Expression, purification, and characterization of thymidylate synthase from Lactococcus lactis
Department of Biochemistry and Molecular Pharmacology
Amino Acid Sequence; Binding Sites; Crystallization; Escherichia coli; *Gene Expression; Hydrogen-Ion Concentration; Lactococcus lactis; Models, Molecular; Molecular Sequence Data; Molecular Structure; Osmolar Concentration; Recombinant Proteins; Structure-Activity Relationship; Thymidylate Synthase
Biochemistry, Biophysics, and Structural Biology | Pharmacology, Toxicology and Environmental Health
The thymidylate synthase (TS) gene from Lactococcus lactis has been highly expressed in Escherichia coli. The TS protein was purified by sequential chromatography on Q-Sepharose and phenyl-Sepharose. Six grams of cell pellet yielded 140 mg of homogeneous TS. TS is a highly conserved enzyme, and several of the conserved amino acid residues that have been implicated in catalytic function are altered in L. lactis TS. By use of a 3-dimensional homology model, we have predicted covariant changes that might compensate for these differences. With the large amounts of L. lactis TS now available, studies can be pursued to understand the structure-function relationships of this enzyme compared to other TSs and to confirm the presumed roles of the compensatory changes predicted in the homology model.
DOI of Published Version
Protein Sci. 1994 Jul;3(7):1114-6. Link to article on publisher's site
Protein science : a publication of the Protein Society
Greene, Patricia J.; Yu, Pak-Lam; Zhao, Jia; Schiffer, Celia A.; and Santi, Daniel V., "Expression, purification, and characterization of thymidylate synthase from Lactococcus lactis" (1994). Biochemistry and Molecular Pharmacology Publications and Presentations. 102.