Discovery of hyperstable carbohydrate-active enzymes through metagenomics of extreme environments

Andrea Strazzulli; Beatrice Cobucci-Ponzano; Roberta Iacono; Rosa Giglio; Luisa Maurelli; Nicola Curci; Corinna Schiano-di-Cola; Annalisa Santangelo; Patrizia Contursi; Vincent Lombard; Bernard Henrissat; Federico M. Lauro; Carlos M.G.A. Fontes; Marco Moracci

doi:10.1111/febs.15080

Discovery of hyperstable carbohydrate-active enzymes through metagenomics of extreme environments

Andrea Strazzulli^*, Beatrice Cobucci-Ponzano, Roberta Iacono, Rosa Giglio, Luisa Maurelli, Nicola Curci, Corinna Schiano-di-Cola, Annalisa Santangelo, Patrizia Contursi, Vincent Lombard, Bernard Henrissat, Federico M. Lauro, Carlos M.G.A. Fontes, Marco Moracci

^*Corresponding author

Institut for Naturvidenskab og Miljø

Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › peer review

Abstract

The enzymes from hyperthermophilic microorganisms populating volcanic sites represent interesting cases of protein adaptation and biotransformations under conditions where conventional enzymes quickly denature. The difficulties in cultivating extremophiles severely limit access to this class of biocatalysts. To circumvent this problem, we embarked on the exploration of the biodiversity of the solfatara Pisciarelli, Agnano (Naples, Italy), to discover hyperthermophilic carbohydrate-active enzymes (CAZymes) and to characterize the entire set of such enzymes in this environment (CAZome). Here, we report the results of the metagenomic analysis of two mud/water pools that greatly differ in both temperature and pH (T = 85 °C and pH 5.5; T = 92 °C and pH 1.5, for Pool1 and Pool2, respectively). DNA deep sequencing and following in silico analysis led to 14 934 and 17 652 complete ORFs in Pool1 and Pool2, respectively. They exclusively belonged to archaeal cells and viruses with great genera variance within the phylum Crenarchaeota, which reflected the difference in temperature and pH of the two Pools. Surprisingly, 30% and 62% of all of the reads obtained from Pool1 and 2, respectively, had no match in nucleotide databanks. Genes associated with carbohydrate metabolism were 15% and 16% of the total in the two Pools, with 278 and 308 putative CAZymes in Pool1 and 2, corresponding to ~ 2.0% of all ORFs. Biochemical characterization of two CAZymes of a previously unknown archaeon revealed a novel subfamily GH5_19 β-mannanase/β-1,3-glucanase whose hemicellulose specificity correlates with the vegetation surrounding the sampling site, and a novel NAD⁺-dependent GH109 with a previously unreported β-N-acetylglucosaminide/β-glucoside specificity. Databases: The sequencing reads are available in the NCBI Sequence Read Archive (SRA) database under the accession numbers SRR7545549 (Pool1) and SRR7545550 (Pool2). The sequences of GH5_Pool2 and GH109_Pool2 are available in GenBank database under the accession numbers MK869723 and MK86972, respectively. The environmental data relative to Pool1 and Pool2 (NCBI BioProject PRJNA481947) are available in the Biosamples database under the accession numbers SAMN09692669 (Pool1) and SAMN09692670 (Pool2).

Originalsprog	Engelsk
Tidsskrift	FEBS Journal
Vol/bind	287
Udgave nummer	6
Sider (fra-til)	1116-1137
Antal sider	22
ISSN	1742-464X
DOI	https://doi.org/10.1111/febs.15080
Status	Udgivet - 1 mar. 2020

Bibliografisk note

Funding Information:
This work was supported by a Short Term Mobility Program of the National Research Council of Italy and a grant from the Ministero dell’Università e della Ricerca Scientifica—Industrial Research Project ‘Integrated agro‐industrial chains with high energy efficiency for the development of eco‐compatible processes of energy and biochemicals production from renewable sources and for the land valorization (Enerbio‐Chem)’ PON01_01966, funded in the frame of Operative National Programme Research and Competitiveness 2007–2013 D. D. Prot. n.01/Ric. 18.1.2010.

Link til dokument

10.1111/febs.15080

Citer dette

Strazzulli, A., Cobucci-Ponzano, B., Iacono, R., Giglio, R., Maurelli, L., Curci, N., Schiano-di-Cola, C., Santangelo, A., Contursi, P., Lombard, V., Henrissat, B., Lauro, F. M., Fontes, C. M. G. A., & Moracci, M. (2020). Discovery of hyperstable carbohydrate-active enzymes through metagenomics of extreme environments. FEBS Journal, 287(6), 1116-1137. https://doi.org/10.1111/febs.15080

