3D solution structure of the DNA-binding domain of Interleukin enhancer binding factor by NMR

• Main Authors: Pei-Phen Liu, 劉沛棻
• Other Authors: Woei-Jer Chuang
• Format: Others
• Language: zh-TW
• Published: 2001
• Online Access: http://ndltd.ncl.edu.tw/handle/33907794719472380206

碩士 === 國立成功大學 === 生物化學研究所 === 89 === Members of the forkhead/winged helix family of transcription factors have been shown to play important roles in development, inflammation, and neoplasia. Members of this family are characterized by a conserved 100-amino acid DNA-binding domain called the forkhead or winged helix domain. Members of the winged helix family contain three a-helices, three b-strands, and two wings. Two major regions are involved in the protein and DNA interactions: helix 3, the recognition site which makes contacts with the major groove of DNA, and wing 2, which interacts with the minor groove of DNA. Since the discovery of the winged helix/fork head motif, a large number of topologically related proteins with diverse biological functions have been characterized by both X-ray crystallography and NMR spectroscopy. Recently, a winged helix transcription factor (RFX1) has been shown to bind the major groove of DNA with wing 1. This surprising observation suggests that the winged helix proteins with similar folds may have different modes for DNA recognition. Therefore, the differences in DNA binding specificity of winged helix family are determined by variations in the contact residues on the recognition element, including wings or/and recognition helix. Interleukin enhancer binding factors (ILFs) are transcription factors that bind to purine-rich regulatory motifs in both human T-cell leukemia virus long terminal region (HILV-1 LTR) and the interleukin-2 (IL-2) promoter. Specifically, ILFs bind to the antigen receptor response element 2 (ARRE-2) of IL-2 promoter. Although the function of ILFs is not well understood, previous studies suggest that ILFs may have a positive role in regulating IL-2 gene expression. To date, three ILFs have been found: ILF-1, ILF-2, and ILF-3 with 655, 609, and 323 amino acids, respectively. Both ILF-1 and ILF-2 contain several amino acid homologies including a region for potential ubiquitin-mediated degradation, a nuclear localization signal, an N-glycosylation motif, and a DNA-binding domain. The DNA-binding domain belongs to the winged helix family since residues from 251 to 348 of ILF-1 and ILF-2 share 35 to 88% similarity with other known members of this family. We have assigned 1H, 13C, and 15N resonances and deduced the solution structure of the DNA-binding domain of ILF by multidimensional NMR spectroscopy. Our structural analysis revealed that the wing 2 region contains an extra a helix which may be responsible the DNA-binding specificity differences among winged helix family. This is the first study to report the presence of a helix in the wing 2 region of winged helix family.