Crystallization and preliminary X-ray crystallographic studies of the biotin carboxyl carrier protein and biotin protein ligase complex from Pyrococcus horikoshii OT3

Biotin protein ligase (BPL) catalyses the biotinylation of the biotin carboxyl carrier protein (BCCP) subunit of acetyl-CoA carboxylase. To elucidate the exact details of the protein-protein interactions in the biotinylation function, the C-terminal half fragment of BCCP (BCCP Delta N76), the R48A mutant of BPL (BPL*) and the R48A K111A double mutant of BPL (BPL**), all of which are from Pyrococcus horikoshii OT3, have been expressed, purified and successfully cocrystallized. Cocrystals of the BPL*-BCCP Delta N76 and BPL**-BCCP Delta N76 complexes as well as crystals of BPL*, BPL** and BCCP Delta N76 were obtained by the oil-microbatch method using PEG 20 000 as a precipitant at 295 K. Complete X-ray diffraction data sets for BPL*-BCCP Delta N76 and BPL**-BCCP Delta N76 crystals were collected at 100 K to 2.7 and 2.0 angstrom resolution, respectively, using synchrotron radiation. They belong to the monoclinic space group P2(1), with similar unit-cell parameters a = 69.85, b = 63.12, c = 75.64 angstrom, beta = 95.9 degrees. Assuming two subunits of the complex per asymmetric unit gives a V-M value of 2.45 angstrom(3) Da(-1) and a solvent content of 50%.