To understand the role of His and Glu in the catalytic activity of Bacillus licheniformis a-amylase (BLA), His235 was replaced with Glu. The mutant enzyme, H235E, was characterized in terms of its mode of action using labeled and unlabeled maltooctaose (Glc8). H235E predominantly produced maltotridecaose (Glc13) from Glc8, exhibiting high substrate transglycosylation activity, with K-m = 0.38 mM and k(cat)/K-m = 20.58 mM(-1) s(-1) for hydrolysis, and K-m2 = 18.38 mM and k(cat2)/K-m2 = 2.57 mM(-1) s(-1) for transglycosylation, while the wild-type BLA exhibited high hydrolysis activity exclusively. Glu235-located on a wide open groove near subsite +1-is likely involved in transglycosylation via formation of an alpha-1,4-glycosidic linkage and may recognize and stabilize the non-reducing end glucose of the acceptor molecule. (C) 2014 Elsevier Inc. All rights reserved.