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Title: Preliminary X-ray Diffraction Analysis of the Cytoplasmic N-terminal Domain of the Na/HCO3 Cotransporter NBCe1-A

Abstract

The N-terminal cytoplasmic domain of the Na{sup +}-coupled HCO{sub 3}{sup -} cotransporter NBCe1-A (NtNBCe1) has been linked with proximal renal tubular acidosis. In a previous purification study of recombinant NtNBCe1, crystal growth at a suboptimal protein concentration (<1 mg ml{sup -1}) yielded small single diamond-shaped crystals that diffracted poorly. In the present study, by increasing the protein concentration 50-fold, the crystal size was doubled and robustness was also improved. Crystal annealing made the crystals suitable for X-ray diffraction. The crystals either belong to space group P3121 or P31 with pseudo P3121 symmetry, with unit-cell parameters a = 51.7, b = 51.7, c = 200.6 Angstroms, {alpha} = {beta} = 90, {gamma} = 120 deg, and diffract X-rays to 3.0 Angstroms resolution. The calculated Matthews number is 1.9 Angstroms{sup 3} Da{sup -1}, with two monomers of molecular weight {approx}83 kDa in the asymmetric unit. The molecular- replacement packing solution shows that the molecules form dimers by a domain-swapping mechanism.

Authors:
;
Publication Date:
Research Org.:
Brookhaven National Laboratory (BNL) National Synchrotron Light Source
Sponsoring Org.:
Doe - Office Of Science
OSTI Identifier:
914352
Report Number(s):
BNL-78920-2007-JA
TRN: US200809%%194
DOE Contract Number:  
DE-AC02-98CH10886
Resource Type:
Journal Article
Resource Relation:
Journal Name: Acta Cryst. F; Journal Volume: 62
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; ANNEALING; CRYSTAL GROWTH; DIMERS; MOLECULAR WEIGHT; MONOMERS; PROTEINS; PURIFICATION; RESOLUTION; SPACE GROUPS; SYMMETRY; X-RAY DIFFRACTION; national synchrotron light source

Citation Formats

Gill,H., and Boron, W. Preliminary X-ray Diffraction Analysis of the Cytoplasmic N-terminal Domain of the Na/HCO3 Cotransporter NBCe1-A. United States: N. p., 2006. Web. doi:10.1107/S1744309106015181.
Gill,H., & Boron, W. Preliminary X-ray Diffraction Analysis of the Cytoplasmic N-terminal Domain of the Na/HCO3 Cotransporter NBCe1-A. United States. doi:10.1107/S1744309106015181.
Gill,H., and Boron, W. Sun . "Preliminary X-ray Diffraction Analysis of the Cytoplasmic N-terminal Domain of the Na/HCO3 Cotransporter NBCe1-A". United States. doi:10.1107/S1744309106015181.
@article{osti_914352,
title = {Preliminary X-ray Diffraction Analysis of the Cytoplasmic N-terminal Domain of the Na/HCO3 Cotransporter NBCe1-A},
author = {Gill,H. and Boron, W.},
abstractNote = {The N-terminal cytoplasmic domain of the Na{sup +}-coupled HCO{sub 3}{sup -} cotransporter NBCe1-A (NtNBCe1) has been linked with proximal renal tubular acidosis. In a previous purification study of recombinant NtNBCe1, crystal growth at a suboptimal protein concentration (<1 mg ml{sup -1}) yielded small single diamond-shaped crystals that diffracted poorly. In the present study, by increasing the protein concentration 50-fold, the crystal size was doubled and robustness was also improved. Crystal annealing made the crystals suitable for X-ray diffraction. The crystals either belong to space group P3121 or P31 with pseudo P3121 symmetry, with unit-cell parameters a = 51.7, b = 51.7, c = 200.6 Angstroms, {alpha} = {beta} = 90, {gamma} = 120 deg, and diffract X-rays to 3.0 Angstroms resolution. The calculated Matthews number is 1.9 Angstroms{sup 3} Da{sup -1}, with two monomers of molecular weight {approx}83 kDa in the asymmetric unit. The molecular- replacement packing solution shows that the molecules form dimers by a domain-swapping mechanism.},
doi = {10.1107/S1744309106015181},
journal = {Acta Cryst. F},
number = ,
volume = 62,
place = {United States},
year = {Sun Jan 01 00:00:00 EST 2006},
month = {Sun Jan 01 00:00:00 EST 2006}
}