Genome‐Wide Screen for Escherichia coli [NiFe]‐Hydrogenase Maturation Factors
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abstract
[NiFe]‐hydrogenases catalyze the reversible oxidation of hydrogen gas at an intricate bimetallic active site to facilitate the anaerobic growth of many bacteria and archaea.(1) The maturation of this enzyme relies on a network of cellular pathways: for instance, the incorporation of nickel and iron cofactors both rely on specific metallochaperones as well as their respective metal homeostasis networks.(2) Therefore, [NiFe]‐hydrogenase activity can serve as a beacon for many related biochemical pathways. Furthermore, the ability to tune [NiFe]‐hydrogenase activity has garnered much interest for potential applications to the hydrogen economy and as a novel antibiotic target.(3,4) However, current methodologies to measure hydrogenase activity are time consuming and require specialized equipment, limiting discoveries of novel hydrogenase‐related applications. In order to answer these issues, we designed, optimized, and validated a [NiFe]‐hydrogenase assay in E. coli that is amenable to high‐throughput screening. We have applied this assay to screen the Keio collection(5) in search of the remaining genes that contribute, directly or indirectly, to [NiFe]‐hydrogenase maturation. Here we report the discovery and characterization of genes that had no previously known function.Support or Funding InformationThis work was supported in part by funding from the Natural Science and Engineering Research Council (Canada) and the Canadian Institutes of Health Research.
