Nucleotide sequence and characteristics of endoglucanase gene engB from Clostridium cellulovorans.
An endoglucanase gene, engB, from Clostridium cellulovorans, previously cloned into pUC19, has been further characterized and its product investigated. The enzyme, EngB, encoded by the gene was secreted into the periplasmic space of Escherichia coli. The enzyme was active against carboxymethylcellulose, xylan and lichenan but not Avicel (crystalline cellulose). The sequenced gene showed an open reading frame of 1323 base pairs and coded for a protein with a molecular mass of 48.6 kDa. The mRNA contained a typical Gram-positive ribosome-binding site sequence GGAGG and a sequence coding for a putative signal peptide. There is high amino acid and base sequence homology between the N-terminal regions of EngB and another C. cellulovorans endoglucanase, EngD, but they differ significantly in their C-termini. Deletion analyses revealed that up to 32 amino acids of the N-terminus and 52 amino acids of the C-terminus were not required for catalytic activity. The conserved reiterated domains at the C-terminus of EngB were similar to those from endoglucanases from other cellulytic bacteria. According to our deletion analyses, this region is not needed for catalytic activity.
Foong
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Department of Biochemistry and Biophysics, University of California, Davis 95616.
Hamamoto
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Shoseyov
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Doi
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D000595:Amino Acid Sequence; D001483:Base Sequence; D002266:Carboxymethylcellulose Sodium / Q000378:metabolism; D002480:Cellulase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D003013:Clostridium / Q000201:enzymology / Q000235:genetics*; D004252:DNA Mutational Analysis; D004926:Escherichia coli / Q000235:genetics; D005936:Glucans / Q000378:metabolism; D008969:Molecular Sequence Data; D016366:Open Reading Frames / Q000235:genetics*; D012689:Sequence Homology, Nucleic Acid; D014990:Xylans / Q000378:metabolism
D005936:Glucans; D014990:Xylans; D002480:Cellulase; D002266:Carboxymethylcellulose Sodium; C015968:lichenin