1.A.43 The Camphor Resistance or Fluoride Channel (Fluc) Family

CrcB of E. coli appears to confer camphor resistance when overexpressed and induces resA expression (Sand et al. 2003). Deletion is not lethal but increases sensitivity to camphor and fluoride. CrcB has 127 aas and 4 TMSs. Its many close homologues are small proteins, usually with 4 TMSs.  These proteins mediate fluoride efflux and resistance (Ji et al. 2014).  The active transporter is a dimer of 4 TMS subunits arranged antiparallel (Stockbridge et al. 2014).

Fluc family fluoride channels are assembled as primitive antiparallel homodimers (McIlwain et al. 2020). Crystallographic studies revealed a cation bound at the center of the protein, where it is coordinated at the dimer interface by mainchain carbonyl oxygens from the mid-membrane breaks in two corresponding TMSs.  This cation is a stably bound sodium ion, and, although it is not a transported substrate, its presence is required for the channel to adopt an open, fluoride conducting conformation. The interfacial site is selective for sodium over other cations, except for Li+, which competes with Na+ for binding, but does not support channel activity. Fluoride, F-, ubiquitous in soil, water and marine environments, is a chronic threat to microorganisms. Many bacteria, archaea, unicellular eukaryotes and plants use F- exporters to lower cytoplasmic F- levels to counteract the anion's toxicity (McIlwain et al. 2020).

Stockbridge et al. 2013 and Li et al. 2013 showed that these ''Fluc'' proteins, purified and reconstituted in liposomes and planar phospholipid bilayers, form constitutively open anion channels with extreme selectivity for F- over Cl-. The active channel is a dimer of identical or homologous subunits arranged in antiparallel transmembrane orientation, or a single protein with two internal 4TMS repeats (as for the yeast proteins (TC# 1.A.43.2.4 and 5). This dual-topological assembly had not previously been seen in ion channels but is known in multidrug transporters of the SMR sub-family of the DMT superfamily (TC# 2.A.7.1) (Pornillos et al. 2005; Pornillos and Chang 2006). 

Crystal structures showed that Fluc channels contain two separate ion-conduction pathways, each with two F- binding sites (Stockbridge et al. 2015). Last et al. 2016 examined the consequences of mutating two conserved F--coordinating phenylalanine residues. Substitution of each phenylalanine specifically extinguished its associated F- binding site in crystal structures and concomitantly inhibited F- permeation. Functional analysis of concatemeric channels, which permit mutagenic manipulation of individual pores, showed that each pore can be separately inactivated without blocking F- conduction through its symmetry-related twin. The results strongly supported a dual-pathway architecture of these channels (Last et al. 2016).



Baker, J.L., N. Sudarsan, Z. Weinberg, A. Roth, R.B. Stockbridge, and R.R. Breaker. (2012). Widespread genetic switches and toxicity resistance proteins for fluoride. Science 335: 233-235.

Hu, K.H., E. Liu, K. Dean, M. Gingras, W. DeGraff, and N.J. Trun. (1996). Overproduction of three genes leads to camphor resistance and chromosome condensation in Escherichia coli. Genetics 143: 1521-1532.

Ji, C., R.B. Stockbridge, and C. Miller. (2014). Bacterial fluoride resistance, Fluc channels, and the weak acid accumulation effect. J Gen Physiol 144: 257-261.

Johnston, N.R. and S.A. Strobel. (2019). Nitrate and Phosphate Transporters Rescue Fluoride Toxicity in Yeast. Chem Res Toxicol 32: 2305-2319.

Last, N.B., L. Kolmakova-Partensky, T. Shane, and C. Miller. (2016). Mechanistic signs of double-barreled structure in a fluoride ion channel. Elife 5:.

Li, S., K.D. Smith, J.H. Davis, P.B. Gordon, R.R. Breaker, and S.A. Strobel. (2013). Eukaryotic resistance to fluoride toxicity mediated by a widespread family of fluoride export proteins. Proc. Natl. Acad. Sci. USA 110: 19018-19023.

