Department of Food Science, University of Guelph, Guelph, ON, Canada N1G2W1.
Protein Eng Des Sel. 2010 Jan;23(1):19-26. doi: 10.1093/protein/gzp066.
Proplasmepsin II (zPMII) represents a unique member of the aspartic proteinase family, with a prosegment-enzyme interaction that is thus far unique among the pepsin-like proteases. The role of the prosegment in aspartic proteinase structure and function was investigated by generating two chimeric proteins, one with the pepsinogen prosegment fused to the mature region of PMII (pepproPMII) and a second with the prosegment of PMII fused to pepsin (PMIIpropep). Both chimeras were expressed using Escherichia coli; however, PMIIpropep was extremely unstable suggesting protein misfolding. Alternatively, pepproPMII was capable of both autoactivation and hydrolysis of a synthetic substrate. Similarly, when the PMII enzyme was expressed without a prosegment, it too exhibited activity against the synthetic enzyme. CD measurements indicated that pepproPMII had reduced thermal stability when compared with zPMII. This reduction of temperature stability may have resulted from the inability of the pepsinogen prosegment to stabilize the C-terminal domain of the PMII enzyme. The ability of PMII to fold in the presence of a completely non-homologous prosegment and in its absence suggests that prosegment is not critical to obtaining a functional enzyme in all pepsin-like enzymes but likely plays a role in protein stabilization.
原浆朊酶 II（zPMII）是天冬氨酸蛋白酶家族中的一个独特成员，其前肽-酶相互作用在胃蛋白酶样蛋白酶中是独一无二的。前肽在天冬氨酸蛋白酶结构和功能中的作用是通过生成两种嵌合蛋白来研究的，一种是将胃蛋白酶原前肽融合到 PMII 的成熟区域（pepproPMII），另一种是将 PMII 的前肽融合到胃蛋白酶（PMIIpropep）。这两种嵌合体都使用大肠杆菌进行表达；然而，PMIIpropep 极不稳定，表明蛋白质错误折叠。另一方面，pepproPMII 既能自动激活又能水解合成底物。同样，当没有前肽表达 PMII 酶时，它也对合成酶表现出活性。CD 测量表明，与 zPMII 相比，pepproPMII 的热稳定性降低。这种温度稳定性的降低可能是由于胃蛋白酶原前肽无法稳定 PMII 酶的 C 末端结构域。PMII 在完全非同源前肽存在和不存在的情况下能够折叠的能力表明，前肽对于所有胃蛋白酶样酶获得功能性酶并非至关重要，但可能在蛋白质稳定中发挥作用。