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title = "Discovery of hyperstable carbohydrate-active enzymes through metagenomics of extreme environments",

abstract = "The enzymes from hyperthermophilic microorganisms populating volcanic sites represent interesting cases of protein adaptation and biotransformations under conditions where conventional enzymes quickly denature. The difficulties in cultivating extremophiles severely limit access to this class of biocatalysts. To circumvent this problem, we embarked on the exploration of the biodiversity of the solfatara Pisciarelli, Agnano (Naples, Italy), to discover hyperthermophilic carbohydrate-active enzymes (CAZymes) and to characterize the entire set of such enzymes in this environment (CAZome). Here, we report the results of the metagenomic analysis of two mud/water pools that greatly differ in both temperature and pH (T = 85 °C and pH 5.5; T = 92 °C and pH 1.5, for Pool1 and Pool2, respectively). DNA deep sequencing and following in silico analysis led to 14 934 and 17 652 complete ORFs in Pool1 and Pool2, respectively. They exclusively belonged to archaeal cells and viruses with great genera variance within the phylum Crenarchaeota, which reflected the difference in temperature and pH of the two Pools. Surprisingly, 30% and 62% of all of the reads obtained from Pool1 and 2, respectively, had no match in nucleotide databanks. Genes associated with carbohydrate metabolism were 15% and 16% of the total in the two Pools, with 278 and 308 putative CAZymes in Pool1 and 2, corresponding to ~ 2.0% of all ORFs. Biochemical characterization of two CAZymes of a previously unknown archaeon revealed a novel subfamily GH5_19 β-mannanase/β-1,3-glucanase whose hemicellulose specificity correlates with the vegetation surrounding the sampling site, and a novel NAD+-dependent GH109 with a previously unreported β-N-acetylglucosaminide/β-glucoside specificity. Databases: The sequencing reads are available in the NCBI Sequence Read Archive (SRA) database under the accession numbers SRR7545549 (Pool1) and SRR7545550 (Pool2). The sequences of GH5_Pool2 and GH109_Pool2 are available in GenBank database under the accession numbers MK869723 and MK86972, respectively. The environmental data relative to Pool1 and Pool2 (NCBI BioProject PRJNA481947) are available in the Biosamples database under the accession numbers SAMN09692669 (Pool1) and SAMN09692670 (Pool2).",

keywords = "archaea, CAZymes, extremozyme, lignocellulose, microbiome",

author = "Andrea Strazzulli and Beatrice Cobucci-Ponzano and Roberta Iacono and Rosa Giglio and Luisa Maurelli and Nicola Curci and Corinna Schiano-di-Cola and Annalisa Santangelo and Patrizia Contursi and Vincent Lombard and Bernard Henrissat and Lauro, {Federico M.} and Fontes, {Carlos M.G.A.} and Marco Moracci",

note = "Funding Information: This work was supported by a Short Term Mobility Program of the National Research Council of Italy and a grant from the Ministero dell{\textquoteright}Universit{\`a} e della Ricerca Scientifica—Industrial Research Project {\textquoteleft}Integrated agro‐industrial chains with high energy efficiency for the development of eco‐compatible processes of energy and biochemicals production from renewable sources and for the land valorization (Enerbio‐Chem){\textquoteright} PON01_01966, funded in the frame of Operative National Programme Research and Competitiveness 2007–2013 D. D. Prot. n.01/Ric. 18.1.2010. ",

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Strazzulli, A, Cobucci-Ponzano, B, Iacono, R, Giglio, R, Maurelli, L, Curci, N, Schiano-di-Cola, C, Santangelo, A, Contursi, P, Lombard, V, Henrissat, B, Lauro, FM, Fontes, CMGA & Moracci, M 2020, 'Discovery of hyperstable carbohydrate-active enzymes through metagenomics of extreme environments', FEBS Journal, bind 287, nr. 6, s. 1116-1137. https://doi.org/10.1111/febs.15080

TY - JOUR

T1 - Discovery of hyperstable carbohydrate-active enzymes through metagenomics of extreme environments

AU - Strazzulli, Andrea

AU - Cobucci-Ponzano, Beatrice

AU - Iacono, Roberta

AU - Giglio, Rosa

AU - Maurelli, Luisa

AU - Curci, Nicola

AU - Schiano-di-Cola, Corinna

AU - Santangelo, Annalisa

AU - Contursi, Patrizia

AU - Lombard, Vincent

AU - Henrissat, Bernard

AU - Lauro, Federico M.

AU - Fontes, Carlos M.G.A.

AU - Moracci, Marco

N1 - Funding Information: This work was supported by a Short Term Mobility Program of the National Research Council of Italy and a grant from the Ministero dell’Università e della Ricerca Scientifica—Industrial Research Project ‘Integrated agro‐industrial chains with high energy efficiency for the development of eco‐compatible processes of energy and biochemicals production from renewable sources and for the land valorization (Enerbio‐Chem)’ PON01_01966, funded in the frame of Operative National Programme Research and Competitiveness 2007–2013 D. D. Prot. n.01/Ric. 18.1.2010.

