Mutations in the amino‐terminal domain of the human poly(ADP‐ribose) polymerase that affect its catalytic activity but not its DNA binding capacity

FEBS Letters

Fribourg, Sébastien¹  Trucco, Carlotta¹  Ménissier-de Murcia, Josiane¹  de Murcia, Gilbert¹  Flatter, Eric¹ 

[1] Ecole Supérieure de Biotechnologie de Strasbourg, UPR A9003 du CNRS, Cancérogenèse et Mutagenèse Moléculaire et Structurale, Boulevard Sébastien Brant, F-67400 Illkirch-Graffenstaden, France

关键词: Random mutagenesis;    Colony screening;    Zinc finger;    DNA binding protein;    NAD metabolism;    PARP;    poly(ADP-ribose) polymerase (EC 2.4.2.30);    PCR;    polymerase chain reaction;    DBD;    DNA-binding domain;    FI;    zinc finger I;    FII;    zinc finger II;    DTT;    dithiothreitol;   

DOI  :  10.1016/S0014-5793(96)01347-6

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Poly-ADP ribosylation of nuclear proteins is activated when poly(ADP-ribose) polymerase (PARP), a nuclear zinc-finger enzyme, binds to single-strand DNA breaks. To understand how the signal emerging from its DNA-binding domain (DBD) bound to such breaks is transduced to its catalytic domain, the structure-function relationship of the DBD was investigated. We have used mutagenesis by the polymerase chain reaction (PCR) to generate a random library of PARP mutants. In this work, we describe the identification of catalytically inactive mutants bearing single point mutations, located outside the two zinc fingers in the DBD, that have conserved their full capacity to bind DNA. The results obtained demonstrate that the DNA-dependent activation of PARP requires not only a capacity to bind DNA but also a number of crucial residues to maintain a conformation of the domain necessary to transfer an ‘activation signal’ to the catalytic domain.

【 授权许可】

Unknown