Fina Khaerunnisa Frima, Muhammad Akbar Thufail, Indri Novia Madhani, Zahrotun Nafisah, Sofi Siti Shofiyah, Ayra Ulpiyana, Fernita Puspasari, Reza Aditama, Ihsanawati Ihsanawati, Dessy Natalia
The α-amylase BmaN1 from Bacillus megaterium NL3 is a member of GH13_45 subfamily that has a conserved C-terminal region of approximately 30 residues. This region features a motif of five aromatic amino acids predicted to play a role in starch binding. This study aimed to unravel the role of the C-terminal region in starch hydrolysis. The full-length and C-terminally truncated forms of BmaN1 (BmaN1∆C) were expressed in Escherichia coli ArcticExpress (DE3), resulting in proteins with molecular weights of 56 kDa and 49 kDa, respectively. They exhibited comparable enzymatic activity in the hydrolysis of soluble starch, displaying versatility across a wide range of pH values, temperatures, and NaCl concentrations. BmaN1 and BmaN1∆C activities were inhibited by acarbose and were reduced by SDS and EDTA. In terms of binding and degrading the starch granules, BmaN1∆C showed lower affinity and activity in comparison to BmaN1. Our study indicates that the C-terminal region of BmaN1 significantly enhances its binding affinity and degrading the raw starches. Copyright © 2024 Frima et al.
Biochemistry and Biomolecular Engineering Research Division, Faculty of Mathematics and Natural Sciences, Institut Teknologi Bandung, Bandung, Indonesia; Department of Chemistry, Faculty of Science, Institut Teknologi Sumatera, Lampung Selatan, Indonesia; Department of Chemistry, Faculty of Science and Marine, Universitas Oso, Pontianak, Indonesia; Biosciences and Biotechnology Research Center, Institut Teknologi Bandung, Bandung, Indonesia