McIlwain, B.C., K. Martin, E.A. Hayter, and R.B. Stockbridge. (2020). An Interfacial Sodium Ion is an Essential Structural Feature of Fluc Family Fluoride Channels. J. Mol. Biol. [Epub: Ahead of Print]

Picoli, C., E. Soleilhac, A. Journet, C. Barette, M. Comte, C. Giaume, F. Mouthon, M.O. Fauvarque, and M. Charvériat. (2019). High-Content Screening Identifies New Inhibitors of Connexin 43 Gap Junctions. Assay Drug Dev Technol 17: 240-248.

Pornillos, O. and G. Chang. (2006). Inverted repeat domains in membrane proteins. FEBS Lett. 580: 358-362.

Pornillos, O., Y.J. Chen, A.P. Chen, and G. Chang. (2005). X-ray structure of the EmrE multidrug transporter in complex with a substrate. Science 310: 1950-1953.

Sand, O., M. Gingras, N. Beck, C. Hall, and N. Trun. (2003). Phenotypic characterization of overexpression or deletion of the Escherichia coli crcA, cspE and crcB genes. Microbiology 149: 2107-2117.

Smith, K.D., P.B. Gordon, A. Rivetta, K.E. Allen, T. Berbasova, C. Slayman, and S.A. Strobel. (2015). Yeast Fex1p Is a Constitutively Expressed Fluoride Channel with Functional Asymmetry of Its Two Homologous Domains. J. Biol. Chem. 290: 19874-19887.

Stockbridge, R.B., A. Koide, C. Miller, and S. Koide. (2014). Proof of dual-topology architecture of Fluc F(-) channels with monobody blockers. Nat Commun 5: 5120.

Stockbridge, R.B., J.L. Robertson, L. Kolmakova-Partensky, and C. Miller. (2013). A family of fluoride-specific ion channels with dual-topology architecture. Elife 2: e01084.

Stockbridge, R.B., L. Kolmakova-Partensky, T. Shane, A. Koide, S. Koide, C. Miller, and S. Newstead. (2015). Crystal structures of a double-barrelled fluoride ion channel. Nature 525: 548-551.


TC#NameOrganismal TypeExample

The camphor resistance protein, CrcB or FluC (Hu et al. 1996; Sand et al. 2003).  Exports fluoride selectively over chloride by an anion open channel mechanism (Stockbridge et al. 2013).  The active transporter is a dimer of 4 TMS subunits arranged in an antiparallel transmembrane orientation (Stockbridge et al. 2014).  In bacteria lacking Fluc, F- accumulates when the external medium is acidified as a predicted function of the transmembrane pH gradient. This weak acid accumulation effect, which results from the high pKa (3.4) and membrane permeability of HF, is abolished by Fluc channels (Ji et al. 2014).  A proper tubulin network is required for functional Cx43 GJ channels, and mefloquineis a gap junction inhibitor (Picoli et al. 2019).



CrcB of E. coli (P37002)


CrcB-like protein of 164 aas and 4 TMSs


CreB of Mobiluncus curtisii (Falcivibrio vaginalis)


Putative fluoride transporter of 122 aas, CrcB


CrcB of Campylobacter jejuni


CreB of 168 aas and 4 TMSs

CreB of Brachybacterium faecium


CreB of 123 aas and 4 TMSs.

CreB of Aequorivita sublithincola


CrcB, putative fluoride channel protein of 124 aas and 4 TMSs

CrcB of Lactobacillus kefiranofaciens


CrcB of 133 aas and 4 TMSs.

CrcB of Halorubrum coriense


Fluc homologue of 453 aas and 9 putative TMSs.

Fluc of Acanthamoeba castellanii


Fluoride ion channel of 128 aas and 4 TMSs, Fluc or CrcB.  The crystal structure is known (PDB5A40; 5A43).

