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

Abstract

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-cellmore » 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%.« less

Authors:
; ; ;  [1]
  1. Advanced Protein Crystallography Research Group, RIKEN SPring-8 Center, Harima Institute, 1-1-1 Kouto, Sayo-cho, Sayo-gun, Hyogo 679-5148 (Japan)
Publication Date:
OSTI Identifier:
22360298
Resource Type:
Journal Article
Journal Name:
Acta Crystallographica. Section F
Additional Journal Information:
Journal Volume: 63; Journal Issue: Pt 4; Other Information: PMCID: PMC2330208; PMID: 17401210; PUBLISHER-ID: fw5133; OAI: oai:pubmedcentral.nih.gov:2330208; Copyright (c) International Union of Crystallography 2007; Country of input: International Atomic Energy Agency (IAEA); Journal ID: ISSN 1744-3091
Country of Publication:
United Kingdom
Language:
English
Subject:
75 CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND SUPERFLUIDITY; CARRIERS; CRYSTALLIZATION; CRYSTALS; INTERACTIONS; RESOLUTION; SOLVENTS; SPACE GROUPS; SYNCHROTRON RADIATION; X-RAY DIFFRACTION

Citation Formats

Bagautdinov, Bagautdin, Matsuura, Yoshinori, Bagautdinova, Svetlana, and Kunishima, Naoki. Crystallization and preliminary X-ray crystallographic studies of the biotin carboxyl carrier protein and biotin protein ligase complex from Pyrococcus horikoshii OT3. United Kingdom: N. p., 2007. Web. doi:10.1107/S1744309107011967.
Bagautdinov, Bagautdin, Matsuura, Yoshinori, Bagautdinova, Svetlana, & Kunishima, Naoki. Crystallization and preliminary X-ray crystallographic studies of the biotin carboxyl carrier protein and biotin protein ligase complex from Pyrococcus horikoshii OT3. United Kingdom. https://doi.org/10.1107/S1744309107011967
Bagautdinov, Bagautdin, Matsuura, Yoshinori, Bagautdinova, Svetlana, and Kunishima, Naoki. Sun . "Crystallization and preliminary X-ray crystallographic studies of the biotin carboxyl carrier protein and biotin protein ligase complex from Pyrococcus horikoshii OT3". United Kingdom. https://doi.org/10.1107/S1744309107011967.
@article{osti_22360298,
title = {Crystallization and preliminary X-ray crystallographic studies of the biotin carboxyl carrier protein and biotin protein ligase complex from Pyrococcus horikoshii OT3},
author = {Bagautdinov, Bagautdin and Matsuura, Yoshinori and Bagautdinova, Svetlana and Kunishima, Naoki},
abstractNote = {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%.},
doi = {10.1107/S1744309107011967},
url = {https://www.osti.gov/biblio/22360298}, journal = {Acta Crystallographica. Section F},
issn = {1744-3091},
number = Pt 4,
volume = 63,
place = {United Kingdom},
year = {2007},
month = {4}
}