This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Tran, L Articles by Wong, S L Search for Related Content PubMed PubMed Citation Articles by Tran, L Articles by Wong, S L

 Previous Article  |  Next Article 

J Bacteriol. 1991 October; 173(20): 6364-6372

research-article

Cloning and expression of a novel protease gene encoding an extracellular neutral protease from Bacillus subtilis.

Department of Biological Sciences, University of Calgary, Alberta, Canada.

ABSTRACT

We have cloned from Bacillus subtilis a novel protease gene (nprB) encoding a neutral protease by using a shotgun cloning approach. The gene product was determined to have a molecular mass of 60 kDa. It has a typical signal peptide-like sequence at the N-terminal region. The expression of nprB can be stimulated by using a B. subtilis strain, WB30, carrying a sacU(h)h mutation. Expression of this protease gene results in production of a 37-kDa protease in the culture medium. The first five amino acid residues from the N terminus of the mature protease were determined to be Ala-Ala-Gly-Thr-Gly. This indicates that the protease is synthesized in a preproenzyme form. The purified protease has a pH optimum of around 6.6, and its activity can be inhibited by EDTA, 1,10-phenanthroline (a zinc-specific chelator), and dithiothreitol. It retained 65% of its activity after treatment at 65 degrees C for 20 min. Sequence comparison indicates that the mature form of this protease has 66% homology with the two thermostable neutral proteases from B. thermoproteolyticus and B. stearothermophilus. It also shares 65, 61, and 56% homology with the thermolabile neutral proteases from B. cereus, B. amyloliquefaciens, and B. subtilis, respectively. The zinc-binding site and the catalytic residues are all conserved among these proteases. Sequence homology extends into the "propeptide" region. The nprB gene was mapped between metC and glyB and was not required for growth or sporulation.

This article has been cited by other articles:

• Thomaides, H. B., Davison, E. J., Burston, L., Johnson, H., Brown, D. R., Hunt, A. C., Errington, J., Czaplewski, L. (2007). Essential Bacterial Functions Encoded by Gene Pairs. J. Bacteriol. 189: 591-602 [Abstract] [Full Text]  
• Park, C. H., Lee, S. J., Lee, S. G., Lee, W. S., Byun, S. M. (2004). Hetero- and Autoprocessing of the Extracellular Metalloprotease (Mpr) in Bacillus subtilis. J. Bacteriol. 186: 6457-6464 [Abstract] [Full Text]  
• Bindel Connelly, M., Young, G. M., Sloma, A. (2004). Extracellular Proteolytic Activity Plays a Central Role in Swarming Motility in Bacillus subtilis. J. Bacteriol. 186: 4159-4167 [Abstract] [Full Text]  
• Obst, M., Oppermann-Sanio, F. B., Luftmann, H., Steinbuchel, A. (2002). Isolation of Cyanophycin-degrading Bacteria, Cloning and Characterization of an Extracellular Cyanophycinase Gene (cphE) from Pseudomonas anguilliseptica Strain BI. THE cphE GENE FROM P. ANGUILLISEPTICA BI ENCODES A CYANOPHYCIN-HYDROLYZING ENZYME. J. Biol. Chem. 277: 25096-25105 [Abstract] [Full Text]  
• Wu, S.-C., Wong, S.-L. (2002). Engineering of a Bacillus subtilis Strain with Adjustable Levels of Intracellular Biotin for Secretory Production of Functional Streptavidin. Appl. Environ. Microbiol. 68: 1102-1108 [Abstract] [Full Text]  
• Tjalsma, H., Bolhuis, A., Jongbloed, J. D. H., Bron, S., van Dijl, J. M. (2000). Signal Peptide-Dependent Protein Transport in Bacillus subtilis: a Genome-Based Survey of the Secretome. Microbiol. Mol. Biol. Rev. 64: 515-547 [Abstract] [Full Text]  
• Rao, M. B., Tanksale, A. M., Ghatge, M. S., Deshpande, V. V. (1998). Molecular and Biotechnological Aspects of Microbial Proteases. Microbiol. Mol. Biol. Rev. 62: 597-635 [Abstract] [Full Text]  
• Wu, S.-C., Ye, R., Wu, X.-C., Ng, S.-C., Wong, S.-L. (1998). Enhanced Secretory Production of a Single-Chain Antibody Fragment from Bacillus subtilis by Coproduction of Molecular Chaperones. J. Bacteriol. 180: 2830-2835 [Abstract] [Full Text]  
• de Kreij, A., Venema, G., van den Burg, B. (2000). Substrate Specificity in the Highly Heterogeneous M4 Peptidase Family Is Determined by a Small Subset of Amino Acids. J. Biol. Chem. 275: 31115-31120 [Abstract] [Full Text]