Publications

• Structure of the formate transporter FocA reveals a pentameric aquaporin-like channel.

  Wang Y., Huang Y., Wang J., Cheng C., Huang W., Lu P., Xu Y.N., Wang P., Yan N., Shi Y.

  FocA is a representative member of the formate-nitrite transporter family, which transports short-chain acids in bacteria, archaea, fungi, algae and parasites. The structure and transport mechanism of the formate-nitrite transporter family remain unknown. Here we report the crystal structure of Es ... >> More

  FocA is a representative member of the formate-nitrite transporter family, which transports short-chain acids in bacteria, archaea, fungi, algae and parasites. The structure and transport mechanism of the formate-nitrite transporter family remain unknown. Here we report the crystal structure of Escherichia coli FocA at 2.25 A resolution. FocA forms a symmetric pentamer, with each protomer consisting of six transmembrane segments. Despite a lack of sequence homology, the overall structure of the FocA protomer closely resembles that of aquaporin and strongly argues that FocA is a channel, rather than a transporter. Structural analysis identifies potentially important channel residues, defines the channel path and reveals two constriction sites. Unlike aquaporin, FocA is impermeable to water but allows the passage of formate. A structural and biochemical investigation provides mechanistic insights into the channel activity of FocA. << Less

  Nature 462:467-472(2009) [PubMed] [EuropePMC]

• Isolation and characterization of hypophosphite-resistant mutants of Escherichia coli: identification of the FocA protein, encoded by the pfl operon, as a putative formate transporter.

  Suppmann B., Sawers G.

  Hypophosphite was used as a toxic analogue to identify genes whose products have a putative function in the transport of formate. Two Tn10-derived insertion mutants were identified that exhibited increased resistance to high concentrations of hypophosphite in the culture medium. The transposon was ... >> More

  Hypophosphite was used as a toxic analogue to identify genes whose products have a putative function in the transport of formate. Two Tn10-derived insertion mutants were identified that exhibited increased resistance to high concentrations of hypophosphite in the culture medium. The transposon was located in the identical position in the focA (formate channel; previously termed orf) gene of the pfl operon in both mutants. A defined chromosomal focA nonsense mutant, which showed minimal polarity effects on pfl gene expression, had the same phenotype as the insertion mutants. Results obtained using a hycA-lacZ fusion to monitor changes in the intracellular formate concentration in a focA mutant indicated that the level of formate inside the cell was elevated compared with the wild type. Moreover, it could be shown that there was a corresponding reduction of approximately 50% in the amount of formate excreted by a focA mutant into the culture medium. Taken together, these results indicate that formate accumulates in anaerobic cells which do not have a functional focA gene product and that one function of FocA may be to export formate from the cell. A further significant result was that hypophosphite could substitute for formate in activating hycA gene expression. This hypophosphite-dependent activation of hycA gene expression was reduced 10-fold in a focA null mutant, suggesting that hypophosphite must first enter the cell before it can act as a signal to activate hycA expression. By analogy, these data suggest that focA may also be functional in the import of formate into anaerobic Escherichia coli cells. Site-specific mutagenesis identified the translation initiation codon of focA as a GUG. Therefore, the FocA polypeptide has a molecular weight of 30,958. FocA shows significant similarity at both the primary and secondary structural levels with the NirC protein of E. coli and the FdhC protein of Methanobacterium formicicum. All three proteins are predicted to be integral membrane proteins. A detailed in vivo TnphoA mutagenesis study predicted that FocA has six membrane-spanning segments. << Less

  Mol. Microbiol. 11:965-982(1994) [PubMed] [EuropePMC]

• A single amino acid exchange converts FocA into a unidirectional efflux channel for formate.

  Kammel M., Trebbin O., Pinske C., Sawers R.G.

  During mixed-acid fermentation, <i>Escherichia coli</i> initially translocates formate out of the cell, but re-imports it at lower pH. This is performed by FocA, the archetype of the formate-nitrite transporter (FNT) family of pentameric anion channels. Each protomer of FocA has a hydrophobic pore ... >> More

  During mixed-acid fermentation, <i>Escherichia coli</i> initially translocates formate out of the cell, but re-imports it at lower pH. This is performed by FocA, the archetype of the formate-nitrite transporter (FNT) family of pentameric anion channels. Each protomer of FocA has a hydrophobic pore through which formate/formic acid is bidirectionally translocated. It is not understood how the direction of formate/formic acid passage through FocA is controlled by pH. A conserved histidine residue (H209) is located within the translocation pore, suggesting that protonation/deprotonation might be linked to the direction of formate translocation. Using a formate-responsive <i>lacZ</i>-based reporter system we monitored changes in formate levels <i>in vivo</i> when H209 in FocA was exchanged for either of the non-protonatable amino acids asparagine or glutamine, which occur naturally in some FNTs. These FocA variants (with N or Q) functioned as highly efficient formate efflux channels and the bacteria could neither accumulate formate nor produce hydrogen gas. Therefore, the data in this study suggest that this central histidine residue within the FocA pore is required for pH-dependent formate uptake into <i>E. coli</i> cells. We also address why H209 is evolutionarily conserved and provide a physiological rationale for the natural occurrence of N/Q variants of FNT channels. << Less

  Microbiology 168:0-0(2022) [PubMed] [EuropePMC]