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[MAGICPAH] Molecular Approaches and MetaGenomic Investigations for optimizing Clean-up of PAH contaminated sites (245226)
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5 Research products, page 1 of 1

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  • Open Access English
    Authors: 
    Mónica Martínez-Martínez; Cristina Coscolín; Gerard Santiago; Jennifer Chow; Peter J. Stogios; Rafael Bargiela; Christoph Gertler; José Navarro-Fernández; Alexander Bollinger; Stephan Thies; +32 more
    Publisher: American Chemical Society
    Country: Spain
    Project: EC | INMARE (634486), EC | KILL●SPILL (312139), EC | ULIXES (266473), NSERC , EC | MAGICPAH (245226)

    Esterases receive special attention because of their wide distribution in biological systems and environments and their importance for physiology and chemical synthesis. The prediction of esterases’ substrate promiscuity level from sequence data and the molecular reasons why certain such enzymes are more promiscuous than others remain to be elucidated. This limits the surveillance of the sequence space for esterases potentially leading to new versatile biocatalysts and new insights into their role in cellular function. Here, we performed an extensive analysis of the substrate spectra of 145 phylogenetically and environmentally diverse microbial esterases, when tested with 96 diverse esters. We determined the primary factors shaping their substrate range by analyzing substrate range patterns in combination with structural analysis and protein–ligand simulations. We found a structural parameter that helps rank (classify) the promiscuity level of esterases from sequence data at 94% accuracy. This parameter, the active site effective volume, exemplifies the topology of the catalytic environment by measuring the active site cavity volume corrected by the relative solvent accessible surface area (SASA) of the catalytic triad. Sequences encoding esterases with active site effective volumes (cavity volume/SASA) above a threshold show greater substrate spectra, which can be further extended in combination with phylogenetic data. This measure provides also a valuable tool for interrogating substrates capable of being converted. This measure, found to be transferred to phosphatases of the haloalkanoic acid dehalogenase superfamily and possibly other enzymatic systems, represents a powerful tool for low-cost bioprospecting for esterases with broad substrate ranges, in large scale sequence data sets. C.C. thanks the Spanish Ministry of Economy, Industry and Competitiveness for a Ph.D. fellowship (Grant BES-2015-073829). This project received funding from the European Union’s Horizon 2020 research and innovation program [Blue Growth: Unlocking the potential of Seas and Oceans] under grant agreement no. 634486 (project acronym INMARE). This research was also supported by the European Community Projects MAGICPAH (FP7-KBBE-2009-245226), ULIXES (FP7-KBBE-2010-266473), and KILLSPILL (FP7-KBBE2012-312139) and grants BIO2011-25012, PCIN-2014-107, BIO2014-54494-R, and CTQ2016-79138-R from the Spanish Ministry of Economy, Industry and Competitiveness. The present investigation was also funded by the Spanish Ministry of Economy, Industry and Competitiveness within the ERA NET IB2, grant no. ERA-IB-14-030 (MetaCat), the UK Biotechnology and Biological Sciences Research Council (BBSRC), grant no. BB/M029085/1, and the German Research Foundation (FOR1296). R.B. and P.N.G. acknowledge the support of the Supercomputing Wales project, which is part-funded by the European Regional Development Fund (ERDF) via the Welsh Government. O.V.G. and P.N.G. acknowledge the support of the Centre of Environmental Biotechnology Project funded by the European Regional Development Fund (ERDF) through the Welsh Government. A.Y. and A.S. gratefully acknowledge funding from Genome Canada (2009-OGI-ABC-1405) and the NSERC Strategic Network grant IBN. A.I.P. was supported by the Counseling of Economy and Employment of the Principality of Asturias, Spain (Grant FC-15-GRUPIN14-107). V.G. acknowledges the joint BSC-CRG-IRB Research Program in Computational Biology. The authors gratefully acknowledge financial support provided by the European Regional Development Fund (ERDF). We acknowledge support by the CSIC Open Access Publication Initiative through its Unit of Information Resources for Research (URICI). Peer reviewed

