The N-terminal domains of the paralogous HycE and NuoCD govern assembly of the respective formate hydrogenlyase and NADH dehydrogenase complexes.

Formate hydrogenlyase (FHL) is the main hydrogen-producing enzyme complex in enterobacteria. It converts formate to CO and H via a formate dehydrogenase and a [NiFe]-hydrogenase. FHL and complex I are evolutionarily related and share a common core architecture.

Dissection of the Hydrogen Metabolism of the Enterobacterium Trabulsiella guamensis: Identification of a Formate-Dependent and Essential Formate Hydrogenlyase Complex Exhibiting Phylogenetic Similarity to Complex I.

is a nonpathogenic enterobacterium that was isolated from a vacuum cleaner on the island of Guam. It has one H-oxidizing Hyd-2-type hydrogenase (Hyd) and encodes an H-evolving Hyd that is most similar to the uncharacterized formate hydrogenlyase (FHL-2 ) complex. The FHL-2 (FHL-2 ) complex is predicted to have 5 membrane-integral and between 4 and 5 cytoplasmic subunits.

The dual-function chaperone HycH improves assembly of the formate hydrogenlyase complex.

The assembly of multi-protein complexes requires the concerted synthesis and maturation of its components and subsequently their co-ordinated interaction. The membrane-bound formate hydrogenlyase (FHL) complex is the primary hydrogen-producing enzyme in and is composed of seven subunits mostly encoded within the operon for [NiFe]-hydrogenase-3 (Hyd-3). The HycH protein is predicted to have an accessory function and is not part of the final structural FHL complex.

Identification of an Isothiocyanate on the HypEF Complex Suggests a Route for Efficient Cyanyl-Group Channeling during [NiFe]-Hydrogenase Cofactor Generation.

[NiFe]-hydrogenases catalyze uptake and evolution of H2 in a wide range of microorganisms. The enzyme is characterized by an inorganic nickel/ iron cofactor, the latter of which carries carbon monoxide and cyanide ligands. In vivo generation of these ligands requires a number of auxiliary proteins, the so-called Hyp family.

The influence of oxygen on [NiFe]-hydrogenase cofactor biosynthesis and how ligation of carbon monoxide precedes cyanation.

The class of [NiFe]-hydrogenases is characterized by a bimetallic cofactor comprising low-spin nickel and iron ions, the latter of which is modified with a single carbon monoxide (CO) and two cyanide (CN-) molecules. Generation of these ligands in vivo requires a complex maturation apparatus in which the HypC-HypD complex acts as a 'construction site' for the Fe-(CN)2CO portion of the cofactor. The order of addition of the CO and CN- ligands determines the ultimate structure and catalytic efficiency of the cofactor; however much debate surrounds the succession of events.

[NiFe]-hydrogenase maturation in vitro: analysis of the roles of the HybG and HypD accessory proteins1.

[NiFe]-hydrogenases (Hyd) bind a nickel-iron-based cofactor. The Fe ion of the cofactor is bound by two cyanide ligands and a single carbon monoxide ligand. Minimally six accessory proteins (HypA-HypF) are necessary for NiFe(CN)2CO cofactor biosynthesis in Escherichia coli.

The [NiFe]-hydrogenase accessory chaperones HypC and HybG of Escherichia coli are iron- and carbon dioxide-binding proteins.

[NiFe]-hydrogenase accessory proteins HypC and HypD form a complex that binds a Fe-(CN)₂CO moiety and CO₂. In this study two HypC homologues from Escherichia coli were purified under strictly anaerobic conditions and both contained sub-stoichiometric amounts of iron (approx. 0.

HypD is the scaffold protein for Fe-(CN)2CO cofactor assembly in [NiFe]-hydrogenase maturation.

[NiFe]-hydrogenases bind a NiFe-(CN)2CO cofactor in their catalytic large subunit. The iron-sulfur protein HypD and the small accessory protein HypC play a central role in the generation of the CO and CN(-) ligands. Infrared spectroscopy identified signatures on an anaerobically isolated HypCD complex that are reminiscent of those in the hydrogenase active site, suggesting that this complex is the assembly site of the Fe-(CN)2CO moiety of the cofactor prior to its transfer to the hydrogenase large subunit.

