ESSENTIAL CELLULAR FUNCTION OF HYPUSINE IN ELF 5A

真核蛋白翻译起始因子5A（eIF-5A）含有 一个羟腐胺赖氨酸的残基，似乎是唯一的细胞蛋白质 这种独特的氨基酸。 羟腐胺赖氨酸是在 通过转移的丁胺部分， 多胺亚精胺与eIF-5A前体中的赖氨酰残基反应， 形成脱氧羟腐胺赖氨酸，然后羟基化形成羟腐胺赖氨酸。 为了鉴定与eIF-5A 相互作用以发挥其生物活性，我们已经产生了一种 亲和法分离。多组氨酸标记的人eIF- 5A前体蛋白（His-tag-ec-eIF-5A）通过利用 pET-24 b载体。含脱氧羟腐胺赖氨酸的形式通过以下制备： 脱氧羟腐胺赖氨酸合酶中His-标签-ec-eIF-5A的修饰 反应His标记的蛋白质与Ni（II）-NTA-琼脂糖结合 柱被用作亲和矩阵以分离和鉴定 与eIF-5A特异性相互作用的细胞蛋白。虽然 已经确定羟腐胺赖氨酸对于真核细胞是至关重要 增殖，羟腐胺赖氨酸的确切生理作用， 含有eIF-5A蛋白的基因尚不清楚。 我们也在研究 eIF-5A前体蛋白与脱氧羟腐胺赖氨酸特异性相互作用 通过使用His-tag-ec-eIF-5A或His-tag- 脱氧羟腐胺赖氨酸合酶作为亲和性结合到Ni（II）树脂上 矩阵 在另一个实验室最近的一项研究中，据报道，eIF-5A 在复制中Rev功能所需的宿主细胞因子 HIV-1我们目前正在研究羟腐胺赖氨酸的作用 eIF-5A和Rev相互作用的修饰以及可能的 羟腐胺赖氨酸对HIV-1复制干预作用 合成.

Eukaryotic protein translation initiation factor 5A (eIF-5A) contains one residue of hypusine and appears to be the only cellular protein with this unique amino acid. Hypusine is produced post- translationally by transfer of the butylamine portion of the polyamine spermidine to a lysyl residue in the eIF-5A precursor to form deoxyhypusine followed by hydroxylation to form hypusine. In an effort to identify cellular proteins with which eIF-5A interacts to exert its biological activity, we have originated an affinity method for their isolation. Polyhistidine-tagged human eIF- 5A precursor protein (His-tag-ec-eIF-5A) was produced by utilizing pET-24b vector. The deoxyhypusine-containing form was prepared by modification of His-tag-ec-eIF-5A in the deoxyhypusine synthase reaction. The his-tagged proteins bound to a Ni (II)-NTA-agarose column are being used as an affinity matrix to isolate and identify cellular proteins that specifically interact with eIF-5A. Although it is well established that hypusine is vital for eukaryotic cell proliferation, the precise physiological role of the hypusine- containing protein eIF-5A is yet unknown. We are also studying the specific interaction of eIF-5A precursor protein with deoxyhypusine synthase in vitro by the use of His-tag-ec-eIF-5A or His-tag- deoxyhypusine synthase bound to the Ni(II) resin as an affinity matrix. In a recent study from another laboratory, eIF-5A was reported to be a host cellular factor required for Rev function in the replication of HIV-1. We are currently investigating the role of hypusine modification in the interaction of eIF-5A and Rev and the possible intervention of HIV-1 replication through inhibition of hypusine synthesis.