  • Open Access English
    Authors: 
    Ana Popovic; Tran Hai; Anatoly Tchigvintsev; Mahbod Hajighasemi; Boguslaw Nocek; Anna N. Khusnutdinova; Greg Brown; Julia Glinos; Robert Flick; Tatiana Skarina; +11 more
    Publisher: Nature Publishing Group
    Country: Italy
    Project: EC | INMARE (634486), EC | MAGICPAH (245226), EC | MICRO B3 (287589), EC | KILL●SPILL (312139), EC | ULIXES (266473), NSERC , UKRI | [14-ERA IB] MetaCat: A me... (BB/M029085/1), EC | MAMBA (226977)

    AbstractMetagenomics has made accessible an enormous reserve of global biochemical diversity. To tap into this vast resource of novel enzymes, we have screened over one million clones from metagenome DNA libraries derived from sixteen different environments for carboxylesterase activity and identified 714 positive hits. We have validated the esterase activity of 80 selected genes, which belong to 17 different protein families including unknown and cyclase-like proteins. Three metagenomic enzymes exhibited lipase activity, and seven proteins showed polyester depolymerization activity against polylactic acid and polycaprolactone. Detailed biochemical characterization of four new enzymes revealed their substrate preference, whereas their catalytic residues were identified using site-directed mutagenesis. The crystal structure of the metal-ion dependent esterase MGS0169 from the amidohydrolase superfamily revealed a novel active site with a bound unknown ligand. Thus, activity-centered metagenomics has revealed diverse enzymes and novel families of microbial carboxylesterases, whose activity could not have been predicted using bioinformatics tools.

  • Open Access
    Authors: 
    Michael Kube; Tatyana N. Chernikova; Yamal Al-Ramahi; Ana Beloqui; Nieves Lopez-Cortez; Marı A-Eugenia Guazzaroni; Hermann J. Heipieper; Sven Klages; O. R. Kotsyurbenko; Ines Langer; +26 more
    Publisher: Springer Science and Business Media LLC
    Countries: Germany, Spain
    Project: EC | MAMBA (226977), NIH | The Midwest Center for St... (3U54GM074942-04S3), NIH | Midwest Center for Struct... (3U54GM094585-02S1), EC | MICRO B3 (287589), EC | ULIXES (266473), EC | MAGICPAH (245226)

    Ubiquitous bacteria from the genus Oleispira drive oil degradation in the largest environment on Earth, the cold and deep sea. Here we report the genome sequence of Oleispira antarctica and show that compared with Alcanivorax borkumensis—the paradigm of mesophilic hydrocarbonoclastic bacteria—O. antarctica has a larger genome that has witnessed massive gene-transfer events. We identify an array of alkane monooxygenases, osmoprotectants, siderophores and micronutrient-scavenging pathways. We also show that at low temperatures, the main protein-folding machine Cpn60 functions as a single heptameric barrel that uses larger proteins as substrates compared with the classical double-barrel structure observed at higher temperatures. With 11 protein crystal structures, we further report the largest set of structures from one psychrotolerant organism. The most common structural feature is an increased content of surface-exposed negatively charged residues compared to their mesophilic counterparts. Our findings are relevant in the context of microbial cold-adaptation mechanisms and the development of strategies for oil-spill mitigation in cold environments. We acknowledge the funding from the EU Framework Program 7 to support Projects MAMBA (226977), ULIXES (266473), MAGIC PAH (245226) and MICROB3 (287589) This work received the support of the Government of Canada through Genome Canada and the Ontario Genomics Institute (grant 2009-OGI-ABC-1405 to A.F.Y. and A.S.), and the U.S. Government National Institutes of Health (grants GM074942 and GM094585 (to A.S. through Midwest Center for Structural Genomics). The study was supported by the Max Planck Society and the Deutsche Forschungsgemeinschaft through project KU 2679/2-1 and BU 890/21-1. © 2013 Macmillan Publishers Limited Peer Reviewed

  • Open Access English
    Authors: 
    Mónica Martínez-Martínez; María Alcaide; Anatoli Tchigvintsev; Oleg N. Reva; Julio Polaina; Rafael Bargiela; María-Eugenia Guazzaroni; Álvaro Chicote; Albert Canet; Francisco Valero; +4 more
    Publisher: American Society for Microbiology
    Country: Spain
    Project: EC | MAGICPAH (245226)

