Abstract
The C-terminal domain of Bacillus sp. strain TS-23 α-amylase (BLA) has been known to be involved in the raw starch-binding activity of the enzyme. Sequence comparison revealed that Thr-527, Trp-545, Trp-561, Lys-576, and Trp-588 in this domain are highly conserved in the aligned enzymes. To understand structure-function relationships in the starch-binding domain of BLA, site-directed mutagenesis was conducted to replace these residues with leucine or isoleucine. The overexpressed enzymes have been purified by nickel-chelate chromatography, and the molecular mass of the purified proteins was approximately 64.5 kDa. Starch-binding assay showed that the binding activities of the single-mutated enzymes were significantly reduced, while the combinational mutations did not lead to a complete loss of the activity.
Similar content being viewed by others
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Lo, F., Chiang, Y., Chi, C. et al. Site-Directed Mutagenesis of the Conserved Threonine, Tryptophan, and Lysine Residues in the Starch-Binding Domain of Bacillus sp. Strain TS-23 α-Amylase. Curr Microbiol 48, 280–284 (2004). https://doi.org/10.1007/s00284-003-4198-y
Issue Date:
DOI: https://doi.org/10.1007/s00284-003-4198-y