Metagenomics and novel gene discovery

Culligan, Eamonn P.; Sleator, Roy D.; Marchesi, Julian R.; Hill, Colin

doi:10.4161/viru.27208

Cited by 100 publications

(38 citation statements)

References 145 publications

(166 reference statements)

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“…Metagenomics. In addition to focusing on individual gastrointestinal pathogens, my lab also employs a much broader metagenomics approach to identifying novel osmotolerance loci in the human gut microbiome (reviewed in 24,25 ). The human distal colon is one of the most complex and densely populated microbial ecosystems on Earth, with cell densities of up to 10 12 per gram of faeces.…”

Section: Acid Tolerancementioning

confidence: 99%

Under the microscope: From pathogens to probiotics and back

Sleator¹

2015

Bioengineered

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Section: Acid Tolerancementioning

confidence: 99%

Under the microscope: From pathogens to probiotics and back

Sleator¹

2015

Bioengineered

Self Cite

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“…According to CAZy (Carbohydrate-Active enZYmes) database classification, endoglucanases are very diverse and are part of 14 glycosyl hydrolase (GH) families12. Among the known strategies131415, metagenomics (culture independent approach) is a unique way to access the hidden information in unexplored microbial lineages and discover novel genes, metabolic pathways, and industrially important products1617 as only 0.1–1% of the microbes are culturable under laboratory conditions.…”

mentioning

confidence: 99%

Biochemical and structural characterization of a novel halotolerant cellulase from soil metagenome

Garg

Srivastava

Brahma

et al. 2016

Sci Rep

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Cellulase catalyzes the hydrolysis of β-1,4-linkages of cellulose to produce industrially relevant monomeric subunits. Cellulases find their applications in pulp and paper, laundry, food and feed, textile, brewing industry and in biofuel production. These industries always have great demand for cellulases that can work efficiently even in harsh conditions such as high salt, heat, and acidic environments. While, cellulases with high thermal and acidic stability are already in use, existence of a high halotolerant cellulase is still elusive. Here, we report a novel cellulase Cel5R, obtained from soil metagenome that shows high halotolerance and thermal stability. The biochemical and functional characterization of Cel5R revealed its endoglucanase activity and high halostability. In addition, the crystal structure of Cel5R determined at 2.2 Å resolution reveals a large number of acidic residues on the surface of the protein that contribute to the halophilic nature of this enzyme. Moreover, we demonstrate that the four free and non-conserved cysteine residues (C65, C90, C231 and C273) contributes to the thermal stability of Cel5R by alanine scanning experiments. Thus, the newly identified endoglucanase Cel5R is a promising candidate for various industrial applications.

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“…This hit rate is signicantly higher than the median rate that is observed for functional metagenomic screening. 39 We later picked the S2A24 clone from the original library and extracted its fosmid using a GeneJET plasmid miniprep kit. Sequencing and annotation of the fosmid conrmed the presence of a novel TA and 37 other unique ORFs.…”

Section: Resultsmentioning

confidence: 99%

Metagenomic discovery of a novel transaminase for valorization of monoaromatic compounds

2018

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The profitability of next-generation biorefineries is acutely contingent on the discovery and utilization of biocatalysts that can valorize lignin. To this end, the metabolic catalogues of diverse microbiota have been mined previously using functional metagenomics in order to identify biocatalysts that can selectively degrade lignin into monoaromatic compounds. Herein, we have further improved the valorization factor of biorefining by deploying functional metagenomics toward the identification of a novel transaminase that can selectively functionalize lignin-derived monoaromatics to produce value-added feedstocks for pharmaceutical synthesis.We implemented a high-throughput colorimetric assay using o-xylylenediamine as the amino donor and successfully identified a transaminase that utilizes the canonical cofactor, pyridoxal 5 0 -phosphate, to aminate as many as 14 monoaromatic aldehydes and ketones. We subsequently identified the optimal conditions for enzyme activity towards the most favoured amino acceptor, benzaldehyde, including temperature, pH and choice of co-solvent. We also evaluated the specificity of the enzyme towards a variety of amino donors, as well as the optimal concentration of the most favoured amino donor. Significantly, the novel enzyme is markedly smaller than typical transaminases, and it is stably expressed in E. coli without any modifications to its amino acid sequence. Finally, we developed and implemented a computational methodology to assess the activity of the novel transaminase. The methodology is generalizable for assessing any transaminase and facilitates in silico screening of enzyme-substrate combinations in order to develop efficient biocatalytic routes to value-added amines. The computational pipeline is an ideal complement to metagenomics and opens new possibilities for biocatalyst discovery.

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Metagenomics and novel gene discovery

Cited by 100 publications

References 145 publications

Under the microscope: From pathogens to probiotics and back

Under the microscope: From pathogens to probiotics and back

Biochemical and structural characterization of a novel halotolerant cellulase from soil metagenome

Metagenomic discovery of a novel transaminase for valorization of monoaromatic compounds

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