    The esterases and lipases from the α/β hydrolase superfamily exhibit an enormous sequence diversity, fold plasticity, and activities. Here, we present the comprehensive sequence and biochemical analyses of seven distinct esterases and lipases from the metagenome of Lake Arreo, an evaporite karstic lake in Spain (42°46=N, 2°59=W; altitude, 655 m). Together with oligonucleotide usage patterns and BLASTP analysis, our study of esterases/lipases mined from Lake Arreo suggests that its sediment contains moderately halophilic and cold-adapted proteobacteria containing DNA fragments of distantly related plasmids or chromosomal genomic islands of plasmid and phage origins. This metagenome encodes esterases/lipases with broad substrate profiles (tested over a set of 101 structurally diverse esters) and habitat-specific characteristics, as they exhibit maximal activity at alkaline pH (8.0 to 8.5) and temperature of 16 to 40°C, and they are stimulated (1.5 to 2.2 times) by chloride ions (0.1 to 1.2 M), reflecting an adaptation to environmental conditions. Our work provides further insights into the potential significance of the Lake Arreo esterases/lipases for biotechnology processes (i.e., production of enantiomers and sugar esters), because these enzymes are salt tolerant and are active at low temperatures and against a broad range of substrates. As an example, the ability of a single protein to hydrolyze triacylglycerols, (non)halogenated alkyl and aryl esters, cinnamoyl and carbohydrate esters, lactones, and chiral epoxides to a similar extent was demonstrated. We gratefully acknowledge the financial support provided by the Spanish Ministry of Economy and Competitiveness (project CSD2007-00005), the European Community project MAGICPAH (FP7-KBBE-2009-245226), the European Regional Development Fund (ERDF), and the Government of Canada through Genome Canada, Ontario Genomics Institute, and Ontario Research Fund (2009-OGI-ABC-1405 and ORF-GL2-01-004). M.-E.G. thanks the CSIC for a JAE fellowship. Peer Reviewed

  • Open Access
    Authors: 
    María Alcaide; Jesús Tornés; Peter J. Stogios; Xiaohui Xu; Christoph Gertler; Rosa Di Leo; Rafael Bargiela; Álvaro Lafraya; María-Eugenia Guazzaroni; Nieves López-Cortés; +9 more
    Publisher: Portland Press
    Country: Spain
    Project: EC | MAMBA (226977), EC | MAGICPAH (245226), EC | ULIXES (266473)

    Several members of the C-C MCP (meta-cleavage product) hydrolase family demonstrate an unusual ability to hydrolyse esters as well as the MCPs (including those from mono- and bi-cyclic aromatics). Although the molecular mechanisms responsible for such substrate promiscuity are starting to emerge, the full understanding of these complex enzymes is far from complete. In the present paper, we describe six distinct α/β hydrolases identified through genomic approaches, four of which demonstrate the unprecedented characteristic of activity towards a broad spectrum of substrates, including p-nitrophenyl, halogenated, fatty acyl, aryl, glycerol, cinnamoyl and carbohydrate esters, lactones, 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoate and 2-hydroxy-6-oxohepta-2,4-dienoate. Using structural analysis and site-directed mutagenesis we have identified the three residues (Ser32, Val130 and Trp144) that determine the unusual substrate specificity of one of these proteins, CCSP0084. The results may open up new research avenues into comparative catalytic models, structural and mechanistic studies, and biotechnological applications of MCP hydrolases. The authors gratefully acknowledge the financial support provided by the European Community projects MAGICPAH (Molecular Approaches and MetaGenomic Investigations for optimizing Clean-up of PAH contaminated sites) [number FP7-KBBE-2009-245226], ULIXES (Unravelling and exploiding Mediterranean Sea microbial diversity and ecology for xenobiotics’ and pollutants’ clean up) [number FP7-KBBE-2010-266473] and MAMBA (Marine Metagenomics for New Biotechnological Applications) [number FP7-KBBE2008-226977]. This work was further funded by the Spanish Ministry of Economy and Competitiveness (formerly MICINN) [grant number BIO2011-25012]. M.-E.G. thanks the CSIC (Consejo Superior de Investigaciones Cient´ıficas) for a JAE (Junta para la Ampliación de Estudios) fellowship. Peer Reviewed

