Production of D-tagatose 3-epimerase of Pseudomonas cichorii ST-24 using recombinant Escherichia coli

被引:31
作者
Ishida, Y
Kamiya, T
Izumori, K [1 ]
机构
[1] Kagawa Univ, Fac Agr, Dept Bioresource Sci, Miki, Kagawa 76107, Japan
[2] Shikoku Res Inst Inc, Dept Biotechnol, Takamatsu, Kagawa 76101, Japan
来源
JOURNAL OF FERMENTATION AND BIOENGINEERING | 1997年 / 84卷 / 04期
关键词
Pseudomonas cichorii; D-tagatose; 3-epimerase; expression; EXPRESSION; GALACTITOL;
D O I
10.1016/S0922-338X(97)89257-4
中图分类号
Q81 [生物工程学(生物技术)]; Q93 [微生物学];
学科分类号
071005 ; 0836 ; 090102 ; 100705 ;
摘要
The D-tagatose 3-epimerase (D-TE) gene of Pseudomonas cichorii ST-24 was expressed in Escherichia coli under the control of the trc promoter. The D-TE production level was highest in E. coli JM105 as a host strain and in NZC medium as a culture medium. Production of D-TE by E. coli JM105 was about 100-fold higher than that of D-TE by P. cichorii ST-24, and the enzyme constituted similar to 5% of the total protein. D-TE from E. coli JM105 was purified by polyethylene glycol precipitation, DEAE-Toyopearl 650M column chromatography, and Sephadex G-150 column chromatography. The overall purification procedure resulted in 16.7-fold purification with 18.2% recovery. The molecular weight of the purified D-TE was estimated to be 33.0 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), and agreed with that of the D-TE from P. cichorii ST-24, D-TE produced by E. coli JM105 had generally the same enzymological properties, i.e., the optimum pH and temperature, pH and thermal stabilities, and K-m value, as that from Pseudomonas sp. ST-24 but the V-max value of D-TE from E. coli was 6 times higher than that from Pseudomonas sp. ST-24. The N-terminal amino acid sequence of the recombinant D-TE agreed with that of the D-TE from P. cichorii ST-24.
引用
收藏
页码:348 / 350
页数:3
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