Tsukagoshi N, Ando Y, Tomita Y, Uchida R, Takemura T, Sasaki T, Yamagata H, Udaka S, Ichihara Y, Takahashi K
Department of Food Science and Technology, Faculty of Agriculture, Nagoya University, Japan.
Gene. 1988 May 30;65(2):285-92. doi: 10.1016/0378-1119(88)90465-9.
A clone, pSPcA2, which carries the full-length swine pepsinogen cDNA was isolated. The coding sequence comprised the signal peptide [15 amino acids (aa)], the activation peptide segment (44 aa) and mature pepsin (327 aa). The deduced amino acid sequence agrees with the published sequence with two exceptions. Asparagine instead of aspartate is present at aa positions 19 and 308. Two types of plasmids, pAS and pUCtacSPc series, were constructed for expressing swine pepsinogen cDNA. These plasmids directed the synthesis of polypeptides which were detected by employing an antibody to swine pepsinogen. However, all the polypeptides formed aggregates and showed no acid protease activity. Only the protein directed by pAS5 regained the acid protease activity after renaturation procedures. The activity was completely inhibited by pepstatin. Furthermore, the renatured pAS5 protein was spontaneously converted to pepsin under acidic conditions. The presence of Arg-8 in the activation peptide segment appears important for the stabilization of the pepsinogen molecule.
分离出了携带猪胃蛋白酶原全长cDNA的克隆体pSPcA2。编码序列包括信号肽（15个氨基酸）、激活肽段（44个氨基酸）和成熟胃蛋白酶（327个氨基酸）。推导的氨基酸序列与已发表序列一致，但有两个例外。在第19和308位氨基酸处，存在天冬酰胺而非天冬氨酸。构建了两种类型的质粒，即pAS和pUCtacSPc系列，用于表达猪胃蛋白酶原cDNA。这些质粒指导合成的多肽通过使用抗猪胃蛋白酶原抗体进行检测。然而，所有多肽都形成了聚集体，且没有酸性蛋白酶活性。只有pAS5指导合成的蛋白质在复性后恢复了酸性蛋白酶活性。该活性被胃蛋白酶抑制剂完全抑制。此外，复性后的pAS5蛋白在酸性条件下会自发转化为胃蛋白酶。激活肽段中Arg-8（精氨酸-8）的存在对于胃蛋白酶原分子的稳定似乎很重要。
