The N‐terminal domains of the paralogous HycE and NuoCD govern assembly of the respective formate hydrogenlyase and NADH dehydrogenase complexes. Issue 3 (4th February 2020)

• Record Type: Journal Article
• Authors: Skorupa, Philipp; Lindenstrauß, Ute; Burschel, Sabrina; Blumenscheit, Christian; Friedrich, Thorsten; Pinske, Constanze
• Is Part Of: FEBS open bio. Volume 10:Issue 3(2020)
• Page Start: 371
• Page End: 385
• Publication Date: 2020-02-04
• Subjects: complex I -- formate hydrogenlyase -- fusion proteins -- HycE -- NADH:ubiquinone oxidoreductase -- NuoCD; Molecular biology -- Periodicals; Cytology -- Periodicals; Life sciences -- Periodicals; Biological Science Disciplines -- Periodicals; Molecular Biology -- Periodicals; Cell Biology -- Periodicals; Cytology; Life sciences; Molecular biology; Periodicals; 572.805
• DOI: 10.1002/2211-5463.12787
• Languages: English
• ISSNs: 2211-5463

Abstract

Abstract : Formate hydrogenlyase (FHL) is the main hydrogen‐producing enzyme complex in enterobacteria. It converts formate to CO2 and H2 via a formate dehydrogenase and a [NiFe]‐hydrogenase. FHL and complex I are evolutionarily related and share a common core architecture. However, complex I catalyses the fundamentally different electron transfer from NADH to quinone and pumps protons. The catalytic FHL subunit, HycE, resembles NuoCD of Escherichia coli 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 NuoCD 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 E. coli and observed that both FHL 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 E. coli NuoC protein fusion did not restore activity, indicating that the domains have complex‐specific functions. A FHL complex in which the N‐ and C‐terminal domains of HycE were physically separated still retained most of its FHL activity, while the separation of NuoCD abolished complex I activity completely. Only the FHL 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. Abstract : Complex I and the formate hydrogenlyase (FHL) complex from Escherichia coli contain either the fusion protein NuoCD or HycE, respectively. We deleted the N‐terminal, cofactor‐free domains from both proteins or genetically separated them into two individual proteins and monitored enzyme activity. The deletion caused instability of both complexes, while after separation, FHL retained activity but complex I was inactive.