The purification, crystallization and preliminary diffraction of a glycerophosphodiesterase from Enterobacter aerogenes.
The metallo-glycerophosphodiesterase from Enterobacter aerogenes (GpdQ) has been cloned, expressed in Escherichia coli and purified. Initial screening of crystallization conditions for this enzyme resulted in the identification of needles from one condition in a sodium malonate grid screen. Removal of the metals from the enzyme and subsequent optimization of these conditions led to crystals that diffracted to 2.9 angstroms and belonged to space group P2(1)3, with unit-cell parameter a = 164.1 angstroms. Self-rotation function analysis and V(M) calculations indicated that the asymmetric unit contains two copies of the monomeric enzyme, corresponding to a solvent content of 79%. It is intended to determine the structure of this protein utilizing SAD phasing from transition metals or molecular replacement.
Jackson
Colin J
CJ
The Research School of Chemistry, Australian National University, ACT 0200, Australia.
Carr
Paul D
PD
Kim
Hye Kyung
HK
Liu
Jian Wei
JW
Ollis
David L
DL
D001426:Bacterial Proteins / Q000737:chemistry / Q000302:isolation & purification; D003001:Cloning, Molecular; D003460:Crystallization; D021902:Enterobacter aerogenes / Q000201:enzymology*; D004926:Escherichia coli / Q000201:enzymology; D008958:Models, Molecular; D010727:Phosphoric Diester Hydrolases / Q000737:chemistry* / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification; D014961:X-Ray Diffraction
D001426:Bacterial Proteins; D011994:Recombinant Proteins; D010727:Phosphoric Diester Hydrolases; C038133:glycerophosphodiester phosphodiesterase