Crystallization and 1.1-A diffraction of chorismate lyase from Escherichia coli.
Chorismate pathway enzymes are important as producers of nonnucleotide aromatic compounds. The enzyme chorismate lyase from Escherichia coli has been crystallized in four distinct forms, three of which have been characterized by X-ray diffraction. Despite widespread screening, all four crystal forms grow from the same chemical conditions. The wild-type enzyme tends to aggregate, even in the presence of reducing agent, and yielded only one crystal form (monoclinic, form 1) that grew in intricate clusters. Chemical modification of the cysteines mitigated problems with aggregation and solubility but did not affect crystal growth behavior. Protein aggregation was largely eliminated by mutating the protein's two cysteines to serines. The double mutant retains full enzymatic activity and crystallizes in three new forms, one of which (triclinic) diffracts to 1.1-A resolution.
Stover
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NIST Biotechnology Division, Gaithersburg, Maryland 20899-8312, USA.
Mayhew
M P
MP
Holden
M J
MJ
Howard
A
A
Gallagher
D T
DT
D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D003460:Crystallization; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D016297:Mutagenesis, Site-Directed; D007652:Oxo-Acid-Lyases / Q000737:chemistry* / Q000235:genetics; D011487:Protein Conformation
D001426:Bacterial Proteins; D007652:Oxo-Acid-Lyases; C075960:chorismate pyruvate lyase