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High resolution AFM topographs of the Escherichia coli water channel aquaporin Z
Author(s) -
Scheuring Simon,
Ringler Philippe,
Borgnia Mario,
Stahlberg Henning,
Müller Daniel J.,
Agre Peter,
Engel Andreas
Publication year - 1999
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1093/emboj/18.18.4981
Subject(s) - aquaporin , tetramer , biology , escherichia coli , biophysics , cleavage (geology) , electron crystallography , crystallography , extracellular , protein structure , membrane , cytoplasm , biochemistry , chemistry , optics , diffraction , gene , enzyme , electron diffraction , paleontology , physics , fracture (geology)
Aquaporins form a large family of membrane channels involved in osmoregulation. Electron crystallography has shown monomers to consist of six membrane spanning α‐helices confirming sequence based predictions. Surface exposed loops are the least conserved regions, allowing differentiation of aquaporins. Atomic force microscopy was used to image the surface of aquaporin Z, the water channel of Escherichia coli . Recombinant protein with an N‐terminal fragment including 10 histidines was isolated as a tetramer by Ni‐affinity chromatography, and reconstituted into two‐dimensional crystals with p 42 1 2 symmetry. Small crystalline areas with p 4 symmetry were found as well. Imaging both crystal types before and after cleavage of the N‐termini allowed the cytoplasmic surface to be identified; a drastic change of the cytoplasmic surface accompanied proteolytic cleavage, while the extracellular surface morphology did not change. Flexibility mapping and volume calculations identified the longest loop at the extracellular surface. This loop exhibited a reversible force‐induced conformational change.