Fluc of Bordetella pertussis


Fluoride transporter,CrcB of 122 aas and 4 TMSs (Baker et al. 2012). It is important for reducing fluoride concentrations in the cell, thus reducing its toxicity. Several of these fluoride exporter genes are regulated by fluoride-regulated riboswitches.  M. extorquens has several F- exporters that are regulated by F--riboswithches because this organims can use halogenated hydrocarbons as carbon sources, and they release the toxic halogen ion into the cytoplasm. They need to pump it out to survive. (Baker et al. 2012)

CrcB of Methylorubrum extorquens (strain DSM 6343 / CIP 106787 / DM4) (Methylobacterium extorquens)


Protein CrcB homologue 2


crcB2 of Bacillus subtilis


Putative fluoride-selective channel of 143 aas and 4 TMSs, CrcB.


CreB of Propionibacterium acnes


CreB homologue of 124 aas


CrcB homologue of Methanocaldococcus fervens


CrcB homologue of 172 aas and 4 TMSs


CrcB of Parvularcula bermudensis


Putative fluoride exporter, CrcB.


CrcB of Pelodictyon luteolum (Chlorobium luteolum)


Putative fluoride exporter, CrcB if 114 aas and 4 TMSs.


CrcB of Thermococcus barophilus


Uncharacterized protein of 151 aas and 4 TMSs.


UP of Rothia mucilaginosa (Stomatococcus mucilaginosus)


Putative fluoride exporter, CrcB of 113 aas and 4 TMSs.


UP of Haloquadratum walsbyi


TC#NameOrganismal TypeExample

CrcB-like protein of 307 aas and 6 TMSs in an apparent 3 + 3 arrangement.


CrcB homologue of Tetrahymena thermophila


Uncharacterized protein of 372 aas and 9 - 10 TMSs


UP of Kazachstania africana (Kluyveromyces africanus)


CrcB domain containing protein of 310 aas and 9 TMSs in a 4 + 5 arrangement, with both halves showing sequence similarity with the 4 TMS CrcB of E. coli.


CrcB homologue of Schizosaccharomyces cryophilus


Plasma membrane fluoride ( > chloride) export channel of 375 aas and 8 TMSs, FEX1 (Li et al. 2013). The two homologous 4 TMS domains are functionally assymetric (Smith et al. 2015).  There are two very similar fex genes in S. cerevisiae, the other having TC# 1.A.43.2.5. Fex1 is consitutively synthesized (Smith et al. 2015).


FEX1 of Saccharomyces cerevisiae


Fluoride exporter, FEX2 of 375 aas and 8 TMSs (Li et al. 2013). Overexpression of five genes in a FEX1/FEX2 deletion strain, SSU1, YHB1, IPP1, PHO87, and PHO90, concerned with nitrate and phosphate transport, increase fluoride tolerance by 2- to 10-fold (Johnston and Strobel 2019).


FEX2 of Saccharomyces cerevisiae


Fluoride exporter, FEX, of 526 aas and 10 putative TMSs (Li et al. 2013).


FEX of Neurospora crassa


Camphor resistance CrcB protein of 461 aas and 9 putative TMSs.


CrcB of Arabidopsis thaliana


Uncharacterized protein of 405 aas and 8 putative TMSs.


UP of Ciona intestinalis (Transparent sea squirt) (Ascidia intestinalis)


Uncharacterized protein of 460 aas and 9 TMSs/

UP of Phytophthora parasitica


TC#NameOrganismal TypeExample

CreB of 346 aas and 4 TMSs.

CreB of Bifidobacterium longum


Putative fluoride channel, CrcB, of 180 aas and 4 TMSs.

CrcB of Scardovia wiggsiae


Putative fluoride ion channnel, CrcB, of 178 aas and 4 TMSs

CrcB of Bifidobacterium longum


Putative fluoride ion channel, CrcB, with 310 aas and 4 TMSs.

CrcB of Bifidobacterium animalis subsp. lactis