Photolabeling of the phosphate binding site of chloroplast coupling factor 1 With [32P]azidonitrophenyl phosphate

FEBS Letters

Garin, Jérôme³  Girault, Guy²  Vignais, Pierre V.¹  Michel, Laurent¹ 

[1] Laboratoire de Biochimie (URA 1130 CNRS), Département de Biologie Moléculaire et Structurale, Centre d'Etudes Nucléaires, 85X, 38041 Grenoble Cedex, France;Section de Bioénergétique, Département de Biologie Cellulaire et Moléculaire, CEN Saclay, 91191 Gif-sur-Yvette Cedex, France;Laboratoire de Chimie de Protéines, Département de Biologie Moléculaire et Structurale, Centre d'Etudes Nucléaires, 85X, 38041 Grenoble Cedex, France

关键词: Chloroplast F1 ATPase;    Phosphate binding site;    Photoaffinity;    4-Azido-2-nitrophenylphosphate;    ANPP;    4-azido-2-nitrophenyl phospate;    MES;    2-(N-morpholino)ethane sulfonic acids;    TFA;    trifluoroacetic acid;    CF1;    catalytic factor (soluble) of the ATP systhase complex from chloroplasts;    TDAB;    tetradecyltrimethylammonium bromide;    EDC;    1-ethyl-3-(3-dimethyl-aminopropyl) carbodiimide;   

DOI  :  10.1016/0014-5793(92)81191-N

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Chloroplast F₁-AT Pase (CF₁) was photolabeled by a radiolabeled photoactivatable derivative of P_i, 4-azido-2-nitrophenyl [³²P]phosphate (ANPP). The radioactivity was localized in the β subunit of CF₁. Upon cleavage of the β subunit by cyanogen bromide, the predominantly labeled peptide was recovered, which was subsequently subjected to tryptic digestion. A tryptic peptide (spanning Ile³¹²-Arg³⁵⁴), was found to contain nearly all the covalently bound radioactivity. By Edman degradation, the labeled amino acid residues were identified as Tyr³²⁸, Val³²⁹ and Pro³³⁰. The labeled β-Tyr³²⁸ of CF₁ is the equivalent of β-Tyr³¹¹ of F₁ from beef heart mitochondria, which was previously found to be photolabeled by ANPP [J. Garin et al. (1989) Biochemistry 28, 1442–1448].

【 授权许可】

Unknown