To provide insight into the potential role of a loop in domain B of several bacterial alpha-amylases, molecular and structural investigation of Bacillus stearothermophilus a-amylase (Amy US] 00) was used as a model. Combination deletion mutants of G(213), I-214 and G(215), 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 G(213)-I-214 or I-214-G(215) 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 degrees 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. (C) 2009 Elsevier Inc. All rights reserved.