Cloning, expression, and carbon catabolite repression of the bamM gene encoding beta-amylase of Bacillus megaterium DSM319

Lee JS, Wittchen KD, Stahl C, Strey J, Meinhardt F

Research article (journal) | Peer reviewed

Abstract

The bamM gene from Bacillus megaterium DSM319 encoding an extracellular beta -amylase was isolated and completely sequenced. Chromosomal inactivation by deletion mutagenesis resulted in total loss of amylolytic activity, indicative of a single starch-degrading enzyme. Functional characterization of the expressed protein revealed a maltogenic enzyme exhibiting optimal activities at pH 7.5 and 50 degreesC. Amylase expression is subject to catabolite repression by glucose. A putative cis-acting catabolite-responsive element (CRE) was identified; it is located within the bamM coding region, matching the position of the predicted signal peptide processing site. Base substitutions introduced by site-directed mutagenesis within the bamM-CRE - retaining unchanged the amino acid sequence - provoked a remarkable relief from carbon catabolite repression (CCR), thereby proving functionality of the CRE for CCR.