Advanced search in
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arrow_drop_down
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[MAGICPAH] Molecular Approaches and MetaGenomic Investigations for optimizing Clean-up of PAH contaminated sites (245226)
Include:
The following results are related to Canada. Are you interested to view more results? Visit OpenAIRE - Explore.
5 Research products, page 1 of 1
  • Open Access English
    Authors: 
    Mónica Martínez-Martínez; Cristina Coscolín; Gerard Santiago; Jennifer Chow; Peter J. Stogios; Rafael Bargiela; Christoph Gertler; José Navarro-Fernández; Alexander Bollinger; Stephan Thies; +32 more
    Publisher: American Chemical Society
    Country: Spain
    Project: EC | INMARE (634486), EC | KILL●SPILL (312139), EC | ULIXES (266473), NSERC , EC | MAGICPAH (245226)

    Esterases receive special attention because of their wide distribution in biological systems and environments and their importance for physiology and chemical synthesis. The prediction of esterases’ substrate promiscuity level from sequence data and the molecular reasons why certain such enzymes are more promiscuous than others remain to be elucidated. This limits the surveillance of the sequence space for esterases potentially leading to new versatile biocatalysts and new insights into their role in cellular function. Here, we performed an extensive analysis of the substrate spectra of 145 phylogenetically and environmentally diverse microbial esterases, when tested with 96 diverse esters. We determined the primary factors shaping their substrate range by analyzing substrate range patterns in combination with structural analysis and protein–ligand simulations. We found a structural parameter that helps rank (classify) the promiscuity level of esterases from sequence data at 94% accuracy. This parameter, the active site effective volume, exemplifies the topology of the catalytic environment by measuring the active site cavity volume corrected by the relative solvent accessible surface area (SASA) of the catalytic triad. Sequences encoding esterases with active site effective volumes (cavity volume/SASA) above a threshold show greater substrate spectra, which can be further extended in combination with phylogenetic data. This measure provides also a valuable tool for interrogating substrates capable of being converted. This measure, found to be transferred to phosphatases of the haloalkanoic acid dehalogenase superfamily and possibly other enzymatic systems, represents a powerful tool for low-cost bioprospecting for esterases with broad substrate ranges, in large scale sequence data sets. C.C. thanks the Spanish Ministry of Economy, Industry and Competitiveness for a Ph.D. fellowship (Grant BES-2015-073829). This project received funding from the European Union’s Horizon 2020 research and innovation program [Blue Growth: Unlocking the potential of Seas and Oceans] under grant agreement no. 634486 (project acronym INMARE). This research was also supported by the European Community Projects MAGICPAH (FP7-KBBE-2009-245226), ULIXES (FP7-KBBE-2010-266473), and KILLSPILL (FP7-KBBE2012-312139) and grants BIO2011-25012, PCIN-2014-107, BIO2014-54494-R, and CTQ2016-79138-R from the Spanish Ministry of Economy, Industry and Competitiveness. The present investigation was also funded by the Spanish Ministry of Economy, Industry and Competitiveness within the ERA NET IB2, grant no. ERA-IB-14-030 (MetaCat), the UK Biotechnology and Biological Sciences Research Council (BBSRC), grant no. BB/M029085/1, and the German Research Foundation (FOR1296). R.B. and P.N.G. acknowledge the support of the Supercomputing Wales project, which is part-funded by the European Regional Development Fund (ERDF) via the Welsh Government. O.V.G. and P.N.G. acknowledge the support of the Centre of Environmental Biotechnology Project funded by the European Regional Development Fund (ERDF) through the Welsh Government. A.Y. and A.S. gratefully acknowledge funding from Genome Canada (2009-OGI-ABC-1405) and the NSERC Strategic Network grant IBN. A.I.P. was supported by the Counseling of Economy and Employment of the Principality of Asturias, Spain (Grant FC-15-GRUPIN14-107). V.G. acknowledges the joint BSC-CRG-IRB Research Program in Computational Biology. The authors gratefully acknowledge financial support provided by the European Regional Development Fund (ERDF). We acknowledge support by the CSIC Open Access Publication Initiative through its Unit of Information Resources for Research (URICI). Peer reviewed