Metabolic deficiences revealed in the biotechnologically important model bacterium Escherichia coli BL21(DE3).

The Escherichia coli B strain BL21(DE3) has had a profound impact on biotechnology through its use in the production of recombinant proteins. Little is understood, however, regarding the physiology of this important E. coli strain.

Malfolded recombinant Tat substrates are Tat-independently degraded in Escherichia coli.

The twin-arginine translocation (Tat) system translocates folded proteins across biological membranes. It has been suggested that the Tat system of Escherichia coli can direct Tat substrates to degradation if they are not properly folded [Matos, C.F.

Tat transport of linker-containing proteins in Escherichia coli.

The twin-arginine translocation (Tat) system serves to translocate folded and often cofactor-containing proteins across biological membranes. The mechanistic limits of the Tat system can be explored by addressing the transport of specifically designed Tat substrates. It thus could be recently shown that unstructured proteins are also accepted by the Tat system, but only if they are polar on their surface.

Recombinant expression of tatABC and tatAC results in the formation of interacting cytoplasmic TatA tubes in Escherichia coli.

The twin-arginine translocation (Tat) system of bacteria and plant plastids serves to translocate folded proteins across energized biological membranes. In Escherichia coli, the three components TatA, TatB, and TatC mediate this membrane passage. Here we demonstrate that TatA can assemble to form clusters of tube-like structures in vivo.

Functional Tat transport of unstructured, small, hydrophilic proteins.

The twin-arginine translocation (Tat) system is a protein translocation system that is adapted to the translocation of folded proteins across biological membranes. An understanding of the folding requirements for Tat substrates is of fundamental importance for the elucidation of the transport mechanism. We now demonstrate for the first time Tat transport for fully unstructured proteins, using signal sequence fusions to naturally unfolded FG repeats from the yeast Nsp1p nuclear pore protein.

The TatBC complex formation suppresses a modular TatB-multimerization in Escherichia coli.

Twin-arginine translocation (Tat) systems allow the translocation of folded proteins across biological membranes of most prokaryotes. In proteobacteria, a TatBC complex binds Tat substrates and initiates their translocation after recruitment of the component TatA. TatA and TatB belong to one protein family, but only TatB forms stable complexes with TatC.

Conservation and variation between Rhodobacter capsulatus and Escherichia coli Tat systems.

The Tat system allows the translocation of folded and often cofactor-containing proteins across biological membranes. Here, we show by an interspecies transfer of a complete Tat translocon that Tat systems are largely, but not fully, interchangeable even between different classes of proteobacteria. The Tat apparatus from the alpha-proteobacterium Rhodobacter capsulatus was transferred to a Tat-deficient Escherichia coli strain, which is a gamma-proteobacterium.

YcdB from Escherichia coli reveals a novel class of Tat-dependently translocated hemoproteins.

The Tat (twin-arginine translocation) system of Escherichia coli serves to translocate folded proteins across the cytoplasmic membrane. The reasons established so far for the Tat dependence are cytoplasmic cofactor assembly and/or heterodimerization of the respective proteins. We were interested in the reasons for the Tat dependence of novel Tat substrates and focused on two uncharacterized proteins, YcdO and YcdB.

Topological studies on the twin-arginine translocase component TatC.

The twin-arginine translocation (Tat) system can translocate folded proteins across biological membranes. Among the known Tat-system components in Escherichia coli, TatC is the only protein with multiple trans-membrane domains. TatC is important for translocon interactions with Tat substrates.

Structural studies on a twin-arginine signal sequence.

Translocation of folded proteins across biological membranes can be mediated by the so-called 'twin-arginine translocation' (Tat) system. To be translocated, Tat substrates require N-terminal signal sequences which usually contain the eponymous twin-arginine motif. Here we report the first structural analysis of a twin-arginine signal sequence, the signal sequence of the high potential iron-sulfur protein from Allochromatium vinosum.