Behaviour of topological marker proteins targeted to the Tat protein transport pathway

Abstract

SummaryThe Escherichia coli Tat system mediates Secindependent export of protein precursors bearing twin arginine signal peptides. Formate dehydrogenase-N is a three-subunit membrane-bound enzyme, in which localization of the FdnG subunit to the membrane is Tat dependent. FdnG was found in the periplasmic fraction of a mutant lacking the membrane anchor subunit FdnI, confirming that FdnG is located at the periplasmic face of the cytoplasmic membrane. However, the phenotypes of gene fusions between fdnG and the subcellular reporter genes phoA (encoding alkaline phosphatase) or lacZ (encoding b-galactosidase) were the opposite of those expected for analogous fusions targeted to the Sec translocase. PhoA fusion experiments have previously been used to argue that the peripheral membrane DmsAB subunits of the Tat-dependent enzyme dimethyl sulphoxide reductase are located at the cytoplasmic face of the inner membrane. Biochemical data are presented that instead show DmsAB to be at the periplasmic side of the membrane. The behaviour of reporter proteins targeted to the Tat system was analysed in more detail. These data suggest that the Tat and Sec pathways differ in their ability to transport heterologous passenger proteins. They also suggest that caution should be

Dual Topology of the Escherichia coli TatA Protein

“…Bolhuis et al (9) have constructed a TatB-TatC fusion protein by connecting the C terminus of TatB to the N terminus of TatC. Because the N terminus of TatC is located in the cytoplasm (20,35,36), the fusion should prohibit the C terminus of TatB changing the cellular location. Interestingly, the TatB-TatC fusion protein functionally complements a null mutant lacking both TatB and TatC in E. coli.…”

Extracellular respiration of dimethyl sulfoxide by Shewanella oneidensis strain MR-1

“…Accessing this source of DMSO for respiration might be difficult using traditional mechanisms. The DMSO reductase enzyme in Escherichia coli (17) and Rhodobacter spp. (18) is periplasmically located.…”

Characterization of the Menaquinone Reduction Site in the Diheme Cytochrome b Membrane Anchor of Wolinella succinogenes NiFe-hydrogenase

“…The quinone reduction inhibitor HQNO 1 is bound to FdnI in the crystal structure of the E. coli FdnGHI complex. The HQNO binding site, which is likely to be identical with the menaquinone reduction site, is situated near the cytoplasmic surface of the membrane, whereas the FdnG and FdnH subunits were shown to be oriented to the periplasmic side (7). It is therefore likely that protons required for menaquinone reduction are taken up from the cytoplasm, whereas protons generated upon formate oxidation are released into the periplasm.…”