Characterization of the catalytic domains of <Emphasis Type="Italic">Trichoderma reesei</Emphasis> endoglucanase I,II, and III,expressed in <Emphasis Type="Italic">Escherichia coli</Emphasis> |
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Authors: | Hikaru Nakazawa Katsunori Okada Ryota Kobayashi Tetsuya Kubota Tomoko Onodera Nobuhiro Ochiai Naoki Omata Wataru Ogasawara Hirofumi Okada Yasushi Morikawa |
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Institution: | (1) Department of Bioengineering, Nagaoka University of Technology, 1603-1 Kamitomioka, Nagaoka 940-21, Japan |
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Abstract: | The genes encoding the catalytic domains (CD) of the three endoglucanases (EG I; Cel7B, EG II; Cel5A, and EG III; Cel12A)
from Trichoderma reesei QM9414 were expressed in Escherichia coli strains Rosetta-gami B (DE3) pLacI or Origami B (DE3) pLacI and were found to produce functional intracellular proteins.
Protein production by the three endoglucanase transformants was evaluated as a function of growth temperature. Maximal productivity
of EG I-CD at 15°C, EG II-CD at 20°C and EG III at 37°C resulted in yields of 6.9, 72, and 50 mg/l, respectively. The endoglucanases
were purified using a simple purification method based on removing E. coli proteins by isoelectric point precipitation. Specific activity toward carboxymethyl cellulose was found to be 65, 49, and
15 U/mg for EG I-CD, EG II-CD, and EG III, respectively. EG II-CD was able to cleave 1,3–1,4-β-d-glucan and soluble cellulose derivatives. EG III was found to be active against cellulose, 1,3–1,4-β-d-glucan and xyloglucan, while EG I-CD was active against cellulose, 1,3–1,4-β-d-glucan, xyloglucan, xylan, and mannan. |
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Keywords: | Keyword" target="_blank">Keyword Endoglucanase Trichoderma reesei Heterologous expression Low temperature Rosetta-gami B (DE3) pLacI |
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