A novel solvent tolerant esterase of GDSGG motif subfamily from solar saltern through metagenomic approach: Recombinant expression and characterization

Jayanath, G.; Mohandas, Sowmya P.; Kachiprath, Bhavya; Solomon, Solly; Puthiyedathu, Sajeevan Thavarool; Singh, I. S. Bright; Philip, Rosamma

doi:10.1016/j.ijbiomac.2018.06.057

Cited by 23 publications

(21 citation statements)

References 31 publications

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“…The results of the multiple sequence alignment of EstCS1 with closely related enzymes revealed that there were Ser155-Asp255-His285 residues as the conserved sequence motif of EstCS1. In addition, the catalytic nucleophile serine is at position 155 in the consensus GXSXG pentapeptide and the HGGG motif in the oxyanion hole upstream of the pentapeptide motif; these are characteristics of family IV (HSL family) lipolytic enzymes (Figure 1A; Arpigny and Jaeger, 1999;Kim et al, 2015;Huo et al, 2018;Jayanath et al, 2018;Noby et al, 2018). Phylogenetic analysis using amino acid sequences of EstCS1 and other lipases/esterases also revealed that EstCS1 fell within the cluster including family IV lipolytic enzymes (Figure 1B).…”

Section: Screening and Subcloning Of The Putative Esterase Genementioning

confidence: 95%

“…The genetic libraries were obtained directly from environmental sources without microbial culture in a laboratory by means of screening methods based on activity and sequence to effectively identify novel enzymes being used in various industries (Lee et al, 2006;Schmeisser et al, 2007;Kim et al, 2009;Uchiyama and Miyazaki, 2009;Berini et al, 2017). Especially, a number of lipases and esterases belonging to family IV have been recently isolated from various environmental sources since the development of metagenomic analyses (Oh et al, 2012;Choi et al, 2013;Privé et al, 2015;Kim, 2017;Huo et al, 2018;Jayanath et al, 2018;Noby et al, 2018).…”

Section: Introductionmentioning

confidence: 99%

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Characterization of a Novel Moderately Thermophilic Solvent-Tolerant Esterase Isolated From a Compost Metagenome Library

et al. 2020

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A novel esterase, EstCS1, was isolated from a compost metagenomics library. The EstCS1 protein, which consists of 309 amino acid residues with an anticipated molecular mass of 34 kDa, showed high amino acid sequence identities to predicted esterases and alpha/beta hydrolases (59%) from some cultured bacteria and to predicted lipases/esterases from uncultured bacteria. The phylogenetic analysis suggested that the EstCS1 belongs to the hormone-sensitive lipase family of lipolytic enzyme classification and contains a catalytic triad including Ser155-Asp255-His285. The Ser155 residue of the catalytic triad in the EstCS1 was located in the consensus active-site motif, GXSXG. Besides, a conserved HGGG motif placed in an oxyanion hole of the hormone-sensitive lipase family was discovered, too. The EstCS1 demonstrated the highest activity toward p-nitrophenyl propionate (C3) and caproate (C6) and was normally stable up to 60 • C with optimal activity at 50 • C. In addition, an optimal activity was observed at pH 8, and the EstCS1 possessed its stability within the pH range between 5 and 10. Interestingly, EstCS1 had an outstanding stability in up to 30% (v/v) organic solvents and activity over 50% in the presence of 50% (v/v) acetone, ethanol, dimethyl sulfoxide (DMSO), and N,N-dimethylformamide. The EstCS1 hydrolyzed sterically hindered tertiary alcohol esters of t-butyl acetate and linalyl acetate. Considering the properties, such as the moderate thermostability, stability against organic solvents, and activity toward esters of tertiary alcohols, the EstCS1 will be worthwhile to be used for organic synthesis and related industrial applications.

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Section: Screening and Subcloning Of The Putative Esterase Genementioning

confidence: 95%

Section: Introductionmentioning

confidence: 99%

Characterization of a Novel Moderately Thermophilic Solvent-Tolerant Esterase Isolated From a Compost Metagenome Library

et al. 2020

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“…The specific activity of Est7S was in the upper range among family VII esterases such as Est-XG2 (17.71 U/mg protein) [17], EstDL30 (161 U/mg protein) [19], and an esterase NaM1 (488.9 U/mg protein) from a desert metagenome [29]. Compared to others, Est7S was in the middle range; an esterase Est2K (family VIII, 17.1 U/mg protein) from a compost metagenome [10], Lip7- [11], EstN7 of B. cohnii strain N1 (family IV, 336.89 U/mg protein) [30], EstSP (family IV, 745.14 U/mg protein) from a soil metagenome [31], and Est7K (family VIII, 790.2 U/mg protein) from the compost metagenome [32].…”

