9F17

Crystal structure of N term His-tag Adenylosuccinate synthetase from Helicobacter pylori

9F17 の概要

• エントリーDOI: 10.2210/pdb9f17/pdb
• 関連するPDBエントリー: 6ZXQ
• 分子名称: Adenylosuccinate synthetase, CALCIUM ION, GUANOSINE-5'-DIPHOSPHATE, ... (7 entities in total)
• 機能のキーワード: adenylosuccinate synthetase, h. pylori, ligase
• 由来する生物種: Helicobacter pylori 26695
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 95041.97

構造登録者

Stefanic, Z. (登録日: 2024-04-18, 公開日: 2024-09-04)

主引用文献

Miskovic, M.Z.,Wojtys, M.,Winiewska-Szajewska, M.,Wielgus-Kutrowska, B.,Matkovic, M.,Domazet Jurasin, D.,Stefanic, Z.,Bzowska, A.,Lescic Asler, I.
Location Is Everything: Influence of His-Tag Fusion Site on Properties of Adenylosuccinate Synthetase from Helicobacter pylori.
Int J Mol Sci, 25:-, 2024

PubMed Abstract: The requirement for fast and dependable protein purification methods is constant, either for functional studies of natural proteins or for the production of biotechnological protein products. The original procedure has to be formulated for each individual protein, and this demanding task was significantly simplified by the introduction of affinity tags. adenylosuccinate synthetase (AdSS) is present in solution in a dynamic equilibrium of monomers and biologically active homodimers. The addition of the His-tag on the C-terminus (C-His-AdSS) was proven to have a negligible effect on the characteristics of this enzyme. This paper shows that the same enzyme with the His-tag fused on its N-terminus (N-His-AdSS) has a high tendency to precipitate. Circular dichroism and X-ray diffraction studies do not detect any structural change that could explain this propensity. However, the dynamic light scattering, differential scanning fluorimetry, and analytical ultracentrifugation measurements indicate that the monomer of this construct is prone to aggregation, which shifts the equilibrium towards the insoluble precipitant. In agreement, enzyme kinetics measurements showed reduced enzyme activity, but preserved affinity for the substrates, in comparison with the wild-type and C-His-AdSS. The presented results reinforce the notion that testing the influence of the tag on protein properties should not be overlooked.

PubMed: 39062851
DOI: 10.3390/ijms25147613

実験手法

X-RAY DIFFRACTION (1.7 Å)