2.A.37 The Monovalent Cation:Proton Antiporter-2 (CPA2) Family

The CPA2 family is a moderately large from bacteria, archaea and eukaryotes. Among the functionally well-characterized members of the family are (1) the KefB/KefC K⁺ efflux proteins of E. coli which may be capable of catalyzing both K⁺/H⁺ antiport and K⁺ uniport, depending on conditions (Bakker et al., 1987; Booth et al., 1996; Munro et al., 1991), (2) the Na⁺/H⁺ antiporter of Enterococcus hirae (Waser et al., 1992) and (3) the K⁺/H⁺ antiporter of S. cerevisiae. It has been proposed that under normal physiological conditions, these proteins may function by essentially the same mechanism (Reizer et al., 1992).

KefC and KefB of E. coli are responsible for glutathione-gated K⁺ efflux (Ferguson et al., 1993, 1997). Each of these proteins consists of a transmembrane hydrophobic N-terminal domain, and a less well-conserved C-terminal hydrophilic domain. Each protein interacts with a second protein encoded by genes that overlap the gene encoding the primary transporter. The KefC ancillary protein is YabF while the KefB ancillary protein is YheR. These ancillary proteins stimulate transport activity about 10-fold (Miller et al., 2000). These proteins are important for cell survival during exposure to toxic metabolites, possibly because they can release K⁺, allowing H⁺ uptake. Activation of the KefB or KefC K⁺ efflux system only occurs in the presence of glutathione and a reactive electrophile such as methylglyoxal or N-ethylmaleimide. Formation of the methylglyoxal-glutathione conjugate, S-lactoylglutathione, is catalyzed by glyoxalase I, and S-lactoylglutathione activates KefB and KefC (MacLean et al., 1998). H⁺ uptake (acidification of the cytoplasm) accompanying or following K⁺ efflux may serve as a further protective mechanism against electrophile toxicity (Booth et al., 1996; Ferguson et al., 1993, 1997; Stumpe et al., 1996). Inhibition of transport by glutathione was enhanced by NADH (Fujisawa et al., 2007).

Gram negative bacteria are protected against toxic electrophilic compounds by glutathione-gated potassium efflux systems (Kef) that modulate cytoplasmic pH. Roosild et al. (2010) have elucidated the mechanism of gating through structural and functional analysis of the E. coli KefC. The revealed mechanism can explain how subtle chemical differences in glutathione derivatives can produce opposite effects on channel function. Kef channels are regulated by potassium transport and NAD-binding (KTN) domains that sense both reduced glutathione, which inhibits Kef activity, and glutathione adducts that form during electrophile detoxification and activate Kef. Roosild et al. (2010) found that reduced glutathione stabilizes an interdomain association between two KTN folds, whereas large adducts sterically disrupt this interaction. F441 is identified as the pivotal residue discriminating between reduced glutathione and its conjugates. They demonstrated a major structural change on the binding of an activating ligand to a KTN-domain protein.

The MagA protein of Magnetospirillum sp. strain AMB-1 has been reported to be required for synthesis of bacterial magnetic particles. The magA gene is subject to transcriptional activation by an iron deficiency (Nakamura et al., 1995).  However, are more recent report has shown that magA mutants of both Magnetospirillum magneticum AMB-1 and M. gryphiswaldense MSR-1 formed wild-type-like magnetosomes without a growth defect (Uebe et al. 2012). Its transport function is not known. The GerN and GrmA proteins of Bacillus cereus and Bacillus megaterium, respectively, are spore germination proteins that can exchange Na⁺ for H⁺ and/or K⁺ (Southworth et al., 2001). The AmhT homologue of Bacillus pseudofirmus transports both K⁺ and NH₄⁺, influences ammonium homeostasis, and is required for normal sporulation and germination. The identification of these proteins as members of the CPA2 family reveals that monovalent cation transport is required for Bacillus spore formation and germination (Tani et al., 1996).

The proteins of the CPA2 family consist of between 333 and 900 amino acyl residues. They exhibit 10-14 transmembrane α-helical spanners (TMSs). Several organisms possess multiple CPA2 paralogues. Thus, E. coli has three, Methanococcus jannaschii has four and Synechocystis sp. has five paralogues. Members of the family exhibit considerable sequence divergence.

Some members of the CPA2 family show sequence similarity with certain members of the CPA1 family (TC #2.A.36), while other members of the CPA2 family show limited sequence similarity with proteins of the K⁺ transporter (Trk) family (TC #2.A.38). CPA1 and CPA2 proteins also are homologous to (but distantly related to) the β-subunits of Na⁺ transporting organic acid decarboxylases (TC #3.B.1). All four families consist of proteins in the same size range with 10 (or possibly more) putative TMSs.

The generalized transport reaction catalyzed by members of the CPA2 family is:

M⁺ (in) + nH⁺ (out) ⇌ M⁺ (out) + nH⁺ (in).

(The carrier-mediated mode)

Some members may also catalyze:

M⁺ (in) ⇌ M⁺ (out).

(The channel-mediated mode)