1887

Abstract

HynSL from ‘deep ecotype’ (AltDE) is an oxygen-tolerant and thermostable [NiFe] hydrogenase. Its two structural genes (), encoding small and large hydrogenase subunits, are surrounded by eight genes (, and ) predicted to encode accessory proteins involved in maturation of the hydrogenase. A 13 kb fragment containing the ten structural and accessory genes along with three additional adjacent genes (, and ) was cloned into an IPTG-inducible expression vector and transferred into an mutant strain lacking its native hydrogenases. Upon induction, HynSL from AltDE was expressed in and was active, as determined by an hydrogen evolution assay. Subsequent genetic analysis revealed that , , and are not essential for assembling an active hydrogenase. However, and can enhance the activity of the heterologously expressed hydrogenase. We used this genetic system to compare maturation mechanisms between AltDE HynSL and its homologue. When the structural genes for the hydrogenase, , were expressed along with known accessory genes ( and ), no active hydrogenase was produced. Further, co-expression of AltDE accessory genes and with the entire set of the genes did not produce an active hydrogenase. However, co-expression of all AltDE accessory genes with the structural genes generated an active hydrogenase. This result demonstrates that the accessory genes from AltDE can complement their counterparts from and that the two hydrogenases share similar maturation mechanisms.

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2011-05-01
2020-08-09
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