title: EstDZ3: A New Esterolytic Enzyme Exhibiting Remarkable Thermostability.
abstract: Lipolytic enzymes that retain high levels of catalytic activity when exposed to a variety of denaturing conditions are of high importance for a number of biotechnological applications. In this study, we aimed to identify new lipolytic enzymes, which are highly resistant to prolonged exposure to elevated temperatures. To achieve this, we searched for genes encoding for such proteins in the genomes of a microbial consortium residing in a hot spring located in China. After performing functional genomic screening on a bacterium of the genusDictyoglomus, which was isolated from this hot spring followingin situenrichment, we identified a new esterolytic enzyme, termed EstDZ3. Detailed biochemical characterization of the recombinant enzyme, revealed that it constitutes a slightly alkalophilic and highly active esterase against esters of fatty acids with short to medium chain lengths. Importantly, EstDZ3 exhibits remarkable thermostability, as it retains high levels of catalytic activity after exposure to temperatures as high as 95°C for several hours. Furthermore, it exhibits very good stability against exposure to high concentrations of a variety of organic solvents. Interestingly, EstDZ3 was found to have very little similarity to previously characterized esterolytic enzymes. Computational modeling of the three-dimensional structure of this new enzyme predicted that it exhibits a typical α/β hydrolase fold that seems to include a "subdomain insertion", which is similar to the one present in its closest homolog of known function and structure, the cinnamoyl esterase Lj0536 fromLactobacillus johnsonii. As it was found in the case of Lj0536, this structural feature is expected to be an important determinant of the catalytic properties of EstDZ3. The high levels of esterolytic activity of EstDZ3, combined with its remarkable thermostability and good stability against a range of organic solvents and other denaturing agents, render this new enzyme a candidate biocatalyst for high-temperature biotechnological applications.
authors.affiliation: Institute of Biology, Medicinal Chemistry and Biotechnology, National Hellenic Research FoundationAthens, Greece; Laboratory of Biotechnology, School of Chemical Engineering, National Technical University of AthensAthens, Greece.
authors.forename: Dimitra
authors.initials: D
authors.lastname: Zarafeta
authors.affiliation: MicroDish B.V. Utrecht, Netherlands.
authors.forename: Zalan
authors.initials: Z
authors.lastname: Szabo
authors.affiliation: Laboratory of Biotechnology, School of Chemical Engineering, National Technical University of Athens Athens, Greece.
authors.forename: Danai
authors.initials: D
authors.lastname: Moschidi
authors.affiliation: Danish Archaea Centre, Department of Biology, Copenhagen University Copenhagen, Denmark.
authors.forename: Hien
authors.initials: H
authors.lastname: Phan
authors.affiliation: Institute of Biology, Medicinal Chemistry and Biotechnology, National Hellenic Research Foundation Athens, Greece.
authors.forename: Evangelia D
authors.initials: ED
authors.lastname: Chrysina
authors.affiliation: Danish Archaea Centre, Department of Biology, Copenhagen University Copenhagen, Denmark.
authors.forename: Xu
authors.initials: X
authors.lastname: Peng
authors.affiliation: MicroDish B.V. Utrecht, Netherlands.
authors.forename: Colin J
authors.initials: CJ
authors.lastname: Ingham
authors.affiliation: Laboratory of Biotechnology, School of Chemical Engineering, National Technical University of Athens Athens, Greece.
authors.forename: Fragiskos N
authors.initials: FN
authors.lastname: Kolisis
authors.affiliation: Institute of Biology, Medicinal Chemistry and Biotechnology, National Hellenic Research Foundation Athens, Greece.
authors.forename: Georgios
authors.initials: G
authors.lastname: Skretas
keywords: Dictyoglomus; biocatalysis; biotechnology; esterase; functional genomics; hyperthermostability