Cloning, expression, and carbon catabolite repression of the bamM gene encoding beta-amylase of Bacillus megaterium DSM319.
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 degrees C. 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.
Lee
J S
JS
Institut für Mikrobiologie, Westfälische Wilhelms-Universität, Münster, Germany.
Wittchen
K D
KD
Stahl
C
C
Strey
J
J
Meinhardt
F
F
D000595:Amino Acid Sequence; D001410:Bacillus megaterium / Q000201:enzymology* / Q000235:genetics; D001483:Base Sequence; D002244:Carbon / Q000378:metabolism*; D003001:Cloning, Molecular; D004794:Enzyme Repression; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D020218:Response Elements; D001614:beta-Amylase / Q000235:genetics*
D002244:Carbon; D001614:beta-Amylase