The importance of an extra loop in the B-domain of an α-amylase from B. stearothermophilus

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• 作者：Bassem Khemakhem ; Mamdouh Ben Ali ; Nushin Aghajari ; Michel Juy ; Richard Haser ; Samir Bejar
• 关键词：α ; -Amylase ; Mutagenesis ; Bacillus stearothermophilus ; Deletion ; Thermostability ; Loop ; B-domain
• 刊名：Biochemical and Biophysical Research Communications
• 出版年：2009
• 出版时间：17 July 2009
• 年：2009
• 卷：385
• 期：1
• 页码：78-83
• 全文大小：483 K

文摘

To provide insight into the potential role of a loop in domain B of several bacterial α-amylases, molecular and structural investigation of Bacillus stearothermophilus α-amylase (Amy US100) was used as a model. Combination deletion mutants of G₂₁₃, I₂₁₄ and G₂₁₅, described as a loop-forming on the surface bacterial amylases, were subjected to biochemical and structural investigation. Thermoactivity, thermostability as well calcium requirement were studied for each mutant.

Thus, deletion of one residue differently affects only the thermostability. Shortening the loop by deletion of G₂₁₃–I₂₁₄ or I₂₁₄–G₂₁₅ improved the thermostability and reduces calcium requirement. However, the deletion of three residues has a negative effect on thermostability and reduces the optimal temperature by 17 °C.

The structural investigation showed that stabilizing deletions contribute to reinforce the architecture of domain B and the active site conformation. The deletion of three residues reduces the flexibility of this region and abolishes a denser hydrogen bond network.