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. 2016 Feb 26;6:19494.
doi: 10.1038/srep19494.

A Novel Cold-Adapted and Highly Salt-Tolerant Esterase From Alkalibacterium Sp. SL3 From the Sediment of a Soda Lake

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Free PMC article

A Novel Cold-Adapted and Highly Salt-Tolerant Esterase From Alkalibacterium Sp. SL3 From the Sediment of a Soda Lake

Guozeng Wang et al. Sci Rep. .
Free PMC article

Abstract

A novel esterase gene (estSL3) was cloned from the Alkalibacterium sp. SL3, which was isolated from the sediment of soda lake Dabusu. The 636-bp full-length gene encodes a polypeptide of 211 amino acid residues that is closely related with putative GDSL family lipases from Alkalibacterium and Enterococcus. The gene was successfully expressed in E. coli, and the recombinant protein (rEstSL3) was purified to electrophoretic homogeneity and characterized. rEstSL3 exhibited the highest activity towards pNP-acetate and had no activity towards pNP-esters with acyl chains longer than C8. The enzyme was highly cold-adapted, showing an apparent temperature optimum of 30 °C and remaining approximately 70% of the activity at 0 °C. It was active and stable over the pH range from 7 to 10, and highly salt-tolerant up to 5 M NaCl. Moreover, rEstSL3 was strongly resistant to most tested metal ions, chemical reagents, detergents and organic solvents. Amino acid composition analysis indicated that EstSL3 had fewer proline residues, hydrogen bonds and salt bridges than mesophilic and thermophilic counterparts, but more acidic amino acids and less hydrophobic amino acids when compared with other salt-tolerant esterases. The cold active, salt-tolerant and chemical-resistant properties make it a promising enzyme for basic research and industrial applications.

Figures

Figure 1
Figure 1. Phylogenetic tree of the amino acid sequences of EstSL3 and its close homologs.
The tree was constructed using the neighbor-joining method (MEGA 4.0). Bootstrap values (n = 1,000 replicates) are reported as percentages. The scale bar represents the number of changes per amino acid position. The sequence accession numbers are given at the end of each species name.
Figure 2
Figure 2. Structure and surface electrostatic potential analysis of EstSL3.
(a) Modeled EstSL3 constructed by I-TASSER with 3w7vA used as the template. (b) The surface electrostatic potential of EstSL3 obtained by Pymol and APBS plugin. (c) The 180° rotated view of (a). (d) The 180° rotated view of (b). (e) The negative and positive electrostatic potentials are indicated by blue and red, respectively.
Figure 3
Figure 3. Enzymatic properties of purified rEstSL3.
(a) Effect of pH on EstSL3 activity. Activities at various pHs were assayed at 37 °C for 5 min. (b) pH stability of EstSL3. Residual activities after incubation at various pHs for 1 h at 37 °C were assayed at pH 9.0 and 30 °C for 5 min. (c) Effect of temperature on EstSL3 activity in Tris-HCl buffer (pH 9.0). (d) Thermostability of EstSL3. Residual activity was assayed at pH 9.0 and 30 °C for 5 min after pre-incubation at 50 °C, 55 °C or 60 °C for different periods of time. The data are shown as means ± SD (n = 3).
Figure 4
Figure 4. Effect of NaCl on rEstSL3 activity and stability.
(a) Effect of different concentrations of NaCl on the activity of rEstSL3. (b) rEstSL3 stability in the presence of 4 M NaCl. The data are shown as means ± SD (n = 3).

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