Crystal Structure and Functional Characterization of an Esterase (EaEST) from Exiguobacterium antarcticum.

A novel microbial esterase, EaEST, from a psychrophilic bacterium Exiguobacterium antarcticum B7, was identified and characterized. To our knowledge, this is the first report describing structural analysis and biochemical characterization of an esterase isolated from the genus Exiguobacterium. Cryst...

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Main Authors: Chang Woo Lee, Sena Kwon, Sun-Ha Park, Boo-Young Kim, Wanki Yoo, Bum Han Ryu, Han-Woo Kim, Seung Chul Shin, Sunghwan Kim, Hyun Park, T Doohun Kim, Jun Hyuck Lee
Format: Article
Language:English
Published: Public Library of Science (PLoS) 2017-01-01
Series:PLoS ONE
Online Access:http://europepmc.org/articles/PMC5268438?pdf=render
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spelling doaj-e1260855eb1b49fab81a468c7e3482362020-11-24T20:45:06ZengPublic Library of Science (PLoS)PLoS ONE1932-62032017-01-01121e016954010.1371/journal.pone.0169540Crystal Structure and Functional Characterization of an Esterase (EaEST) from Exiguobacterium antarcticum.Chang Woo LeeSena KwonSun-Ha ParkBoo-Young KimWanki YooBum Han RyuHan-Woo KimSeung Chul ShinSunghwan KimHyun ParkT Doohun KimJun Hyuck LeeA novel microbial esterase, EaEST, from a psychrophilic bacterium Exiguobacterium antarcticum B7, was identified and characterized. To our knowledge, this is the first report describing structural analysis and biochemical characterization of an esterase isolated from the genus Exiguobacterium. Crystal structure of EaEST, determined at a resolution of 1.9 Å, showed that the enzyme has a canonical α/β hydrolase fold with an α-helical cap domain and a catalytic triad consisting of Ser96, Asp220, and His248. Interestingly, the active site of the structure of EaEST is occupied by a peracetate molecule, which is the product of perhydrolysis of acetate. This result suggests that EaEST may have perhydrolase activity. The activity assay showed that EaEST has significant perhydrolase and esterase activity with respect to short-chain p-nitrophenyl esters (≤C8), naphthyl derivatives, phenyl acetate, and glyceryl tributyrate. However, the S96A single mutant had low esterase and perhydrolase activity. Moreover, the L27A mutant showed low levels of protein expression and solubility as well as preference for different substrates. On conducting an enantioselectivity analysis using R- and S-methyl-3-hydroxy-2-methylpropionate, a preference for R-enantiomers was observed. Surprisingly, immobilized EaEST was found to not only retain 200% of its initial activity after incubation for 1 h at 80°C, but also retained more than 60% of its initial activity after 20 cycles of reutilization. This research will serve as basis for future engineering of this esterase for biotechnological and industrial applications.http://europepmc.org/articles/PMC5268438?pdf=render
collection DOAJ
language English
format Article
sources DOAJ
author Chang Woo Lee
Sena Kwon
Sun-Ha Park
Boo-Young Kim
Wanki Yoo
Bum Han Ryu
Han-Woo Kim
Seung Chul Shin
Sunghwan Kim
Hyun Park
T Doohun Kim
Jun Hyuck Lee
spellingShingle Chang Woo Lee
Sena Kwon
Sun-Ha Park
Boo-Young Kim
Wanki Yoo
Bum Han Ryu
Han-Woo Kim
Seung Chul Shin
Sunghwan Kim
Hyun Park
T Doohun Kim
Jun Hyuck Lee
Crystal Structure and Functional Characterization of an Esterase (EaEST) from Exiguobacterium antarcticum.
PLoS ONE
author_facet Chang Woo Lee
Sena Kwon
Sun-Ha Park
Boo-Young Kim
Wanki Yoo
Bum Han Ryu
Han-Woo Kim
Seung Chul Shin
Sunghwan Kim
Hyun Park
T Doohun Kim
Jun Hyuck Lee
author_sort Chang Woo Lee
title Crystal Structure and Functional Characterization of an Esterase (EaEST) from Exiguobacterium antarcticum.
title_short Crystal Structure and Functional Characterization of an Esterase (EaEST) from Exiguobacterium antarcticum.
title_full Crystal Structure and Functional Characterization of an Esterase (EaEST) from Exiguobacterium antarcticum.
title_fullStr Crystal Structure and Functional Characterization of an Esterase (EaEST) from Exiguobacterium antarcticum.
title_full_unstemmed Crystal Structure and Functional Characterization of an Esterase (EaEST) from Exiguobacterium antarcticum.
title_sort crystal structure and functional characterization of an esterase (eaest) from exiguobacterium antarcticum.
publisher Public Library of Science (PLoS)
series PLoS ONE
issn 1932-6203
publishDate 2017-01-01
description A novel microbial esterase, EaEST, from a psychrophilic bacterium Exiguobacterium antarcticum B7, was identified and characterized. To our knowledge, this is the first report describing structural analysis and biochemical characterization of an esterase isolated from the genus Exiguobacterium. Crystal structure of EaEST, determined at a resolution of 1.9 Å, showed that the enzyme has a canonical α/β hydrolase fold with an α-helical cap domain and a catalytic triad consisting of Ser96, Asp220, and His248. Interestingly, the active site of the structure of EaEST is occupied by a peracetate molecule, which is the product of perhydrolysis of acetate. This result suggests that EaEST may have perhydrolase activity. The activity assay showed that EaEST has significant perhydrolase and esterase activity with respect to short-chain p-nitrophenyl esters (≤C8), naphthyl derivatives, phenyl acetate, and glyceryl tributyrate. However, the S96A single mutant had low esterase and perhydrolase activity. Moreover, the L27A mutant showed low levels of protein expression and solubility as well as preference for different substrates. On conducting an enantioselectivity analysis using R- and S-methyl-3-hydroxy-2-methylpropionate, a preference for R-enantiomers was observed. Surprisingly, immobilized EaEST was found to not only retain 200% of its initial activity after incubation for 1 h at 80°C, but also retained more than 60% of its initial activity after 20 cycles of reutilization. This research will serve as basis for future engineering of this esterase for biotechnological and industrial applications.
url http://europepmc.org/articles/PMC5268438?pdf=render
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