Abstract
Hydrogen is one of the most common elements on Earth. The enzymes converting molecular hydrogen into protons and electrons are the hydrogenases. Hydrogenases are ubiquitously distributed in all three domains of life where they play a central role in cell metabolism. So far, the recovery of hydrogenases has been restricted to culture-dependent and sequence-based approaches. We have recently developed the only activity-based screen for seeking H2-uptake enzymes from metagenomes without having to rely on enrichment and isolation of hydrogen-oxidizing microorganisms or prior metagenomic sequencing. When screening 14,400 fosmid clones from three hydrothermal vent metagenomes using this solely activity-based approach, four clones with H2-uptake activity were identified with specific activities of up to 258 ± 19 nmol H2/min/mg protein of partially purified membrane fractions. The respective metagenomic fragments exhibited mostly very low or no similarities to sequences in the public databases. A search with hidden Markov models for different hydrogenase groups showed no hits for three of the four metagenomic inserts, indicating that they do not encode for classical hydrogenases. Our activity-based screen serves as a powerful tool for the discovery of (novel) hydrogenases which would not have been identified by the currently available techniques. This screen can be ideally combined with culture- and sequence-based approaches to investigate the tremendous hydrogen-converting potential in the environment.
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Acknowledgements
We thank the Captain and crews of the R/V Meteor and ROV Kiel6000 for helping us to obtain hydrothermal vent samples. Furthermore, we are indebted to Dr. Kai Thormann for providing the suicide vector pNPTS138R6KT to construct the mutant and the Shewanella oneidensis MR-1 strain as well as Dr. Harald Huber for providing the A. aeolicus DNA. We also greatly appreciate that Prof. Svein Valla supplied the pRS44 broad host range fosmid vector and the appendant pTA66 plasmid. We thank Nicolas Rychlik for constructing the S. oneidensis ΔhyaB mutant and for support with developing the screen as well as numerous technical assistant-students for picking clones. We also thank Dominik Danso for help with performing the hidden Markov-Model searches.
Funding
This work was supported by the research grant DFG PE1549-6/1 from the German Science Foundation.
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Adam, N., Perner, M. Novel hydrogenases from deep-sea hydrothermal vent metagenomes identified by a recently developed activity-based screen. ISME J 12, 1225–1236 (2018). https://doi.org/10.1038/s41396-017-0040-6
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DOI: https://doi.org/10.1038/s41396-017-0040-6
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