Section: Purification Of Est7smentioning

confidence: 84%

“…The activity ratio for C10 to C4 was 0.13, between Est7K (0.035) [32] and Est2K (0.4) [10]. Est7S preferred a shortchain fatty acid (C2 or C4) as the substrate, likely many esterases, such as EstSP (C2) [31], EstN7 (C2) [30], BlEst1 (C2) of B. licheniformis [33], EstSP2 (C2) of Sphingomonas glacialis [34], and an esterase (C2) of Xanthomonas oryzae [35], but contrary to Lpc53E1 (C16) from the metagenome of the marine sponge Haliclona simulans [36] and KM12 lipase (C12) of B. licheniformis KM12 [37]. Based on the substrate preference, Est7S is a typical carboxylesterase rather than a lipase [4].…”

Section: Properties Of Est7smentioning

confidence: 99%

Characterization of an alkaline esterase from an enriched metagenomic library derived from an oil-spill area

Baek

Lee

et al. 2019

JABC

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A novel esterase gene (est7S) was cloned from an enriched metagenomic library derived from an oil-spill area. The gene encoded a protein of 505 amino acids, and the molecular mass of the Est7S was estimated to be 54,512 Da with no signal peptide. Est7S showed the highest identity of 40% to an esterase from a sludge metagenome compared to the characterized enzymes with their properties, although it showed 99% identity to a carboxylesterase in the genome sequence of Alcanivorax borkumensis SK2. Est7S had catalytic triad residues, Ser183, Glu312, and His420, and the GESAG motif in most family VII lipolytic enzymes. Est7S was purified from the crude extract of clone SM7 using Sephacryl S-200 HR and HiTrap Q column chromatographies. The purified Est7S was optimally active at 50 o C and pH 10.0. Est7S showed a high specific activity of 366.7 U/mg protein. It preferred short length esters, particularly p-nitrophenyl acetate, efficiently hydrolyzed R-and S-enantiomers of methyl-3-hydroxy-2-methylpropionate, and glyceryl tributyrate. These properties of Est7S may provide potential merits in biotechnological applications such as detergent and paper processing under alkaline conditions.

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“…5B). The completely inhibited in enzymatic activity of EstQ7 by phenylmethylsulfonyl uoride (PMSF), known as serine hydrolase inhibitor, indicating a serine residue is conserved in the pentapeptide GXSXG motif of its active site (Sameh et al 2007;Jayanath et al 2018).…”

Section: Effects Of Additives On the Activity Of Estq7mentioning

confidence: 99%

Identification and characterization of a novel carboxylesterase EstQ7 from a soil metagenomic library

et al. 2021

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A novel lipolytic gene, estq7, was identi ed from a fosmid metagenomic library. The recombinant enzyme EstQ7 consists of 370 amino acids with an anticipated molecular mass of 42 kDa. Multiple sequence alignments showed that EstQ7 contained a pentapeptide motif GHSMG, and a putative catalytic triad Ser174-Asp306-His344. Interestingly, EstQ7 was found to have very little similarity to the characterized lipolytic enzymes. Phylogenetic analysis revealed that EstQ7 may be a member of a novel family of lipolytic enzymes. Biochemical characterization of the recombinant enzyme revealed that it constitutes a slightly alkalophilic, moderate thermophilic and highly active carboxylesterase against short chain fatty acid esters with optimum temperature 50 ℃ and pH 8.2. The Km and kcat values toward p-nitrophenyl acetate were determined to be 0.17 mM and 1910 S -1 , respectively. Moreover, EstQ7 was demonstrated to have acyltransferase activity by GC-MS analysis. Homology modeling of the three-dimensional structure of this new enzyme showed that it exhibits a typical α/β hydrolase fold, and the catalytic triad residues are spatially close. Molecular docking reveled the interactions between the enzyme and the ligand. The high levels of lipolytic activity of EstQ7, combined with its moderate thermophilic property, and acyltransferase activity, render this novel enzyme a promising candidate biocatalyst for food, pharmaceutical and biotechnological applications.

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A novel solvent tolerant esterase of GDSGG motif subfamily from solar saltern through metagenomic approach: Recombinant expression and characterization

Cited by 23 publications

References 31 publications

Characterization of a Novel Moderately Thermophilic Solvent-Tolerant Esterase Isolated From a Compost Metagenome Library

Characterization of a Novel Moderately Thermophilic Solvent-Tolerant Esterase Isolated From a Compost Metagenome Library

Characterization of an alkaline esterase from an enriched metagenomic library derived from an oil-spill area

Identification and characterization of a novel carboxylesterase EstQ7 from a soil metagenomic library

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