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

Journal Article · Sun Apr 01 04:00:00 EDT 2007 · Acta Crystallographica. Section F

DOI:https://doi.org/10.1107/S1744309107011967· OSTI ID:22360298

Bagautdinov, Bagautdin; Matsuura, Yoshinori; Bagautdinova, Svetlana; Kunishima, Naoki ^[1]

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1. Advanced Protein Crystallography Research Group, RIKEN SPring-8 Center, Harima Institute, 1-1-1 Kouto, Sayo-cho, Sayo-gun, Hyogo 679-5148 (Japan)

A truncated form of biotin carboxyl carrier protein containing the C-terminal half fragment (BCCPΔN76) and the biotin protein ligase (BPL) with the mutation R48A (BPL*) or the double mutation R48A K111A (BPL**) were successfully cocrystallized in the presence of ATP and biotin. The BPL*–BCCPΔN76 and BPL**–BCCPΔN76 crystals belong to space group P2{sub 1} and diffract X-rays to 2.7 and 2.0 Å resolution, respectively. 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 (BCCPΔN76), 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*–BCCPΔN76 and BPL**–BCCPΔN76 complexes as well as crystals of BPL*, BPL** and BCCPΔN76 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*–BCCPΔN76 and BPL**–BCCPΔN76 crystals were collected at 100 K to 2.7 and 2.0 Å resolution, respectively, using synchrotron radiation. They belong to the monoclinic space group P2{sub 1}, with similar unit-cell parameters a = 69.85, b = 63.12, c = 75.64 Å, β = 95.9°. Assuming two subunits of the complex per asymmetric unit gives a V{sub M} value of 2.45 Å{sup 3} Da{sup −1} and a solvent content of 50%.

• OSTI ID: 22360298
• Journal Information: Acta Crystallographica. Section F, Journal Name: Acta Crystallographica. Section F Journal Issue: Pt 4 Vol. 63; ISSN ACSFCL; ISSN 1744-3091
• Country of Publication: United Kingdom
• Language: English

Related Subjects

75 CONDENSED MATTER PHYSICS
SUPERCONDUCTIVITY AND SUPERFLUIDITY
CARRIERS
CRYSTALLIZATION
CRYSTALS
INTERACTIONS
RESOLUTION
SOLVENTS
SPACE GROUPS
SYNCHROTRON RADIATION
X-RAY DIFFRACTION