Na(+)/H(+) exchangers are a family of ubiquitous membrane proteins. In higher eukaryotes they regulate cytosolic pH by removing an intracellular H(+) in exchange for an extracellular Na(+). In yeast and Escherichia coli, Na(+)/H(+) exchangers function in the opposite direction to remove intracellular Na(+) in exchange for extracellular H(+). Na(+)/H(+) exchangers display an internal pH-sensitivity that varies with the different antiporter types. Only recently have investigations examined the amino acids involved in pH-sensitivity and in cation binding and transport. Histidine residues are good candidates for H(+)-sensing amino acids, since they can ionize within the physiological pH range. Histidine residues have been shown to be important in the function of the E. coli Na(+)/H(+) exchanger NhaA and in the yeast Na(+)/H(+) exchanger sod2. In E. coli, His(225) of NhaA may function to interact with, or regulate, the pH-sensory region of NhaA. In sod2, His(367) is also critical to transport and may be a functional analogue of His(225) of NhaA. Histidine residues are not critical for the function of the mammalian Na(+)/H(+) exchanger, although an unusual histidine-rich sequence of the C-terminal tail has some influence on activity. Other amino acids involved in cation binding and transport by Na(+)/H(+) exchangers are only beginning to be studied. Amino acids with polar side chains such as aspartate and glutamate have been implicated in transport activity of NhaA and sod2, but have not been studied in the mammalian Na(+)/H(+) exchanger. Further studies are needed to elucidate the mechanisms involved in pH-sensitivity and cation binding and transport by Na(+)/H(+) exchangers.