Optimal production of a novel endo-acting beta-1,4-xylanase cloned from Saccharophagus degradans 2-40 into Escherichia coli BL21(DE3).
To date, gene xyn10C from Saccharophagus degradans 2-40 has only been identified to encode a potential xylanase. In the present study, xyn10C was cloned and overexpressed in Escherichia coli BL21(DE3). The protein produced by xyn10C, Xyn10C, was expressed in a soluble active form and found to be an endotype beta-1,4-xylanase that preferentially produces xylobiose from xylan. Recombinant cell fermentation revealed that induction of the gene at low temperatures fostered expression of the recombinant xylanase with high volumetric and specific activities. Additionally, low growth rates were favorable for producing soluble active xylanase via a reduction in the formation of inclusion bodies. Furthermore, the optimal concentration of isopropyl-beta-D-thiogalactopyranoside for induction was found to be 100 microm after two hours of precultivation at 37 degrees C. Finally, enzyme production conducted using a fermentor with a working volume of 1.5-l resulted in slightly higher specific activities of xylanase when compared with the generation of enzymes in flasks with a working volume of 100ml.
Ko
Ja Kyong
JK
School of Life Sciences and Biotechnology, Korea University, Seoul 136-713, Republic of Korea.
Jung
Min Woo
MW
Kim
Kyoung Heon
KH
Choi
In-Geol
IG
D034241:Alteromonadaceae / Q000187:drug effects / Q000201:enzymology*; D049109:Cell Proliferation / Q000187:drug effects; D002851:Chromatography, High Pressure Liquid; D002855:Chromatography, Thin Layer; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D043364:Endo-1,4-beta Xylanases / Q000096:biosynthesis* / Q000235:genetics*; D004926:Escherichia coli / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D005285:Fermentation / Q000187:drug effects; D006868:Hydrolysis / Q000187:drug effects; D007544:Isopropyl Thiogalactoside / Q000494:pharmacology; D011994:Recombinant Proteins / Q000096:biosynthesis; D013696:Temperature; D013997:Time Factors; D014990:Xylans / Q000378:metabolism
D011994:Recombinant Proteins; D014990:Xylans; D007544:Isopropyl Thiogalactoside; D043364:Endo-1,4-beta Xylanases