NDLI: Two novel deep-sea sediment metagenome-derived esterases: residue 199 is the determinant of substrate specificity and preference

Content Provider	Springer Nature : BioMed Central
Author	Huo, Ying-Yi Jian, Shu-Ling Cheng, Hong Rong, Zhen Cui, Heng-Lin Xu, Xue-Wei
Abstract	Background The deep-sea environment harbors a vast pool of novel enzymes. Owing to the limitations of cultivation, cultivation-independent has become an effective method for mining novel enzymes from the environment. Based on a deep-sea sediment metagenomics library, lipolytic-positive clones were obtained by activity-based screening methods. Results Two novel esterases, DMWf18-543 and DMWf18-558, were obtained from a deep-sea metagenomic library through activity-based screening and high-throughput sequencing methods. These esterases shared 80.7% amino acid identity with each other and were determined to be new members of bacterial lipolytic enzyme family IV. The two enzymes showed the highest activities toward p-nitrophenyl (p-NP) butyrate at pH 7.0 and 35–40 °C and were found to be resistant to some metal ions (Ba2+, Mg2+, and Sr2+) and detergents (Triton X-100, Tween 20, and Tween 80). DMWf18-543 and DMWf18-558 exhibited distinct substrate specificities and preferences. DMWf18-543 showed a catalytic range for substrates of C2–C8, whereas DMWf18-558 presented a wider range of C2–C14. Additionally, DMWf18-543 preferred p-NP butyrate, whereas DMWf18-558 preferred both p-NP butyrate and p-NP hexanoate. To investigate the mechanism underlying the phenotypic differences between the esterases, their three-dimensional structures were compared by using homology modeling. The results suggested that residue Leu199 of DMWf18-543 shortens and blocks the substrate-binding pocket. This hypothesis was confirmed by the finding that the DMWf18-558-A199L mutant showed a similar substrate specificity profile to that of DMWf18-543. Conclusions This study characterized two novel homologous esterases obtained from a deep-sea sediment metagenomic library. The structural modeling and mutagenesis analysis provided insight into the determinants of their substrate specificity and preference. The characterization and mechanistic analyses of these two novel enzymes should provide a basis for further exploration of their potential biotechnological applications.
Related Links	https://microbialcellfactories.biomedcentral.com/counter/pdf/10.1186/s12934-018-0864-4.pdf
Ending Page	12
Page Count	12
Starting Page	1
File Format	HTM / HTML
ISSN	14752859
DOI	10.1186/s12934-018-0864-4
Journal	Microbial Cell Factories
Issue Number	1
Volume Number	17
Language	English
Publisher	BioMed Central
Publisher Date	2018-01-30
Access Restriction	Open
Subject Keyword	Applied Microbiology Biotechnology Microbiology Microbial Genetics and Genomics Enzymology Genetic Engineering Metagenomics library Esterase Family IV Deep-sea Substrate-binding pocket Homology modeling
Content Type	Text
Resource Type	Article
Subject	Applied Microbiology and Biotechnology Bioengineering Biotechnology
Journal Impact Factor	4.3/2023
5-Year Journal Impact Factor	5.5/2023

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