Abstract
An Egyptian marine bacterium, isolated from Hamam Pheroon, South Sinai region was able to produce thermostable proteases, the isolate was identified morphologically, biochemically, and confirmed molecularly by 16S rRNA sequencing with 99% similarity to Alcaligenes faecalis. It exhibited optimum activity of 328.3 U/mg after ten min, incubation at 65C and pH 7. Both ammonium sulphate and sephadex G-100 purification methods enhanced the yield of Alcaligenes faecalis strain HFW-9081 to 125 and 121% as well as the specific activity to 458.9 and 590 U/mg, respectively, compared to cell free supernatant. However, relative protease activity was reduced to 35.8% when H2O2 was added. On the other hand, the activities increased 7.5 folds when Tween-80 was used as a surfactant. Genetic background of the protease genes in Alcaligenes faecalis was analyzed using bioinformatics database for the proteases amino acids sequences in the desired bacteria; and it specified that Alcaligenes faecalis has four different protease genes; these genes encode for various peptidases family groups. The variation in the peptidase family groups provides the protease enzymes with many features making them able to remain active under various environmental stresses. The overall results showed promising thermostable proteases isolated from local marine Egyptian bacterium; that can be used potentially in many industrial applications.
Highlights
Bacterial culture was sent to SolGent
The following molecular techniques were done as follow
Primary selection of the isolates was according to the biggest clear zone using
Summary
Pure isolate was characterized morphologically using scanning electron microscope (SEM) and based on the criteria of Bergey’s Manual of Systematic. 16S rRNA gene of the new isolate was amplified using universal 16S rRNA primers. South Korea for 16S rRNA gene sequencing. The following molecular techniques were done as follow: DNA extraction and isolation was done using. The ribosomal rRNA gene ( referred to as rDNA) was amplified using the polymerase chain reaction (PCR). 27F and 1492R were incorporated in the reaction mixture. The reaction conditions were, initial denaturation step at 94 C for. 3 min, after which the Taq polymerase was added, and the reaction continued for the 35 cycles of 45 sec of denaturation at 94 C, 60 sec for the annealing at. Final extension was done for 10 min at 72 C, after which the reaction hold at. Crude enzymes were assayed for proteolytic activity in triplicate as described by Guangrong et al (2006) using casein as the substrate
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
