3WSC

Crystal structure of alginate-binding protein Algp7

3WSC の概要

• エントリーDOI: 10.2210/pdb3wsc/pdb
• 分子名称: Alginate-binding protein (2 entities in total)
• 機能のキーワード: two up-and-down four-helical bundles, alginate binding, sphingomonas, alginate-binding protein
• 由来する生物種: Sphingomonas
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 63402.18

構造登録者

Temtrirath, K.,Murata, K.,Hashimoto, W. (登録日: 2014-03-07, 公開日: 2015-02-18, 最終更新日: 2024-03-20)

主引用文献

Temtrirath, K.,Murata, K.,Hashimoto, W.
Structural insights into alginate binding by bacterial cell-surface protein
Carbohydr.Res., 404C:39-45, 2014

PubMed Abstract: A gram-negative Sphingomonas sp. strain A1 inducibly forms a mouth-like pit on the cell surface in the presence of alginate and directly incorporates polymers into the cytoplasm via the pit and ABC transporter. Among the bacterial proteins involved in import of alginate, a cell-surface EfeO-like Algp7 shows an ability to bind alginate, suggesting its contribution to accumulate alginate in the pit. Here, we show identification of its positively charged cluster involved in alginate binding using X-ray crystallography, docking simulation, and site-directed mutagenesis. The tertiary structure of Algp7 was determined at a high resolution (1.99Å) by molecular replacement, although no alginates were included in the structure. Thus, an in silico model of Algp7/oligoalginate was constructed by docking simulation using atomic coordinates of Algp7 and alginate oligosaccharides, where some charged residues were found to be potential candidates for alginate binding. Site-directed mutagenesis was conducted and five purified mutants K68A, K69A, E194A, N221A, and K68A/K69A were subjected to a binding assay. UV absorption difference spectroscopy along with differential scanning fluorimetry analysis indicated that K68A/K69A exhibited a significant reduction in binding affinity with alginate than wild-type Algp7. Based on these data, Lys68/Lys69 residues of Algp7 probably play an important role in binding alginate.

PubMed: 25665777
DOI: 10.1016/j.carres.2014.11.008

実験手法

X-RAY DIFFRACTION (1.992 Å)