Characterization of the catalytic domains of Trichoderma reesei endoglucanase I, II, and III, expressed in Escherichia coli.
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 degrees C, EG II-CD at 20 degrees C and EG III at 37 degrees 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-beta-D-glucan and soluble cellulose derivatives. EG III was found to be active against cellulose, 1,3-1,4-beta-D-glucan and xyloglucan, while EG I-CD was active against cellulose, 1,3-1,4-beta-D-glucan, xyloglucan, xylan, and mannan.
Nakazawa
Hikaru
H
Department of Bioengineering, Nagaoka University of Technology, 1603-1 Kamitomioka, Nagaoka, 940-21, Japan.
Okada
Katsunori
K
Kobayashi
Ryota
R
Kubota
Tetsuya
T
Onodera
Tomoko
T
Ochiai
Nobuhiro
N
Omata
Naoki
N
Ogasawara
Wataru
W
Okada
Hirofumi
H
Morikawa
Yasushi
Y