Purification, crystallization and preliminary crystallographic analysis of the glycine-cleavage system component T-protein from Pyrococcus horikoshii OT3.

The glycine-cleavage system component T-protein is a folate-dependent enzyme that catalyzes the formation of ammonia and 5,10-CH2-tetrahydrofolate from the aminomethyl intermediate bound to the lipoate cofactor of H-protein. T-protein from Pyrococcus horikoshii OT3 has been cloned, overexpressed in Escherichia coli, purified and crystallized by the microbatch method using PEG 4000 as a precipitant at 296 K. X-ray diffraction data have been collected to 1.50 A resolution at 100 K using synchrotron radiation. The crystals belong to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 78.980, b = 95.708, c = 118.331 A. Assuming one homodimer per asymmetric unit gives a VM value of 2.4 A3 Da(-1) and a solvent content of 49.0%.