The Aquaporin (AQP) Family

The cells of all life forms are surrounded by a membrane and filled with water. Although simple lipid bilayers exhibit limited water permeability, membranes of red cells, cells in renal proximal tubules and in certain other tissues are extremely permeable to water. Water-selective membrane channel proteins were predicted in these tissues to explain the high water permeability (low activation energy) and reversible inhibition by mercuric ions. Aquaporin-1 (AQP1), initially found in human red blood cells, was the first identified water pore. The discovery of AQP1 has led to the identification of many more (>300) family members in bacteria, fungi, plants, insects, and higher animals, including humans. Since members of the AQP family are found in almost every organism, the number of proteins belonging to this family increases rapidly as the sequences of new genomes become available. The phylogenetic tree shows two major clusters of aquaporins, the AQPs (pure water channels) and GLPs (channels for water and small non-ionic solutes). The two major families emerged billions of years ago from an ancient bacterial channel protein.

AQPs have to be highly specific for water to prevent other solutes and ions from also crossing the membrane. Protons present a particularly difficult challenge, because the positive charge of a proton can move along a column of water by hydrogen bond exchange. Since proton fluxes across cellular membranes drive physiological processes, such as membrane fusion, vesicular transport, solute transport and ATP  synthesis, proton leakage across the membrane must be avoided. Work on AQPs has significantly influenced investigations of the biophysics of water permeation across cell membranes, the physiology of fluid transport in the kidney and other organs, and the pathophysiological basis of inherited and acquired disorders of water balance. For his discovery of water channels, Peter Agre (Johns Hopkins University School of Medicine) received the 2003 Nobel Prize in Chemistry.


Aquaporin-0 (AQP0)

See Aquaporin-0 in the Cell Adhesion section.

Aquaporin-1 (AQP1)

Aquaporin-1 (AQP1), discovered in 1988, is the third most abundant protein in red blood cell membranes. It functions as a very specific water conducting pore, and it is also expressed in segments of the kidney that are known to exhibit very high water permeability. The first insight into the structure and function of AQP1 came from sequence analyses and expression studies in Xenopus laevis oocytes. The AQP1 monomer contains 269 amino acid residues, which form two tandem repeats of three membrane-spanning α-helices with amino- and carboxy-termini located on the cytoplasmic side of the membrane. In the hourglass model, connecting loops B (cytoplasmic) and E (extracellular) each contain the consensus motif Asn-Pro-Ala (NPA) and dip into the membrane from the opposite sides where they overlap, forming a single transmembrane aqueous pathway through each subunit of the AQP1 tetramer. Reconstitution of highly purified human red cell AQP1 into well-ordered membrane crystals has permitted definition of AQP1 structure at increasingly higher levels of resolution. Finally, it was possible to build a model for AQP1 into a 3.8 Å density map, generating the first atomic structure for a member of the AQP family. Multiple highly conserved amino acid residues stabilize the novel fold of AQP1. The aqueous pathway is lined with conserved hydrophobic residues that permit rapid water permeation, whereas the water selectivity is due to a constriction of the pore diameter to about 3 Å over a span of one residue. The atomic model provided a possible molecular explanation to a longstanding puzzle in physiology – how membranes can be freely permeable to water but impermeable to protons.

The 2D crystals of AQP1 were of limited size, typically 1 µm in diameter, making high-resolution data collection difficult especially at highly tilted conditions, whereas diffraction patterns from non-tilted crystals showed clear spots to a higher resolution than 3.5 Å. A helium-cooled electron microscope helped to improve the resolution of the structural analysis by reducing the effects of radiation damage. By selecting only images and electron diffraction patterns taken from well-ordered crystals, a 3D density map at a resolution of 3.8 Å in the membrane plane and 4.6 Å normal to the membrane was calculated. The AQP1 monomer contains six tilted, membrane-spanning α-helices forming a right-handed bundle. The 3.8 Å map clearly resolved protrusions corresponding to individual side chains on membrane-spanning α-helices. These permitted unambiguous assignment of the α-helical structure.


AQP1 is a homotetramer (left panels). Stability may result from accommodation of each monomer as a tight-fitting wedge within the tetramer. The relative insolubility of AQP1 in certain detergents indicates that surrounding lipids may be tightly attached. Sufficient space lies between adjacent tetramers in the 2D crystals to accommodate at least one lipid molecule that may be important in crystal formation. Beginning at the four-fold axis of the tetramer, the helices are arranged: 2-1-3 (first repeat), 5-4-6 (second repeat). In the first repeat, helices 1 and 2 run almost parallel to each other, and are tilted to approximately the same angle. Helix 3 is oriented almost perpendicular to the axis roughly defined by the top-on view of the first two helices. The highly tilted α-helices form a right-handed helical bundle, the coiled-coil interactions being stabilized by large helical crossing angles. As predicted by the hourglass model loop B dips into the membrane forming a short pore-lining helix (HB). The arrangement of the first and second repeat exhibits a pseudo two-fold axis running parallel to the membrane plane at the center of the molecule (right panels).


Blocking proton transfer is believed to require interruption of the continuous chain of hydrogen bonds along a single file of water by hydrogen-binding sites at the pore surface. The pore helices HB and HE and the neighbouring Asn residues in the two Asn-Pro-Ala motifs may be critical elements for this process. The AQP1 pore helices are oriented with the C-termini facing out. Due to the positive electrostatic field generated by the dipole moments of the pore helices in AQP1, the oxygen atom in a water molecule coming close to the membrane center orients to the side of the Asn-Pro-Ala motif (top panel). The pore helices HB and HE are held in the middle of the membrane by interactions between the Pro residues of the two Asn-Pro-Ala motifs. The amido groups of Asn 76 and 192 are fixed to extend into the pore at the constriction. The oxygen atom of the water molecule here will form hydrogen bonds with these amido groups (bottom left panel). This reorients the two hydrogen atoms of the water molecule at the pore constriction perpendicular to the channel axis because of the arrangement of the molecular orbital for water (bottom right panel). Thus, the two hydrogen atoms of the water molecule are prevented from forming hydrogen bonds with adjacent water molecules in the single-file column. The water molecule in the pore constriction can form hydrogen bonds via oxygen but not through hydrogen atoms. Thus, water molecules can permeate the pore with a minimal energy barrier, whereas transfer of protons is blocked by hydrogen-bond isolation from bulk water (H-bond isolation mechanism).

» Murata et al. (2000) Nature 407: 599-605.