7KCT

Crystal Structure of the Hydrogenobacter thermophilus 2-Oxoglutarate Carboxylase (OGC) Biotin Carboxylase (BC) Domain Dimer in Complex with Adenosine 5'-Diphosphate Magnesium Salt (MgADP), Adenosine 5'-Diphosphate (ADP, and Bicarbonate Anion (Hydrogen Carbonate/HCO3-)

7KCT の概要

• エントリーDOI: 10.2210/pdb7kct/pdb
• 関連するPDBエントリー: 7KBL 7KC7
• 分子名称: 2-oxoglutarate carboxylase small subunit, ADENOSINE-5'-DIPHOSPHATE, MAGNESIUM ION, ... (5 entities in total)
• 機能のキーワード: biotin carboxylase, biotin-dependent carboxylase, pyruvate carboxylase, atp-grasp, aquificales, rtca, dimer interface, bicarbonate, sequence determining positions, structural waters, thermophile, carbon fixation, thermophilic protein, dimer, wet interface, ligase
• 由来する生物種: Hydrogenobacter thermophilus
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 111173.39

構造登録者

Buhrman, G.K.,Rose, R.B.,Enriquez, P.,Truong, V. (登録日: 2020-10-07, 公開日: 2021-01-27, 最終更新日: 2023-10-18)

主引用文献

Buhrman, G.,Enriquez, P.,Dillard, L.,Baer, H.,Truong, V.,Grunden, A.M.,Rose, R.B.
Structure, Function, and Thermal Adaptation of the Biotin Carboxylase Domain Dimer from Hydrogenobacter thermophilus 2-Oxoglutarate Carboxylase.
Biochemistry, 60:324-345, 2021

PubMed Abstract: 2-Oxoglutarate carboxylase (OGC), a unique member of the biotin-dependent carboxylase family from the order Aquificales, captures dissolved CO via the reductive tricarboxylic acid (rTCA) cycle. Structure and function studies of OGC may facilitate adaptation of the rTCA cycle to increase the level of carbon fixation for biofuel production. Here we compare the biotin carboxylase (BC) domain of OGC with the well-studied mesophilic homologues to identify features that may contribute to thermal stability and activity. We report three OGC BC X-ray structures, each bound to bicarbonate, ADP, or ADP-Mg, and propose that substrate binding at high temperatures is facilitated by interactions that stabilize the flexible subdomain B in a partially closed conformation. Kinetic measurements with varying ATP and biotin concentrations distinguish two temperature-dependent steps, consistent with biotin's rate-limiting role in organizing the active site. Transition state thermodynamic values derived from the Eyring equation indicate a larger positive Δ and a less negative Δ compared to those of a previously reported mesophilic homologue. These thermodynamic values are explained by partially rate limiting product release. Phylogenetic analysis of BC domains suggests that OGC diverged prior to Aquificales evolution. The phylogenetic tree identifies mis-annotations of the Aquificales BC sequences, including the pyruvate carboxylase structure. Notably, our structural data reveal that the OGC BC dimer comprises a "wet" dimerization interface that is dominated by hydrophilic interactions and structural water molecules common to all BC domains and likely facilitates the conformational changes associated with the catalytic cycle. Mutations in the dimerization domain demonstrate that dimerization contributes to thermal stability.

PubMed: 33464881
DOI: 10.1021/acs.biochem.0c00815

実験手法

X-RAY DIFFRACTION (2.02 Å)