PY - 2020/3/1

Y1 - 2020/3/1

N2 - The enzymes from hyperthermophilic microorganisms populating volcanic sites represent interesting cases of protein adaptation and biotransformations under conditions where conventional enzymes quickly denature. The difficulties in cultivating extremophiles severely limit access to this class of biocatalysts. To circumvent this problem, we embarked on the exploration of the biodiversity of the solfatara Pisciarelli, Agnano (Naples, Italy), to discover hyperthermophilic carbohydrate-active enzymes (CAZymes) and to characterize the entire set of such enzymes in this environment (CAZome). Here, we report the results of the metagenomic analysis of two mud/water pools that greatly differ in both temperature and pH (T = 85 °C and pH 5.5; T = 92 °C and pH 1.5, for Pool1 and Pool2, respectively). DNA deep sequencing and following in silico analysis led to 14 934 and 17 652 complete ORFs in Pool1 and Pool2, respectively. They exclusively belonged to archaeal cells and viruses with great genera variance within the phylum Crenarchaeota, which reflected the difference in temperature and pH of the two Pools. Surprisingly, 30% and 62% of all of the reads obtained from Pool1 and 2, respectively, had no match in nucleotide databanks. Genes associated with carbohydrate metabolism were 15% and 16% of the total in the two Pools, with 278 and 308 putative CAZymes in Pool1 and 2, corresponding to ~ 2.0% of all ORFs. Biochemical characterization of two CAZymes of a previously unknown archaeon revealed a novel subfamily GH5_19 β-mannanase/β-1,3-glucanase whose hemicellulose specificity correlates with the vegetation surrounding the sampling site, and a novel NAD+-dependent GH109 with a previously unreported β-N-acetylglucosaminide/β-glucoside specificity. Databases: The sequencing reads are available in the NCBI Sequence Read Archive (SRA) database under the accession numbers SRR7545549 (Pool1) and SRR7545550 (Pool2). The sequences of GH5_Pool2 and GH109_Pool2 are available in GenBank database under the accession numbers MK869723 and MK86972, respectively. The environmental data relative to Pool1 and Pool2 (NCBI BioProject PRJNA481947) are available in the Biosamples database under the accession numbers SAMN09692669 (Pool1) and SAMN09692670 (Pool2).

AB - The enzymes from hyperthermophilic microorganisms populating volcanic sites represent interesting cases of protein adaptation and biotransformations under conditions where conventional enzymes quickly denature. The difficulties in cultivating extremophiles severely limit access to this class of biocatalysts. To circumvent this problem, we embarked on the exploration of the biodiversity of the solfatara Pisciarelli, Agnano (Naples, Italy), to discover hyperthermophilic carbohydrate-active enzymes (CAZymes) and to characterize the entire set of such enzymes in this environment (CAZome). Here, we report the results of the metagenomic analysis of two mud/water pools that greatly differ in both temperature and pH (T = 85 °C and pH 5.5; T = 92 °C and pH 1.5, for Pool1 and Pool2, respectively). DNA deep sequencing and following in silico analysis led to 14 934 and 17 652 complete ORFs in Pool1 and Pool2, respectively. They exclusively belonged to archaeal cells and viruses with great genera variance within the phylum Crenarchaeota, which reflected the difference in temperature and pH of the two Pools. Surprisingly, 30% and 62% of all of the reads obtained from Pool1 and 2, respectively, had no match in nucleotide databanks. Genes associated with carbohydrate metabolism were 15% and 16% of the total in the two Pools, with 278 and 308 putative CAZymes in Pool1 and 2, corresponding to ~ 2.0% of all ORFs. Biochemical characterization of two CAZymes of a previously unknown archaeon revealed a novel subfamily GH5_19 β-mannanase/β-1,3-glucanase whose hemicellulose specificity correlates with the vegetation surrounding the sampling site, and a novel NAD+-dependent GH109 with a previously unreported β-N-acetylglucosaminide/β-glucoside specificity. Databases: The sequencing reads are available in the NCBI Sequence Read Archive (SRA) database under the accession numbers SRR7545549 (Pool1) and SRR7545550 (Pool2). The sequences of GH5_Pool2 and GH109_Pool2 are available in GenBank database under the accession numbers MK869723 and MK86972, respectively. The environmental data relative to Pool1 and Pool2 (NCBI BioProject PRJNA481947) are available in the Biosamples database under the accession numbers SAMN09692669 (Pool1) and SAMN09692670 (Pool2).

KW - archaea

KW - CAZymes

KW - extremozyme

KW - lignocellulose

KW - microbiome

U2 - 10.1111/febs.15080

DO - 10.1111/febs.15080

M3 - Journal article

C2 - 31595646

AN - SCOPUS:85074341447

SN - 1742-464X

VL - 287

SP - 1116

EP - 1137

JO - F E B S Journal

JF - F E B S Journal

IS - 6

ER -