Isolation and characterization of a heavy metal-resistant, thermophilic esterase from a Red Sea Brine Pool

Abstract

The Red Sea Atlantis II brine pool is an extreme environment that displays multiple harsh conditions such as high temperature, high salinity and high concentrations of multiple, toxic heavy metals. The survival of microbes in such an environment by utilizing resistant enzymes makes them an excellent source of extremophilic enzymes. We constructed a fosmid metagenomic library using DNA isolated from the deepest and most secluded layer of this pool. We report the isolation and biochemical characterization of an unusual esterase: EstATII. EstATII is thermophilic (optimum temperature, 656C), halotolerant (maintains its activity in up to 4.5 M NaCl) and maintains at least 60% of its activity in the presence of a wide spectrum of heavy metals. The combination of biochemical characteristics of the Red Sea Atlantis II brine pool esterase, i.e., halotolerance, thermophilicity and resistance to heavy metals, makes it a potentially useful biocatalyst.U ntil recently and despite its uniqueness, the Red Sea has received little attention compared with other marine environments. The Red Sea formed 3-5 million years ago when the Arabian and African plates began to split 1 , and it is characterized by high temperature and salinity due to the high rate of evaporation, lack of major river inflows and low rate of rainfall. The Red Sea is characterized by the presence of deep-sea, hypersaline anoxic basins called brine pools, which are large bodies of water at the bottom of the ocean that are characterized by high temperature and salinity. To date, 25 brine pools have been found in the Red Sea 1,2 . Atlantis II Deep (Figure 1) is the largest brine pool in the Red Sea, and it has the highest temperature and is the most dynamic 1,3 . This brine pool has a maximum depth of 2,194 m and is stratified into several layers that increase in temperature and salinity with increasing depth: the brine-seawater interface, the upper convective layer, the middle convective layer and the lower convective layer (LCL) 1,3 . The lowest layer, LCL, is characterized by a temperature of 68.2uC, pH value of 5.3 and salinity of 270 psu, which is 7.5 times that of normal seawater 1,3 . Atlantis II Deep is nearly anoxic and contains high concentrations of iron, zinc, copper and other heavy metals 1,3 . Together, these extreme conditions make the Atlantis II brine pool an attractive site for mining for biocatalysts, such as lipolytic enzymes, which are predicted to possess desirable traits that include but are not limited to thermo-tolerance, halo-tolerance, pH plasticity and resistance to inhibition by heavy metals.Industrialized societies are moving toward white (industrial) biotechnology, which has proven to be environmentally sound and commercially efficient 4 . Doing so poses a continuous demand for novel biocatalysts, preferably biocatalysts that demonstrate high activity over a wide range of conditions such as temperature, salinity, pH and metal concentrations. Biocatalysts of microbial origin represent the majority of biocatalysts that are used in indu...

Characterization of a novel marine microbial esterase and its use to make D-methyl lactate

“…PHE14 was also quite stable in the presence of multiple organic solvents, surfactants and metal ions, thus making it a very good esterase candidate for organic synthesis that requires an organic solvent, surfactant or metal ion. Similar behavior was observed with other microbial esterases [18][19][20][21]. PHE14 was also characterized to be a salt-tolerant microbial esterase.…”

Functional Characterization of a Robust Marine Microbial Esterase and Its Utilization in the Stereo-Selective Preparation of Ethyl (S)-3-Hydroxybutyrate

“…(Table 3). However, the hydrolytic activity of one esterase EstATII from a Red Sea brine pool was inhibited by Co 2+ (94.8 %) and Zn 2+ (85.2 %) [27]. Similar results were reported for one esterase EstIM1 from a metagenomic library of mountain soil which was inhibited by Co 2+ (87 %) and even completely inhibited by Zn 2+ (18 %) [24].…”

A Novel Cold Active Esterase from a Deep Sea Sponge Stelletta normani Metagenomic Library

“…The esterase was found to be closely related to the aforementioned E40 esterase (66% amino acid homology), which was itself isolated from metagenomic DNA from a depth of 154 m Sea (Li et al, 2012;Li P. Y. et al, 2015). 7N9 and E40 both possess the two highly conserved GDSAG and HGG motifs which group them into the correspondent subfamily of lipase family IV (Mohamed et al, 2013;Ramnath et al, 2017; Figure 3). In contrast however the 7N9 esterase has a much lower optimal temperature (20 • C) than the E40 esterase (45 • C) and is therefore the first truly cold-adapted esterase in this lipase subfamily.…”

“…The E40 esterase is part of the GDSAG motif subfamily within the lipase family IV; phylogenetic comparisons (Figure 2) and multiple sequence alignments (Figure 3) indicate that the 7N9 esterase is part of the same subfamily of lipolytic enzymes as it also contains the characteristic GDSAG motif (hormone sensitive family, Hsl). Furthermore, the esterase also contains the highly conserved HisGly-Gly (HGG) motif, which together with the GDSAG motif is involved in oxyanion hole formation (Mohamed et al, 2013;Ramnath et al, 2017).…”