2.A.36 The Monovalent Cation:Proton Antiporter-1 (CPA1) Family

The CPA1 family is a large family of proteins derived from Gram-positive and Gram-negative bacteria, blue-green bacteria, archaea, yeast, plants and animals. Transporters from eukaryotes have been functionally characterized, and all of these catalyze Na⁺:H⁺ exchange. Their primary physiological functions may be in (1) cytoplasmic pH regulation, extruding the H⁺ generated during metabolism, and (2) salt tolerance (in plants), due to Na⁺ uptake into vacuoles. Bacterial homologues are also Na⁺:H⁺ antiporters, but some also catalyze Li⁺:H⁺ antiport or Ca²⁺:H⁺ antiport under some conditions (Waditee et al., 2001).

The phylogenetic tree for the CPA1 family shows three principal clusters. The first cluster includes proteins derived exclusively from animals, and all of the functionally characterized members of the family belong to this cluster. Of the two remaining clusters, one includes all bacterial homologues while the other includes one from Arabidopsis thaliana, one from Homo sapiensand two from yeast (S. cerevisiaeand S. pombe). Several organisms possess multiple paralogues; for example seven paralogues are found in C. elegans, and five are known for humans. Most of these paralogues are very similar in sequence, and they belong to the animal specific cluster.

Using the mammalian NHE1 (2.A.36.1.1), it has been found that TMSs 4 and 9 as well as the extracellular loop between TMSs 3 and 4 are important for drug (amiloride- and benzoyl guanidinium-based derivatives) sensitivities. Mutations in these regions also affect transport activities. M4 and M9 therefore contain critical sites for both drug and cation recognition.

One homologue, Nhe (TC #2.A.36.1.4), is a chloride-dependent Na⁺:H⁺ antiporter in which residues 1-375 of the 438 aas are identical to Nhe-1 (TC #2.A.36.1.1). The C-terminal 63 residues are unique (Sangan et al., 2002). It is found in the apical membranes of crypt cells of the rat distal colon. This protein was reported to exhibit 6 putative TMSs and is encoded by a 2.5 kb mRNA present in many tissues (Sangan et al., 2002). However, the WHAT program predicts 10 TMSs. nhe transfected fibroblasts exhibit Cl^--dependent Na⁺-dependent intracellular pH recovery to an acid load that was blocked by 5-ethylisopropylamiloride and 5'-nitro-2-(3-phenylpropylamino)benzoate (a Cl^- channel blocker).

Numerous members of the CPA1 family have been sequenced, and these proteins vary substantially in size. The bacterial proteins have 527-549 amino acyl residues while eukaryotic proteins are generally larger, varying in size from 541-894 residues. They exhibit 10-12 putative transmembrane a-helical spanners (TMSs). A recently proposed topological model (Wakabayashi et al., 2000) suggests that in addition to 12 TMSs, a region between TMSs 9 and 10 dips into the membrane to line the pore. However, one homologue, Nhx1 of S. cerevisiae, has an extracellular glycosylated C-terminus (Wells and Rao, 2001). Some members show limited sequence similarity with members of the CPA2 family although this similarity is insufficient to establish homology. PSI-BLAST results with a single iteration provide further evidence that these two large families comprise part of a single superfamily.

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