Forefronts in Nephrology 399

Molecular basis of water selectivity on aquaporin-1

KAZUYOSHI MURATA, KAORU MITSUOKA, TERAHISA HIRAI, THOMAS WALZ, PETER AGRE,J. BERNARD HEYMANN, ANDREAS ENGEL, and YOSHINORI FUJIYOSHI1

Physiological Institute, Okazaki, and Department of Biophysics, Faculty of Science, Kyoto University, Kyoto, Japan;Department of Cell Biology, Harvard Medical School, Boston, Massachusetts, Department of BiologicalChemistry and Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA; andM.E. Muller-Institute for Microscopic Structural Biology at Biozentrum, University of Basel, Basel, Switzerland

the cytoplasmic surface. The remainder of loop B formsWe analyzed the structure of aquaporin-1 (AQP1)the cytoplasmic connection to helix 3, which crosses theby electron crystallography. The scaffold of the AQP1membrane adjacent to helix 1, on the opposite side frommolecule was formed by two helical bundles of 1-3 andhelix 2. Helix 3 is the last structural element of the first4-6. These two helical bundles are unusual in that theAQP1 repeat that terminates on the extracellular sur-three helices form a roughly linear arrangement but notface. The second AQP1 repeat adopts essentially theaccording to their position in the primary structure, thatsame fold as the first and the two repeats are connectedis, 1-2-3 and 4-5-6, but the first helix of each bundle isby loop C.sandwiched between the other two helices of the bundle,

The analyzed structure reveals key amino acid resi-that is, 2-1-3 and 5-4-6. Two helices of each bundle,dues that are conserved throughout the AQP family andhelices 1 and 2 of the first bundle and helices 4 and 5 ofstabilize a novel and unusual fold of AQP1. The waterthe second bundle, run almost parallel to each other,pathway is lined with conserved hydrophobic residuestilting roughly in the same direction in the membrane.that serve for the rapid water transport, whereas theHowever, the third helix of the first bundle, helix 3, iswater selectivity is due to a constriction of the small poreoriented perpendicular to the axis defined by the firstdiameter over a length of only one amino acid residue.two helices of the bundle, and the same is found for

helix 6 in the second helical bundle. The short helix The structure enables us to theorize that it is a molecularHB from the first AQP1 repeat strongly interacts with mechanism for the effective blocking of proton transporttransmembrane helix 6 from the second AQP1 repeat, while maintaining a high water permeability, thus offer-while the short helix HE from the second AQP1 repeat ing an explanation of a long-standing puzzle.interacts with transmembrane helix 3 from the firstAQP1 repeat. This cross-interaction leads to an intimate BIBLIOGRAPHYlink of the two protein halves. de Groot BL, Heymann JB, Engel A, et al: The fold of human

After helix 2, which crosses the membrane adjacent aquaporin 1. J Mol Biol 300:987–994, 2000Engel A, Fujiyoshi Y, Agre P: The importance of aquaporin waterto helix 1, near the fourfold axis of the AQP1 tetramer,

channel protein structures. EMBO J 19:800–806, 2000loop B, immediately folds back into the membrane posi-Fujiyoshi Y: The structural study of membrane proteins by electron

tioning its NPA motif is in the middle between the two crystallography. Adv Biophys 35:25–80, 1998Mitsuoka K, Murata K, Walz T, et al: The structure of aquaporin-1helical bundles 1-3 and 4-6, close to the center of the

at 4.5-A resolution reveals short alpha-helices in the center of thelipid bilayer. After the NPA motif, loop B forms the shortmonomer. J Struct Biol 128:34–43, 1999

�-helix HB, which runs close to helix 6 of the second Murata K, Mitsuoka K, Hirai T, et al: Structural determinants ofwater permeation through aquaporin-1. Nature 407:599–605, 2000AQP1 repeat and brings the polypeptide chain back to

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