FEBS Letters | |
Characterization of a peripheral‐type benzodiazepine‐binding site in the mitochondria of Chinese hamster ovary cells | |
Ferrara, P.1  Riond, J.1  Vita, N.1  Le Fur, G.1  | |
[1] Sanofi Elf Bio-Recherches, BP 137, Labège 31328 Cédex, France | |
关键词: Benzodiazepine; Receptor; Purification; HPLC; Photolabeling; | |
DOI : 10.1016/0014-5793(89)80229-7 | |
学科分类:生物化学/生物物理 | |
来源: John Wiley & Sons Ltd. | |
【 摘 要 】
The isoquinoline carboxamide derivative [3H]PK11195, a ligand for the peripheral-type benzodiazepine (BZD) receptor, binds to Chinese hamster ovary (CHO) cell mitochondria in a specific and saturable manner. Scatchard analysis showed the presence of a single-binding site with an apparent dissociation constant (K d) of 12.0 ± 1.0 nM and a maximal binding capacity of 23.0 ± 2.0 pmol/mg protein. The pharmacological characterization of this CHO BZD-binding site, based on the displacement of [3H]PK11195 by several drugs of known binding specificity, indicated that it is of the peripheral-type. The photoaffinity probe [3H]PK14105, a nitrophenyl derivative of [3H]PK11195, specifically labeled a 17 kDa CHO mitochondrial protein. This 17 kDa protein was purified from digitonin-solubilized mitochondria by gel-filtration chromatography and two reverse-phase HPLC steps. The purified material migrated as a single band on silver stained or autoradiographed SDS-polyacrylamide gels, and had an amino acid composition corresponding to a 17 kDa protein rich in Leu, Val, Ala, Gly, and Pro. Analysis of the amino-terminal sequence of the purified 17 kDa protein revealed a blocked amino-terminus.
【 授权许可】
Unknown
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