  • Open Access English
    Authors: 
    Ana Popovic; Tran Hai; Anatoly Tchigvintsev; Mahbod Hajighasemi; Boguslaw Nocek; Anna N. Khusnutdinova; Greg Brown; Julia Glinos; Robert Flick; Tatiana Skarina; +11 more
    Publisher: Nature Publishing Group
    Country: Italy
    Project: EC | INMARE (634486), EC | MAGICPAH (245226), EC | MICRO B3 (287589), EC | KILL●SPILL (312139), EC | ULIXES (266473), NSERC , UKRI | [14-ERA IB] MetaCat: A me... (BB/M029085/1), EC | MAMBA (226977)

    AbstractMetagenomics has made accessible an enormous reserve of global biochemical diversity. To tap into this vast resource of novel enzymes, we have screened over one million clones from metagenome DNA libraries derived from sixteen different environments for carboxylesterase activity and identified 714 positive hits. We have validated the esterase activity of 80 selected genes, which belong to 17 different protein families including unknown and cyclase-like proteins. Three metagenomic enzymes exhibited lipase activity, and seven proteins showed polyester depolymerization activity against polylactic acid and polycaprolactone. Detailed biochemical characterization of four new enzymes revealed their substrate preference, whereas their catalytic residues were identified using site-directed mutagenesis. The crystal structure of the metal-ion dependent esterase MGS0169 from the amidohydrolase superfamily revealed a novel active site with a bound unknown ligand. Thus, activity-centered metagenomics has revealed diverse enzymes and novel families of microbial carboxylesterases, whose activity could not have been predicted using bioinformatics tools.

  • Open Access
    Authors: 
    Michael Kube; Tatyana N. Chernikova; Yamal Al-Ramahi; Ana Beloqui; Nieves Lopez-Cortez; Marı A-Eugenia Guazzaroni; Hermann J. Heipieper; Sven Klages; O. R. Kotsyurbenko; Ines Langer; +26 more
    Publisher: Springer Science and Business Media LLC
    Countries: Germany, Spain
    Project: EC | MAMBA (226977), NIH | The Midwest Center for St... (3U54GM074942-04S3), NIH | Midwest Center for Struct... (3U54GM094585-02S1), EC | MICRO B3 (287589), EC | ULIXES (266473), EC | MAGICPAH (245226)

    Ubiquitous bacteria from the genus Oleispira drive oil degradation in the largest environment on Earth, the cold and deep sea. Here we report the genome sequence of Oleispira antarctica and show that compared with Alcanivorax borkumensis—the paradigm of mesophilic hydrocarbonoclastic bacteria—O. antarctica has a larger genome that has witnessed massive gene-transfer events. We identify an array of alkane monooxygenases, osmoprotectants, siderophores and micronutrient-scavenging pathways. We also show that at low temperatures, the main protein-folding machine Cpn60 functions as a single heptameric barrel that uses larger proteins as substrates compared with the classical double-barrel structure observed at higher temperatures. With 11 protein crystal structures, we further report the largest set of structures from one psychrotolerant organism. The most common structural feature is an increased content of surface-exposed negatively charged residues compared to their mesophilic counterparts. Our findings are relevant in the context of microbial cold-adaptation mechanisms and the development of strategies for oil-spill mitigation in cold environments. We acknowledge the funding from the EU Framework Program 7 to support Projects MAMBA (226977), ULIXES (266473), MAGIC PAH (245226) and MICROB3 (287589) This work received the support of the Government of Canada through Genome Canada and the Ontario Genomics Institute (grant 2009-OGI-ABC-1405 to A.F.Y. and A.S.), and the U.S. Government National Institutes of Health (grants GM074942 and GM094585 (to A.S. through Midwest Center for Structural Genomics). The study was supported by the Max Planck Society and the Deutsche Forschungsgemeinschaft through project KU 2679/2-1 and BU 890/21-1. © 2013 Macmillan Publishers Limited Peer Reviewed

  • Open Access English
    Authors: 
    Mónica Martínez-Martínez; María Alcaide; Anatoli Tchigvintsev; Oleg N. Reva; Julio Polaina; Rafael Bargiela; María-Eugenia Guazzaroni; Álvaro Chicote; Albert Canet; Francisco Valero; +4 more
    Publisher: American Society for Microbiology
    Country: Spain
    Project: EC | MAGICPAH (245226)

