4FNP

Crystal structure of GH36 alpha-galactosidase AgaA A355E from Geobacillus stearothermophilus

4FNP の概要

• エントリーDOI: 10.2210/pdb4fnp/pdb
• 関連するPDBエントリー: 4FNQ 4FNR 4FNS 4FNT 4FNU
• 分子名称: Alpha-galactosidase AgaA, SULFATE ION (3 entities in total)
• 機能のキーワード: glycoside hydrolase, hydrolase
• 由来する生物種: Geobacillus stearothermophilus
• タンパク質・核酸の鎖数: 4
• 化学式量合計: 334352.19

構造登録者

Merceron, R.,Foucault, M.,Haser, R.,Mattes, R.,Watzlawick, H.,Gouet, P. (登録日: 2012-06-20, 公開日: 2012-10-03, 最終更新日: 2024-02-28)

主引用文献

Merceron, R.,Foucault, M.,Haser, R.,Mattes, R.,Watzlawick, H.,Gouet, P.
The molecular mechanism of the thermostable alpha-galactosidases AgaA and AgaB explained by X-ray crystallography and mutational studies
J.Biol.Chem., 287:39642-39652, 2012

PubMed Abstract: The α-galactosidase AgaA from the thermophilic microorganism Geobacillus stearothermophilus has great industrial potential because it is fully active at 338 K against raffinose and can increase the yield of manufactured sucrose. AgaB has lower affinity for its natural substrates but is a powerful tool for the enzymatic synthesis of disaccharides by transglycosylation. These two enzymes have 97% identity and belong to the glycoside hydrolase (GH) family GH36 for which few structures are available. To understand the structural basis underlying the differences between these two enzymes, we determined the crystal structures of AgaA and AgaB by molecular replacement at 3.2- and 1.8 Å-resolution, respectively. We also solved a 2.8-Å structure of the AgaA(A355E) mutant, which has enzymatic properties similar to those of AgaB. We observe that residue 355 is located 20 Å away from the active site and that the A355E substitution causes structural rearrangements resulting in a significant displacement of the invariant Trp(336) at catalytic subsite -1. Hence, the active cleft of AgaA is narrowed in comparison with AgaB, and AgaA is more efficient than AgaB against its natural substrates. The structure of AgaA(A355E) complexed with 1-deoxygalactonojirimycin reveals an induced fit movement; there is a rupture of the electrostatic interaction between Glu(355) and Asn(335) and a return of Trp(336) to an optimal position for ligand stacking. The structures of two catalytic mutants of AgaA(A355E) complexed with raffinose and stachyose show that the binding interactions are stronger at subsite -1 to enable the binding of various α-galactosides.

PubMed: 23012371
DOI: 10.1074/jbc.M112.394114

実験手法

X-RAY DIFFRACTION (2.803 Å)