Structure of the water channel AgpZ from Escherichia coli revealed by electron crystallography

P. Ringler; M. J. Borgnia; H. Stahlberg; P. C. Maloney; P. Agre; A. Engel

doi:10.1006/jmbi.1999.3031

Structure of the water channel AgpZ from Escherichia coli revealed by electron crystallography

P. Ringler, M. J. Borgnia, H. Stahlberg, P. C. Maloney, P. Agre, A. Engel

Research output: Contribution to journal › Article › peer-review

56 Scopus citations

Abstract

Molecular water channels (aquaporins) allow living cells to adapt to osmotic variations by rapid and specific diffusion of water molecules. Aquaporins are present in animals, plants, algae, fungi and bacteria. Here we present an electron microscopic analysis of the most ancient water channel described so far: the aquaporin Z (AqpZ) of Escherichia coli. A recombinant AqpZ with a poly(histidine) tag at the N terminus has been constructed, overexpressed and purified to homogeneity. Solubilized with octylglucoside, the purified AqpZ remains associated as a homotetramer, and assembles into highly ordered two-dimensional tetragonal crystals with unit cell dimensions a = b = 95 Å, γ = 90°when reconstituted by dialysis in the presence of lipids. Three-dimensional reconstruction of negatively stained lattices revealed the p42₁2 packing arrangement that is also observed with the human erythrocyte water channel (AQP1). The 8 Å projection map of the AqpZ tetramer in frozen hydrated samples is similar to that of AQP1, consistent with the high sequence homology between these proteins.

Original language	English (US)
Pages (from-to)	1181-1190
Number of pages	10
Journal	Journal of molecular biology
Volume	291
Issue number	5
DOIs	https://doi.org/10.1006/jmbi.1999.3031
State	Published - Sep 3 1999

Keywords

2D crystallization
Aquaporin
Electron crystallography
Major integral protein family
STEM

ASJC Scopus subject areas

Structural Biology
Molecular Biology

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10.1006/jmbi.1999.3031

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title = "Structure of the water channel AgpZ from Escherichia coli revealed by electron crystallography",

abstract = "Molecular water channels (aquaporins) allow living cells to adapt to osmotic variations by rapid and specific diffusion of water molecules. Aquaporins are present in animals, plants, algae, fungi and bacteria. Here we present an electron microscopic analysis of the most ancient water channel described so far: the aquaporin Z (AqpZ) of Escherichia coli. A recombinant AqpZ with a poly(histidine) tag at the N terminus has been constructed, overexpressed and purified to homogeneity. Solubilized with octylglucoside, the purified AqpZ remains associated as a homotetramer, and assembles into highly ordered two-dimensional tetragonal crystals with unit cell dimensions a = b = 95 {\AA}, γ = 90°when reconstituted by dialysis in the presence of lipids. Three-dimensional reconstruction of negatively stained lattices revealed the p4212 packing arrangement that is also observed with the human erythrocyte water channel (AQP1). The 8 {\AA} projection map of the AqpZ tetramer in frozen hydrated samples is similar to that of AQP1, consistent with the high sequence homology between these proteins.",

keywords = "2D crystallization, Aquaporin, Electron crystallography, Major integral protein family, STEM",

author = "P. Ringler and Borgnia, {M. J.} and H. Stahlberg and Maloney, {P. C.} and P. Agre and A. Engel",

note = "Funding Information: The work of Ms Bettina Wolpensinger who operated the STEM to record dark-field micrographs for mass determination and the expert assistance of Dr Shirley M{\"u}ller are gratefully acknowledged. The authors are indebted to Dr J. Bernard Heymann for critical proofreading of the manuscript and helpful discussion. The work was supported by the M.E. M{\"u}ller Foundation of Switzerland and the Swiss National Foundation (grant no. 31–42435.94 to A.E.). P.R. thanks the French INSERM, and the Kanton Basel for post-doctoral fellowships.",

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doi = "10.1006/jmbi.1999.3031",

language = "English (US)",

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T1 - Structure of the water channel AgpZ from Escherichia coli revealed by electron crystallography

AU - Ringler, P.

AU - Borgnia, M. J.

AU - Stahlberg, H.

AU - Maloney, P. C.

AU - Agre, P.

AU - Engel, A.

N1 - Funding Information: The work of Ms Bettina Wolpensinger who operated the STEM to record dark-field micrographs for mass determination and the expert assistance of Dr Shirley Müller are gratefully acknowledged. The authors are indebted to Dr J. Bernard Heymann for critical proofreading of the manuscript and helpful discussion. The work was supported by the M.E. Müller Foundation of Switzerland and the Swiss National Foundation (grant no. 31–42435.94 to A.E.). P.R. thanks the French INSERM, and the Kanton Basel for post-doctoral fellowships.

PY - 1999/9/3

Y1 - 1999/9/3

N2 - Molecular water channels (aquaporins) allow living cells to adapt to osmotic variations by rapid and specific diffusion of water molecules. Aquaporins are present in animals, plants, algae, fungi and bacteria. Here we present an electron microscopic analysis of the most ancient water channel described so far: the aquaporin Z (AqpZ) of Escherichia coli. A recombinant AqpZ with a poly(histidine) tag at the N terminus has been constructed, overexpressed and purified to homogeneity. Solubilized with octylglucoside, the purified AqpZ remains associated as a homotetramer, and assembles into highly ordered two-dimensional tetragonal crystals with unit cell dimensions a = b = 95 Å, γ = 90°when reconstituted by dialysis in the presence of lipids. Three-dimensional reconstruction of negatively stained lattices revealed the p4212 packing arrangement that is also observed with the human erythrocyte water channel (AQP1). The 8 Å projection map of the AqpZ tetramer in frozen hydrated samples is similar to that of AQP1, consistent with the high sequence homology between these proteins.

AB - Molecular water channels (aquaporins) allow living cells to adapt to osmotic variations by rapid and specific diffusion of water molecules. Aquaporins are present in animals, plants, algae, fungi and bacteria. Here we present an electron microscopic analysis of the most ancient water channel described so far: the aquaporin Z (AqpZ) of Escherichia coli. A recombinant AqpZ with a poly(histidine) tag at the N terminus has been constructed, overexpressed and purified to homogeneity. Solubilized with octylglucoside, the purified AqpZ remains associated as a homotetramer, and assembles into highly ordered two-dimensional tetragonal crystals with unit cell dimensions a = b = 95 Å, γ = 90°when reconstituted by dialysis in the presence of lipids. Three-dimensional reconstruction of negatively stained lattices revealed the p4212 packing arrangement that is also observed with the human erythrocyte water channel (AQP1). The 8 Å projection map of the AqpZ tetramer in frozen hydrated samples is similar to that of AQP1, consistent with the high sequence homology between these proteins.

KW - 2D crystallization

KW - Aquaporin

KW - Electron crystallography

KW - Major integral protein family

KW - STEM

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JO - Journal of molecular biology

JF - Journal of molecular biology

IS - 5

ER -

Structure of the water channel AgpZ from Escherichia coli revealed by electron crystallography

Abstract

Keywords

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