The Atlantis II deep is the largest brine pool in the Red Sea of around 60 km2. It is located in the central Red Sea (21° 21' N., 38° 04' E.) at a depth of 2200 meters from the sea surface. The lower part of this pool, the lower convective layer (LCL), has extremely harsh environmental conditions characterized by a temperature of up to 68°C, pH 5.3, saturated salt (252 psu), anoxic condition, and high concentration of heavy metals. These harsh abiotic factors exert strong selective pressure in favor of microbial communities adapted to survive in such environment. Therefore, proteins and enzymes involved in the metabolic pathways and physiological processes of these microorganisms should have novel structural and enzymatic activities that could be of interest in biotechnological applications. In this work we utilized an integrated metagenomic approach to identify sequences of potential secreted subtilisin-like serine proteases in the LCL metagenomic datasets, and also functionally screened the LCL metagenomic fosmid library using skimmed milk (casein) as a substrate. Out of 1,167,000 LCL reads (average length 400 nucleotides/read), we obtained 286 sequences that matched potential secreted subtilisin-like serine proteases. The 286 matched reads were assembled into 31 contigs. In the functional analysis, we screened 10,656 clones that provided 9 positive candidates as judged by characteristic clear halo around the positive colonies. Pyrosequencing of 2 of the plates harboring functionally positive clones provided 5 open reading frames that matched subtilisin-like serine protease. This work provided a pivotal step toward identification of subtilisin-like serine proteases with novel catalytic activities.


School of Sciences and Engineering

Degree Name

MS in Biotechnology

First Advisor

El Dorry, Hamza

Document Type



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