Expression of a celE Gene from Clostridium thermocellum in Bacillus

Rustam Aminov, N. P. Golovchenko, K. Ohmiya

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Abstract

The level of expression of the Clostridium thermocellum celE gene in the asporogenous Bacillus subtilis strain 1A718 did not exceed the endogenous background level. However, when transformed into sporogenous strains, celE-containing constructs allowed the cells to express a high level of thermostable carboxy-methylcellulase (CMCase) activity which was detected exclusively in the culture supernatant. The sporulation efficiency was impaired in the celE-carrying strains. Most of the thermostable CMCase activity in the recombinant strains was attributed to the stationary phase of growth, and production of the enzyme could be further enhanced by increasing the cultivation temperature from 37 degrees C to 42 degrees C. Even when expressed in an extracellular proteases deficient mutant, the protein product was cleaved in the P-T-rich Linker sequence (2 sites) and at a site downstream of the putative signal peptidase recognition site. As a consequence, the enzymatically active protein could be isolated only in a truncated form. Plasmid pHE9102, the celE-containing construct, undergoes significant structural rearrangements in Bacillus stearothermophilus strains, preventing any detectable expression.
Original languageEnglish
JournalJournal of fermentation and bioengineering (Print)
Volume79
Issue number6
Pages (from-to)530-537
Number of pages8
ISSN0922-338X
DOIs
Publication statusPublished - 1995
Externally publishedYes

Keywords

  • FOOD
  • BIOTECHNOLOGY
  • LACTOBACILLUS-PLANTARUM
  • CRYSTALLINE CELLULOSE
  • ENDOGLUCANASE GENES
  • CATALYTIC ACTIVITY
  • ZYMOMONAS-MOBILIS
  • ESCHERICHIA-COLI
  • PLASMID PUB110
  • SUBTILIS
  • SECRETION
  • SEQUENCE
  • CLOSTRIDIUM THERMOCELLUM
  • BACILLUS
  • CELE
  • EXPRESSION
  • Clostridium thermocellum
  • Bacillus
  • celE
  • expression
  • biotechnology industry
  • MOLECULAR CLONING
  • PROTEIN PURIFICATION
  • Eubacteria Bacteria Microorganisms (Bacteria, Eubacteria, Microorganisms) - Endospore-forming Gram-Positives [07810] endospore-forming gram-positive rods and cocci Bacillus subtilis Clostridium perfringens
  • 10052, Biochemistry methods - Nucleic acids, purines and pyrimidines
  • 10062, Biochemistry studies - Nucleic acids, purines and pyrimidines
  • 10064, Biochemistry studies - Proteins, peptides and amino acids
  • 10504, Biophysics - Methods and techniques
  • 10506, Biophysics - Molecular properties and macromolecules
  • 13012, Metabolism - Proteins, peptides and amino acids
  • 31000, Physiology and biochemistry of bacteria
  • 31500, Genetics of bacteria and viruses
  • Biochemistry and Molecular Biophysics
  • Genetics
  • Metabolism
  • Methods and Techniques
  • Physiology
  • BACTERIA
  • CELLULASES
  • CLOSTRIDIUM
  • ENZYMES
  • GENE EXPRESSION
  • GENES
  • GENETICS
  • GLYCOSIDASES
  • Biotechnology

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