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Expression, purification, crystallization and preliminary X‐ray crystallographic analysis of tomato β‐galactosidase 4
Author(s) -
Eda Masahiro,
Ishimaru Megumi,
Tada Toshiji
Publication year - 2015
Publication title -
acta crystallographica section f
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 37
ISSN - 2053-230X
DOI - 10.1107/s2053230x14027800
Subject(s) - crystallization , crystallography , x ray , expression (computer science) , chemistry , physics , computer science , optics , organic chemistry , programming language
Plant β‐galactosidases play important roles in carbohydrate‐reserve mobilization, cell‐wall expansion and degradation, and turnover of signalling molecules during ripening. Tomato β‐galactosidase 4 (TBG4) not only has β‐galactosidase activity but also has exo‐β‐(1,4)‐galactanase activity, and prefers β‐(1,4)‐galactans longer than pentamers as its substrates; most other β‐galactosidases only have the former activity. Recombinant TBG4 protein expressed in the yeast Pichia pastoris was crystallized by the sitting‐drop vapour‐diffusion method using PEG 10 000 as a precipitant. The crystals belonged to the orthorhombic space group P 2 1 2 1 2 1 , with unit‐parameters a = 92.82, b = 96.30, c = 159.26 Å, and diffracted to 1.65 Å resolution. Calculation of the Matthews coefficient suggested the presence of two monomers per asymmetric unit ( V M = 2.2 Å 3 Da −1 ), with a solvent content of 45%.