Metal ions-binding T4 lysozyme as an intramolecular protein purification tag compatible with X-ray crystallography.
Phage T4 lysozyme is a well folded and highly soluble protein that is widely used as an insertion tag to improve solubility and crystallization properties of poorly behaved recombinant proteins. It has been used in the fusion protein strategy to facilitate crystallization of various proteins including multiple G protein-coupled receptors, lipid kinases, or sterol binding proteins. Here, we present a structural and biochemical characterization of its novel, metal ions-binding mutant (mbT4L). We demonstrate that mbT4L can be used as a purification tag in the immobilized-metal affinity chromatography and that, in many respects, it is superior to the conventional hexahistidine tag. In addition, structural characterization of mbT4L suggests that mbT4L can be used as a purification tag compatible with X-ray crystallography.
Institute of Organic Chemistry and Biochemistry, Czech Academy of Sciences, Prague, Czech Republic.
Evzen
E
Boura
Institute of Organic Chemistry and Biochemistry, Czech Academy of Sciences, Prague, Czech Republic.
Adriana
A
Baumlova
Institute of Organic Chemistry and Biochemistry, Czech Academy of Sciences, Prague, Czech Republic.
Dominika
D
Chalupska
Institute of Organic Chemistry and Biochemistry, Czech Academy of Sciences, Prague, Czech Republic.
Anna
A
Dubankova
Institute of Organic Chemistry and Biochemistry, Czech Academy of Sciences, Prague, Czech Republic.
Martin
M
Klima
D017122:Bacteriophage T4 / Q000201:enzymology / Q000235:genetics; D002846:Chromatography, Affinity / Q000379:methods*; D018360:Crystallography, X-Ray / Q000379:methods; D009113:Muramidase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D009154:Mutation
D009113:Muramidase
crystal structure; endolysin; histidine tag; lysozyme; phage T4; protein purification