Folding of egg white proteins as a post-translational processing.

利用体外翻译系统和变性复性系统研究了激素调控下鸡输卵管中合成的卵清白色蛋白--卵转铁蛋白和卵清蛋白的折叠机制，发现两步复性法可以有效地复性复杂蛋白--卵转铁蛋白。在第一步中，将蛋白质的还原和变性形式在含有还原型谷胱甘肽的非变性缓冲液中在低温下孵育;在第二步中，在氧化型谷胱甘肽存在下，在较高温度下将还原形式再氧化。在这些条件下，完全还原形式的卵转铁蛋白和它的半分子几乎定量再氧化，以恢复铁结合能力和构象，非常相似的天然形式。圆二色光谱表明，在低温下，完全还原的形式有部分折叠的构象，这是波动像“熔融球”状态。整个ovotransferrin和两个半分子之间的再氧化动力学比较支持独立的N-和C-末端domains.With卵清蛋白约40%的尿素变性蛋白复性后，在非变性条件下孵育18小时的天然形式。同样，在麦胚翻译系统中，只有一部分mRNA定向翻译产物具有天然构象。

Egg white proteins, ovotransferrin and ovalbumin, which are synthesized in hen oviducts under hormonal regulation, were investigated for their folding mechanisms using in vitro translation system as well as refolding systems of denatured forms.A two-step procedure was found to be useful for the efficient refolding of a complex protein, ovotransferrin. In the first step, the reduced and denatured form of the protein was incubated at a low temperature in a nondenaturing buffer containing reduced glutathione; in the second step, the reduced form was reoxidized at a higher temperature in the presence of oxidized glutathione. Under these conditions, the fully reduced forms of ovotransferrin and its half-molecules were almost quantitatively reoxidized to regain iron-binding abilities and conformations, very similar to the native form. The circular dichroism spectra revealed that at low temperatures the fully reduced forms have partially folded conformations, which are fluctuating like "molten globule" states. The reoxidization kinetics compared between whole ovotransferrin and the two half-molecules supported independent refolding of the N- and C-terminal domains.With respect to ovalbumin about 40% of urea-denatured protein was renatured to the native form after 18 hr incubation under non-denaturing conditions. Likewise, only a part of the mRNA-directed translation product in wheat germ translation system was found to take a native-like conformation.