Open AccessOverexpression, crystallization and preliminary X‐ray crystallographic analysis of the C‐terminal cytosolic domain of mouse anoctamin 1
Author(s) -
Park Sang Ho,
Chung Ho Kyung,
Kim Do Jin,
Han Mi Ra,
Park Mi Seul,
Oh Uhtaek,
Kim HyunJung,
Han Byung Woo
Publication year - 2011
Publication title -
acta crystallographica section f
Language(s) - English
Resource type - Journals
ISSN - 1744-3091
DOI - 10.1107/s1744309111027989
Subject(s) - crystallization , crystallography , terminal (telecommunication) , domain (mathematical analysis) , cytosol , chemistry , microbiology and biotechnology , biology , biochemistry , computer science , enzyme , mathematics , organic chemistry , mathematical analysis , telecommunications
Transmembrane protein 16A (TMEM16A, also known as anoctamin 1; ANO1) is a bona fide Ca 2+ ‐activated chloride channel that is activated by intracellular Ca 2+ ‐ and Ca 2+ ‐mobilizing stimuli and plays important roles in a variety of physiological functions. To elucidate the structural features of ANO1, structural analysis of the C‐terminal cytosolic domain of mouse ANO1 (mANO1‐CTD) was initiated. mANO1‐CTD was overexpressed in Escherichia coli and was crystallized at 297 K using a reservoir solution consisting of 0.2 M sodium acetate trihydrate, 0.1 M Tris–HCl pH 8.5 and 30%( w / v ) PEG 4000. X‐ray diffraction data were collected to 2.3 Å resolution. The crystals belonged to the orthorhombic space group P 2 1 2 1 2 1 , with unit‐cell parameters a = 73.96, b = 103.73, c = 114.71 Å. If it is assumed that eight copies of a monomer molecule are present in the crystallographic asymmetric unit, the crystal volume per protein mass ( V M ) is 2.38 Å 3 Da −1 and the solvent content is 48.38%. Attempts to solve the structure of mANO1‐CTD by the MAD method using selenomethionine‐labelled mANO1‐CTD or heavy‐atom‐derivatized crystals are in progress.