally available resources would be better with efficiency at degrading diesel. Use of like microorganisms during bioremediation operation might supply worthy advances in this important biotechnological/ biological domain.