肌細胞核因子(Myocyte Nuclear Factor, MNF)是一種在老鼠體內會選擇性的表現在myogenic stem cell的轉錄因子。而肌細胞核因子所調節的基因主要在負責協調myogenic stem cell的增殖與分化，例如當肌細胞受傷時肌細胞核因子便會有大量表現的現象發生。在生物體內的肌細胞核因子可經由選擇性接合(alternative splicing)產生兩種不同的cDNA，他們各自轉譯為肌細胞核因子α及肌細胞核因子β，轉譯所得的蛋白質分別含617及414個胺基酸；他們的DNA結合區域雖是相同的，然而肌細胞核因子α及肌細胞核因子β在功能上的表現卻是完全不同的；他們在體外試驗中，對於與DNA結合的能力上有著截然不同的特徵；另外，transient-transfection assays中所表現出調控轉錄的特性也有所差異。
肌細胞核因子α、肌細胞核因子βDNA結合區域的範圍包括第289到389個殘基，且屬於winged-helix family的成員；從胺基酸的序列比對發現，肌細胞核因子之DNA結合區域與已知超過200個的winged-helix family具有35﹪~89﹪的相似性；雖然這個DNA結合區域十分具有保留性，但近來卻發現不同的winged-helix family蛋白質能以其獨特的方式辨認特異的DNA序列，繼而對各自下游的基因進行調控作用。為了釐清肌細胞核因子的DNA結合區域在結構上與功能上的關係，我們首先以E.coli系統表現出肌細胞核因子的DNA結合區域，產量大約是40~50mg/L，再利用多維異核核磁共振光譜(heteronuclear multidimensional NMR spectroscopy)推論出肌細胞核因子的DNA結合區域之三級結構；根據三級結構的分析指出，在原本應屬於第二個翅膀狀結構的區域產生了一個新的α-helix結構，這或許與winged helix family的DNA結合能力之專一性有關；另外肌細胞核因子的二級結構與間白素結合因子(ILF-DBD)很相似；綜合結構上的結果，我們比較了肌細胞核因子與ILF-DBD及其他winged-helix family的蛋白質在DNA結合能力的專一性上之不同,並且提出一些新的論點，解釋這個家族與DNA結合能力的專一性之可能原因。

Myocyte nuclear factors (MNFs) are transcription factors that are expressed selectivity in mouse myogenic stem cells. MNFs regulate the genes that coordinate the proliferation and differentiation of myogenic stem cells after muscle injury. Two MNF isoforms are found, MNF-α and MNF-β each contains 617 and 414 amino acids, respectively. They are derived from a single MNF gene by alternative splicing. However, the expressions of MNF-α and MNF-β are differentially regulated. They possess distinctive functional properties with respect to DNA binding in vitro and transcriptional regulatory activity in transient—transfection assays. The DNA-binding domains of MNFs belongs to the winged helix/forkhead family since the residues from 289 to 389 of MNFs share 35﹪to 89﹪similarity with other known members of this family. Members of this family are characterized by a conserved 100-amino acid DNA-binding domain that contain threeα-helices (H1, H2, and H3), threeβ-strands (S1 and S2), and two wing-like loops (W1 and W2). Although the DNA binding domains of the winged helix family proteins are conserved, they can recognize specific DNA sequence to regulate their own downstream genes. In order to elucidate the structural and functional relationships of the DNA-binding domain of MNFs, we expressed the protein in E.coli with a yield of 40-50mg/L. We also assigned 1H, 13C and 15N resonances and deduced the secondary structure of the DNA-binding domain of MNF from multidimensional NMR spectroscopy. The secondary structures of the DNA-binding domain of MNF are similar to those of interleukin enhancer binding factor (ILF). Based on the resulting structures, their DNA-binding specificities were compared. Secondary structure analysis revealed that the wing 2 region contains an extra α-helix, which may be responsible for the DNA-binding specificity differences among members of winged helix family. This result suggests the possible structural diversity in the winged helix/forkhead proteins.

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