A feruloyl esterase derived from a leachate metagenome library

Rashamuse, Konanani; Sanyika, Walter Tendai; Ronneburg, Tina; Brady, Dean

doi:10.5483/bmbrep.2012.45.1.14

Cited by 10 publications

(2 citation statements)

References 37 publications

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“…This could be cleaved to form mature protein (with a maximum cleavage site probability of 0.7 between Ala23 and His24). This could be a transmembrane protein as reported in similar study by Rashamuse and co-worker wherein 398 aa esterase Est C was obtained after the cleavage of 29 aa signal peptide from a protein of 423 aa (Rashamuse et al 2009 , 2011 ).…”

Section: Resultsmentioning

confidence: 60%

Isolation and in silico characterization of novel esterase gene with β-lactamase fold isolated from metagenome of north western Himalayas

Sudan

Vakhlu

2014

3 Biotech

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An esterase-producing clone Aph2 was isolated from the Apharwat soil metagenomic library, a mountain peak in NW Himalayas. ORF 2 (Est Ac) of clone Aph2 corresponds to 271 aa protein and showed 26 % sequence similarity to carboxylesterase gene of Synechococcus sp. JA-2-3B. Est Ac contains nucleophilic Ser in S68-X-X-K71 motif of β-lactamases with Tyr Y103. The conserved sequences are common with family VIII carboxylesterase and class C β-lactamase sequences. Phylogenetic analysis revealed that Est Ac sequence is closely related to esterase than to β-lactamases. In silico 3D protein structure of Est Ac was generated using MODELLER software (9.10 version). Model was generated on the basis of carboxylesterase template (PDB:1CI8) of Est B (Burkholderia gladioli) and the stereochemical parameters of the model generated were satisfactory. Docking with diisopropyl-fluorophosphate confirmed catalytic activity of Ser68 present in S-X-X-K motif.

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Section: Resultsmentioning

confidence: 60%

Isolation and in silico characterization of novel esterase gene with β-lactamase fold isolated from metagenome of north western Himalayas

Sudan

Vakhlu

2014

3 Biotech

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“…In principle, culture-independent metagenomics approaches allow for direct tapping of the entire genetic pool held within natural microbial populations, which cannot be comprehensively accessed using classical enrichment approaches [Nacke et al, 2011;Rondon et al, 2000]. The capacity of the culture-independent metagenomics approach has been demonstrated by the discovery of novel enzyme genes with potential industrial applications [Chandrasekharaiah et al, 2011;Hu et al, 2010;Rashamuse et al, 2009aRashamuse et al, , 2012.…”

Section: Introductionmentioning

confidence: 99%

Accessing Carboxylesterase Diversity from Termite Hindgut Symbionts through Metagenomics

et al. 2012

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A shotgun metagenomic library was constructed from termite hindgut symbionts and subsequently screened for esterase activities. A total of 68 recombinant clones conferring esterolytic phenotypes were identified, of which the 14 most active were subcloned and sequenced. The nucleotide lengths of the esterase-encoding open reading frames (ORFs) ranged from 783 to 2,592 bp and encoded proteins with predicted molecular masses of between 28.8 and 97.5 kDa. The highest identity scores in the GenBank database, from a global amino acid alignment ranged from 39 to 83%. The identified ORFs revealed the presence of the G-X-S-X-D, G-D-S-X, and S-X-X-K sequence motifs that have been reported to harbour a catalytic serine residue in other previously reported esterase primary structures. Five of the ORFs (EstT5, EstT7, EstT9, EstT10, and EstT12) could not be classified into any of the original eight esterase families. One of the ORFs (EstT9) showed a unique primary structure consisting of an amidohydrolase-esterase fusion. Six of the 14 esterase-encoding genes were recombinantly expressed in Escherichia coli and the purified enzymes exhibited temperature optima of between 40–50°C. Substrate-profiling studies revealed that the characterised enzymes were ‘true’ carboxylesterases based on their preferences for short to medium chain length p-nitrophenyl ester substrates. This study has demonstrated a successful application of a metagenomic approach in accessing novel esterase-encoding genes from the gut of termites that could otherwise have been missed by classical culture enrichment approaches.

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