Identification of the regions involved in DNA binding by the mouse PEBP2α protein

FEBS Letters

Dyson, H.Jane²  Grosschedl, Rudolf¹  Munnerlyn, Audrey¹  Pérez-Alvarado, Gabriela C.²  Wright, Peter E.² 

[1] Howard Hughes Medical Institute and Departments of Microbiology and Biochemistry, University of California, San Francisco, CA 94143, USA;Department of Molecular Biology and the Skaggs Institute for Chemical Biology, The Scripps Research Institute, 10550 N. Torrey Pines Road, La Jolla, CA 92037, USA

关键词: Polyoma virus enhancer binding protein 2α;    Runt domain;    Specific DNA binding;    Conformational change;    Nuclear magnetic resonance spectroscopy;    Chemical shift deviation mapping;    PEBP2α;    polyoma virus enhancer binding protein 2α;    AML1;    acute myeloid leukemia 1;    NMR;    nuclear magnetic resonance;    HSQC;    heteronuclear single quantum coherence;    NOESY;    nuclear Overhauser effect spectroscopy;    NOE;    nuclear Overhauser effect;    TOCSY;    total correlation spectroscopy;   

DOI  :  10.1016/S0014-5793(00)01296-5

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

The polyomavirus enhancer binding protein 2α (PEBP2α) is a DNA binding transcriptional regulatory protein that binds conserved sites in the polyomavirus enhancer, mammalian type C retroviral enhancers and T-cell receptor gene enhancers. Binding of PEBP2α and homologous proteins to the consensus DNA sequence TGPyGGTPy is mediated through a protein domain known as the runt domain. Although recent NMR studies of DNA-bound forms of the runt domain have shown an immunoglobulin-like (Ig) fold, the identification of residues of the protein that are involved in DNA binding has been obscured by the low solubility of the runt domain. Constructs of the mouse PEBP2αA1 gene were generated with N- and C-terminal extensions beyond the runt homology region. The construct containing residues Asp90 to Lys225 of the sequence (PEBP2α90–225) yielded soluble protein. The residues that participate in DNA binding were determined by comparing the NMR spectra of free and DNA-bound PEBP2α90–225. Analysis of the changes in the NMR spectra of the two forms of the protein by chemical shift deviation mapping allowed the unambiguous determination of the regions that are responsible for specific DNA recognition by PEBP2α. Five regions in PEBP2α90–225 that are localized at one end of the β-barrel were found to interact with DNA, similar to the DNA binding interactions of other Ig fold proteins.

【 授权许可】

Unknown