Protein disulfide isomerase does not act as an unfoldase in the disassembly of cholera toxin
Protein disulfide isomerase does not act as an unfoldase in the disassembly of cholera toxin
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DOI：
10.1042/bsr20181320
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发表时间：
2018-10-31
影响因子：
4
通讯作者：
Teter, Ken
中科院分区：
文献类型：
--
作者：
Cherubin, Patrick;Guyette, Jessica;Teter, Ken
霍乱毒素(CT)由一个二硫键连接的A1/A2异二聚体和一个环状的、细胞结合的B同源五聚体组成。催化A1亚单位必须与CTA2/CTB5解离，才能显示其细胞活性。全毒素的分解需要A1/A2二硫键的还原，但还原的CTA1不会自发地与CTA2/CTB5分离：蛋白质二硫键异构酶(PDI)负责将CTA1从CT全毒素中的非共价组装中置换出来。与PDI的接触将CTA1从抗蛋白酶的构象转变为蛋白酶敏感的构象，这被认为代表了PDI介导的CTA1的展开。仅基于这一发现，PDI被广泛认为是一种“解折叠酶”，通过解开全毒素相关的A1亚基来触发毒素分解。与PDI功能的这种展开酶模型相比，我们报告了PDI使CTA1蛋白酶敏感的能力与其在毒素分解中的作用无关。在CTA1中，促进PDI诱导的蛋白酶敏感性的多种条件不支持PDI介导的CT全毒素的分解。此外，阻止PDI诱导的CTA1蛋白酶敏感性的转移并不影响PDI介导的CT全毒素的分解。变性的PDI仍然可以将CTA1转化为酶敏感状态，并且需要等量或过多的摩尔分数的PDI才能有效地将CTA1转化为酶敏感状态，并有效地分解CT全毒素。这些观察结果表明，PDI的“去折叠酶”特性在CT的分解中不起作用，也不代表酶的活性。
Cholera toxin (CT) is composed of a disulfide-linked A1/A2 heterodimer and a ring-like, cell-binding B homopentamer. The catalytic A1 subunit must dissociate from CTA2/CTB5 to manifest its cellular activity. Reduction of the A1/A2 disulfide bond is required for holotoxin disassembly, but reduced CTA1 does not spontaneously separate from CTA2/CTB5: protein disulfide isomerase (PDI) is responsible for displacing CTA1 from its non-covalent assembly in the CT holotoxin. Contact with PDI shifts CTA1 from a protease-resistant conformation to a protease-sensitive conformation, which is thought to represent the PDI-mediated unfolding of CTA1. Based solely on this finding, PDI is widely viewed as an 'unfoldase' that triggers toxin disassembly by unfolding the holotoxin-associated A1 subunit. In contrast with this unfoldase model of PDI function, we report the ability of PDI to render CTA1 protease-sensitive is unrelated to its role in toxin disassembly. Multiple conditions that promoted PDI-induced protease sensitivity in CTA1 did not support PDI-mediated disassembly of the CT holotoxin. Moreover, preventing the PDI-induced shift in CTA1 protease sensitivity did not affect PDI-mediated disassembly of the CT holotoxin. Denatured PDI could still convert CTA1 into a protease-sensitive state, and equal or excess molar fractions of PDI were required for both efficient conversion of CTA1 into a protease-sensitive state and efficient disassembly of the CT holotoxin. These observations indicate the 'unfoldase' property of PDI does not play a functional role in CT disassembly and does not represent an enzymatic activity.