Cloning and Expression in<i>Escherichia coli</i>of Thermophilic<i>Clostridium</i>sp. F1 Genes Related to Cellulose Hydrolysis

Sakka, Kazuo; Furuse, Shinji; Shimada, Kyo

doi:10.1080/00021369.1989.10869431

Cited by 4 publications

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“…One approach for analysing the components of the complex consists of cloning and expressing the genes coding for the various enzymes in a suitable host. The cloning of 20 or so individual genes encoding CM-cellulases, xylanases and /8glucosidases from C. thermocellum into Escherichia coli has facilitated isolation of genes and production of individual cellulase components (Schwarz et al, 1985;Hazelwood et al, 1988;Sakka et al, 1989). A number of the recombinant cellulases, e.g.…”

Section: Introductionmentioning

confidence: 99%

Purification and characterization of endoglucanase Ss from Clostridium thermocellum

Fauth

Romaniec

Kobayashi

et al. 1991

Biochemical Journal

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The extracellular cellulolytic enzymes of the thermophilic anaerobe Clostridium thermocellum occur as a protein complex or aggregate known as the cellulosome. By using a combination of ion-exchange, adsorption and hydrophobic-interaction chromatography, it was possible to isolate from extracellular broth a specific endoglucanase of interest without the use of denaturants. The endoglucanase was identified as the cellulosomal subunit Ss by the use of specific antibodies. The enzyme has an Mr of 83,000, an isoelectric point of 3.55, optimum pH of 6.6 and optimum temperature of 70 degrees C. It hydrolyses CM-cellulose and, at a higher rate, the cellodextrins, cellotetraose and cellopentaose, but does not hydrolyse a crystalline cellulose such as Avicel. Cellobiose and cellotriose are also immune to attack. It differs from endoglucanases previously isolated by others and a 76,000-Mr endoglucanase recently isolated in this laboratory.

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Section: Introductionmentioning

confidence: 99%

Purification and characterization of endoglucanase Ss from Clostridium thermocellum

Fauth

Romaniec

Kobayashi

et al. 1991

Biochemical Journal

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“…For example, xylanases and their genes have been reported of thermophilic anaerobic bacteria such as C. stercorarium, 4 -6) Clostridium thermocellum,7 -10) and Caldocellum saccharolyticum. We cloned several xylanase genes in E. coli from C. stercorarium F-9 5 ) and C. thermocellum F _1, 9) which had been isolated and identified in this laboratory. We have sequenced one of them, the xynA gene, from C. stercorarium F-9 (this gene and its translated product are referred to as xynA F _ 9 and XynA F _ 9 , respectively, in this paper) and found that xynA F _ 9 encoded a protein composed of 511 amino acids with a molecular weight of 56,519 including a signal peptide.…”

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confidence: 99%

Purification and Characterization of Xylanase A fromClostridium stercorariumF-9 and a RecombinantEscherichia coli

Sakka

Kojima

Kondo

et al. 1994

Bioscience, Biotechnology, and Biochemistry

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Xylanase A encoded by the Clostridium stercorarium F-9 xynA gene was purified to homogeneity from a recombinant clone of Escherichia coli. The N-terminal amino acid sequence and molecular weight (54,000) estimated by SDS-PAGE of the purified enzyme were consistent with those deduced from the nucleotide sequence [Biosci. Biotech. Biochem., 57, 273-277 (1993)]. A xylanase was also purified to homogeneity from a culture supernatant of C. stercorarium F-9. Its N-terminal amino acid sequence, molecular weight, and enzymatic properties were quite in agreement with those of the recombinant enzyme, indicating that the xynA gene was predominantly expressed as a xylanase gene in C. stercorarium F-9. The purified enzyme hydrolyzed xylotriose to yield xylobiose and xylose while it was less active toward xylobiose. It was optimally active at 75 degrees C and pH 7.0 Km and Vmax were estimated to be 1.9 mg/ml and 2.8 mumol of xylose equivalent/min/micrograms for oat spelt xylan, respectively.

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“…strain FI, which was isolated in this laboratory, and found that one of them (termed ce1C307) was homologous with celC of C. thermocellum. 6 ) It seemed to be interesting to sequence the nucleotides of celC307 because clarifying the difference in the nucleotide sequences between celC and celC307 might be helpful to study the gene structure in respect to conserved or essential parts. In this paper, we describe the nucleotide sequence of celC307 including the promoter region and the expression of the gene in E. coli.…”

Section: Thermophilic Cellulases and Their Genes Inmentioning

confidence: 99%

Nucleotide Sequence ofcelC307Encoding EndoglucanaseC307ofClostridiumsp. Strain F1

Sakka

Shimanuki

Shimada

1991

Agricultural and Biological Chemistry

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Cloning and Expression inEscherichia coliof ThermophilicClostridiumsp. F1 Genes Related to Cellulose Hydrolysis

Cited by 4 publications

References 0 publications

Purification and characterization of endoglucanase Ss from Clostridium thermocellum

Purification and characterization of endoglucanase Ss from Clostridium thermocellum

Purification and Characterization of Xylanase A fromClostridium stercorariumF-9 and a RecombinantEscherichia coli

Nucleotide Sequence ofcelC307Encoding EndoglucanaseC307ofClostridiumsp. Strain F1

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