Molecular cloning and characterization of an endoxylanase gene of Bacillus sp. in Escherichia coli.
A gene encoding an endoxylanase of Bacillus sp. was cloned and expressed in Escherichia coli. The entire nucleotide sequence of a 1,620 bp SmaI fragment containing the endoxylanase gene was determined. The endoxylanase gene was 639 bp long and encoded 213 amino acids which showed up to 96% amino acid homology with other endoxylanases. The endoxylanase produced by E. coli harboring pKJX4 was purified by ion-exchange chromatography (DE-52 and CM-52) and its N-terminal sequence was determined to be Ala-Gly-Thr-Asp-Tyr-Trp-Gln-Asn-Trp-Thr-Asp-Gly-Gly-Gly-Thr. The endoxylanase expressed in E. coli was identical to that of the original Bacillus sp. whose molecular weight was approximately 20,400. Most of the produced endoxylanase was localized in the periplasmic space of E. coli. When the endoxylanase was reacted with 2% oat spelts xylan (w/v) at 40 degrees C for 10 h, the major product was xylobiose which is known to be a selective growth stimulant to one of the healthy intestinal microflora, Bifidobacteria.
Jeong
K J
KJ
Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Kusong-dong Yusong-ku, Taejon, South Korea.
Lee
P C
PC
Park
I Y
IY
Kim
M S
MS
Kim
S C
SC
D000595:Amino Acid Sequence; D001407:Bacillus / Q000201:enzymology* / Q000235:genetics*; D001483:Base Sequence; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D043364:Endo-1,4-beta Xylanases; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics*; D005798:Genes, Bacterial / Q000235:genetics; D007700:Kinetics; D008969:Molecular Sequence Data; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D014990:Xylans / Q000378:metabolism; D014995:Xylosidases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*
D014990:Xylans; D014995:Xylosidases; D043364:Endo-1,4-beta Xylanases