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Media type:
E-Article
Title:
The N‐terminal domains of the paralogous HycE and NuoCD govern assembly of the respective formate hydrogenlyase and NADH dehydrogenase complexes
Description:
<jats:p>Formate hydrogenlyase (<jats:styled-content style="fixed-case">FHL</jats:styled-content>) is the main hydrogen‐producing enzyme complex in enterobacteria. It converts formate to <jats:styled-content style="fixed-case">CO</jats:styled-content><jats:sub>2</jats:sub> and H<jats:sub>2</jats:sub> via a formate dehydrogenase and a [NiFe]‐hydrogenase. <jats:styled-content style="fixed-case">FHL</jats:styled-content> and complex I are evolutionarily related and share a common core architecture. However, complex I catalyses the fundamentally different electron transfer from <jats:styled-content style="fixed-case">NADH</jats:styled-content> to quinone and pumps protons. The catalytic <jats:styled-content style="fixed-case">FHL</jats:styled-content> subunit, HycE, resembles Nuo<jats:styled-content style="fixed-case">CD</jats:styled-content> of <jats:italic>Escherichia coli</jats:italic> complex I; a fusion of NuoC and NuoD present in other organisms. The C‐terminal domain of HycE harbours the [NiFe]‐active site and is similar to other hydrogenases, while this domain in Nuo<jats:styled-content style="fixed-case">CD</jats:styled-content> is involved in quinone binding. The N‐terminal domains of these proteins do not bind cofactors and are not involved in electron transfer. As these N‐terminal domains are separate proteins in some organisms, we removed them in <jats:italic>E. coli</jats:italic> and observed that both <jats:styled-content style="fixed-case">FHL</jats:styled-content> and complex I activities were essentially absent. This was due to either a disturbed assembly or to complex instability. Replacing the N‐terminal domain of HycE with a 180 amino acid <jats:italic>E. coli</jats:italic> NuoC protein fusion did not restore activity, indicating that the domains have complex‐specific functions. A <jats:styled-content style="fixed-case">FHL</jats:styled-content> complex in which the N‐ and C‐terminal domains of HycE were physically separated still retained most of its <jats:styled-content style="fixed-case">FHL</jats:styled-content> activity, while the separation of Nuo<jats:styled-content style="fixed-case">CD</jats:styled-content> abolished complex I activity completely. Only the <jats:styled-content style="fixed-case">FHL</jats:styled-content> complex tolerates physical separation of the HycE domains. Together, the findings strongly suggest that the N‐terminal domains of these proteins are key determinants in complex assembly.</jats:p>