Crystallization and preliminary X-ray diffraction analysis of 5,10-methylenetetrahydrofolate dehydrogenase/cyclohydrolase from Thermoplasma acidophilum DSM 1728

Abstract

The methylenetetrahydrofolate dehydrogenase/cyclohydrolase (MTHFDC) from the thermoacidophilic archaeon Thermoplasma acidophilum is a 30.6 kDa molecular-mass enzyme that sequentially catalyzes the conversion of formyltetrahydrofolate to methylenetetrahydrofolate, with a preference for NADP as a cofactor, rather than NAD. In order to elucidate the functional and structural features of MTHFDC from archaeons at a molecular level, it was overexpressed in Escherichia coli and crystallized in the presence of its cofactor, NADP, at 295 K using polyethylene glycol (PEG) 4000 as a precipitant. The crystal is a member of the monoclinic space group P2(1), with the following unit cell parameters: a=66.333 angstrom, b=52.868 angstrom, c=86.099 angstrom, and beta= 97.570 degrees, and diffracts to a resolution of at least 2.40 angstrom at the synchrotron. Assuming a dimer in the crystallographic asymmetric unit, the calculated Matthews parameter (V-M) was 2.44 angstrom(3) /Da and the solvent content was 49.7%.