EXPRESSION, PURIFICATION, AND CHARACTERIZATION OF HUMAN CYTOSOLIC SERINE HYDROXYMETHYLTRANSFERASE

A cDNA which codes for human cytosolic serine hydroxymethyltransferase (Garrow et al., 1993, J. Biol. Chem. 268, 11910-11916) has been clone d into a pT7-7 vector as a NdeI-EcoRI insert, HMS174 (de3) cells were transformed with this plasmid and, after induction with isopropyl thio galactoside, expressed a catalytically active serine hydroxymethyltran sferase. The enzyme was purified and shown to be the expressed human e nzyme by N-terminal amino acid sequencing. About 225 mg of pure enzyme can be obtained from a 20-liter culture. Spectral characteristics of the bound pyridoxal phosphate were essentially identical to the spectr al properties of rabbit cytosolic serine hydroxymethyltransferase. Kin etic constants for the natural substrates L-serine and tetrahydrofolat e were also similar to the values obtained previously for the rabbit c ytosolic enzyme. (C) 1995 Academic Press, Inc.