An FKBP12 binding assay based upon biotinylated FKBP12

被引：7

作者：

Carreras, CW

Fu, H

Santi, DV

机构：

[1] Kosan Biosci Inc, Dept Pharmacol Sci, Hayward, CA 94545 USA

[2] Kosan Biosci Inc, Dept Chem, Hayward, CA 94545 USA

来源：

ANALYTICAL BIOCHEMISTRY | 2001年 / 298卷 / 01期

关键词：

FKBP; FK506; immunosuppression; biotin; birA;

D O I：

10.1006/abio.2001.5343

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

A binding assay was developed for measuring the affinity of FRBP12 ligands. A biotinylation signal sequence was fused to the 5' end of the human FKBP12 gene, and the fusion protein was expressed in Escherichia coli with biotin ligase. The fusion protein was immobilized in avidin-coated multiwell plates, and varying concentrations of test ligands were allowed to compete with [H-3]FK506 for FKBP12 sites on the plate. The assay provided K-d values for FK520, 32-hydroxy-ethyl indolyl FK520, and 18-ene, 20-oxa FK520 that are in agreement with previously reported values. The assay provides a convenient and rapid method for the assessment of FKBP12 binding by small molecules. (C) 2001 Academic Press.

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收藏

页码：57 / 61

页数：5

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