    The esterases and lipases from the α/β hydrolase superfamily exhibit an enormous sequence diversity, fold plasticity, and activities. Here, we present the comprehensive sequence and biochemical analyses of seven distinct esterases and lipases from the metagenome of Lake Arreo, an evaporite karstic lake in Spain (42°46=N, 2°59=W; altitude, 655 m). Together with oligonucleotide usage patterns and BLASTP analysis, our study of esterases/lipases mined from Lake Arreo suggests that its sediment contains moderately halophilic and cold-adapted proteobacteria containing DNA fragments of distantly related plasmids or chromosomal genomic islands of plasmid and phage origins. This metagenome encodes esterases/lipases with broad substrate profiles (tested over a set of 101 structurally diverse esters) and habitat-specific characteristics, as they exhibit maximal activity at alkaline pH (8.0 to 8.5) and temperature of 16 to 40°C, and they are stimulated (1.5 to 2.2 times) by chloride ions (0.1 to 1.2 M), reflecting an adaptation to environmental conditions. Our work provides further insights into the potential significance of the Lake Arreo esterases/lipases for biotechnology processes (i.e., production of enantiomers and sugar esters), because these enzymes are salt tolerant and are active at low temperatures and against a broad range of substrates. As an example, the ability of a single protein to hydrolyze triacylglycerols, (non)halogenated alkyl and aryl esters, cinnamoyl and carbohydrate esters, lactones, and chiral epoxides to a similar extent was demonstrated. We gratefully acknowledge the financial support provided by the Spanish Ministry of Economy and Competitiveness (project CSD2007-00005), the European Community project MAGICPAH (FP7-KBBE-2009-245226), the European Regional Development Fund (ERDF), and the Government of Canada through Genome Canada, Ontario Genomics Institute, and Ontario Research Fund (2009-OGI-ABC-1405 and ORF-GL2-01-004). M.-E.G. thanks the CSIC for a JAE fellowship. Peer Reviewed

  • Open Access
    Authors: 
    María Alcaide; Jesús Tornés; Peter J. Stogios; Xiaohui Xu; Christoph Gertler; Rosa Di Leo; Rafael Bargiela; Álvaro Lafraya; María-Eugenia Guazzaroni; Nieves López-Cortés; +9 more
    Publisher: Portland Press
    Country: Spain
    Project: EC | MAMBA (226977), EC | MAGICPAH (245226), EC | ULIXES (266473)

    Several members of the C-C MCP (meta-cleavage product) hydrolase family demonstrate an unusual ability to hydrolyse esters as well as the MCPs (including those from mono- and bi-cyclic aromatics). Although the molecular mechanisms responsible for such substrate promiscuity are starting to emerge, the full understanding of these complex enzymes is far from complete. In the present paper, we describe six distinct α/β hydrolases identified through genomic approaches, four of which demonstrate the unprecedented characteristic of activity towards a broad spectrum of substrates, including p-nitrophenyl, halogenated, fatty acyl, aryl, glycerol, cinnamoyl and carbohydrate esters, lactones, 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoate and 2-hydroxy-6-oxohepta-2,4-dienoate. Using structural analysis and site-directed mutagenesis we have identified the three residues (Ser32, Val130 and Trp144) that determine the unusual substrate specificity of one of these proteins, CCSP0084. The results may open up new research avenues into comparative catalytic models, structural and mechanistic studies, and biotechnological applications of MCP hydrolases. The authors gratefully acknowledge the financial support provided by the European Community projects MAGICPAH (Molecular Approaches and MetaGenomic Investigations for optimizing Clean-up of PAH contaminated sites) [number FP7-KBBE-2009-245226], ULIXES (Unravelling and exploiding Mediterranean Sea microbial diversity and ecology for xenobiotics’ and pollutants’ clean up) [number FP7-KBBE-2010-266473] and MAMBA (Marine Metagenomics for New Biotechnological Applications) [number FP7-KBBE2008-226977]. This work was further funded by the Spanish Ministry of Economy and Competitiveness (formerly MICINN) [grant number BIO2011-25012]. M.-E.G. thanks the CSIC (Consejo Superior de Investigaciones Cient´ıficas) for a JAE (Junta para la Ampliación de Estudios) fellowship. Peer Reviewed