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

• Published in: Applied microbiology and biotechnology Vol. 56; no. 1/2; pp. 205 - 211
• Main Authors: Lee, J.S, Wittchen, K.D, Stahl, C, Strey, J, Meinhardt, F
• Format: Journal Article
• Language: English
• Published: Berlin Springer 01.07.2001; Springer Nature B.V
• Subjects: Amino Acid Sequence; Amino acids; Amylases; Bacillus megaterium; Bacillus megaterium - enzymology; Bacillus megaterium - genetics; Bacteriology; bamM gene; Base Sequence; beta-amylase; beta-Amylase - genetics; Biological and medical sciences; Biology of microorganisms of confirmed or potential industrial interest; Biotechnology; Carbon; Carbon - metabolism; Catabolite repression; catabolite-responsive element; Chromosome deletion; Cloning; Cloning, Molecular; enzyme activity; Enzyme Repression; Fundamental and applied biological sciences. Psychology; genes; Genetics; glucose; Microbiology; Mission oriented research; Molecular Sequence Data; Mutagenesis, Site-Directed; Regulatory sequences; Response Elements; signal peptide; Signal processing; Site-directed mutagenesis; β-Amylase; Nucleotide sequence; Enzyme; Bacillaceae; Gene expression; Carbon; β-Amylase; Bacillales; Enzymatic activity; Gene; Glycosidases; Bacteria; Hydrolases; Regulatory sequence; Mutation; O-Glycosidases; Bacillus megaterium; Catabolic repression
• ISSN: 0175-7598; 1432-0614
• DOI: 10.1007/s002530100645

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.