CHARACTERIZATION OF RECOMBINANT HUMAN NEURON-SPECIFIC ENOLASE AND ITS APPLICATION TO ENZYME-IMMUNOASSAY

被引：9

作者：

AOKI, T ^{[1
]}

KIMURA, M ^{[1
]}

KANETA, M ^{[1
]}

KAZAMA, H ^{[1
]}

MORIKAWA, J ^{[1
]}

WATABE, H ^{[1
]}

机构：

[1] EIKEN CHEM CO LTD,TOKYO,JAPAN

来源：

TUMOR BIOLOGY | 1993年 / 14卷 / 05期

关键词：

NEURON-SPECIFIC ENOLASE; GAMMA-ENOLASE; RECOMBINANT PROTEIN; PROTEIN PURIFICATION; TUMOR MARKER; ENZYME IMMUNOASSAY;

D O I：

10.1159/000217838

中图分类号：

R73 [肿瘤学];

学科分类号：

100214 ;

摘要：

Human gamma-enolase cDNA prepared by reverse transcriptase-polymerase chain reaction was cloned into the Escherichia coli expression vector pKK223-3. The resulting plasmid, pHTK503, expressed human gamma-enolase as a 46-kDa protein in SDS-PAGE, and in the cells as the active gammagamma form (designated as recombinant human NSE; R-NSE). R-NSE was purified from E. coli by several chromatographic elutions. Finally, 6.0 mg of R-NSE from 8.1 g cells was purified with a specific activity of 86 units/mg protein. The structural properties of R-NSE were compared with the NSE purified from human brain tissue (B-NSE). The biochemical and enzymatic characteristics were essentially the same, except for the isoelectric point (4.5 for B-NSE and 4.7 for R-NSE). In an NSE immunoassay system, R-NSE and standard NSE were almost equal in reactivity to the anti-NSE antibody. These results indicate that R-NSE can be used as standard assay material.

引用

页码：261 / 270

页数：10

共 32 条

[1]

AOKI T, 1992, CHEM PHARM BULL, V40, P1236

[2]

ARAKAWA T, 1985, J BIOL CHEM, V260, P4435

[3]

BIRNBOIM HC, 1979, NUCLEIC ACIDS RES, V7, P1513

[4] REGULATION OF RIBOSOMAL-RNA PROMOTERS WITH A SYNTHETIC LAC OPERATOR [J].