Spores, the infectious agents of many Firmicutes, are remarkably resilient cell forms. Even distant relatives can have similar spore architectures although some display unique features; they all incorporate protective proteinaceous envelopes. We previously found that Bacillus spores can achieve these protective properties through extensive disulfide cross-linking of self-assembled arrays of cysteine-rich proteins. We predicted that this could be a mechanism employed by spore formers in general, even those from other genera. Here, we tested this by revealing in nanometer detail how the outer envelope (exosporium) in Clostridium sporogenes (surrogate for C. botulinum group I), and in other clostridial relatives, forms a hexagonally symmetric semipermeable array. A cysteine-rich protein, CsxA, when expressed in Escherichia coli, self-assembles into a highly thermally stable structure identical to that of the native exosporium. Like the exosporium, CsxA arrays require harsh “reducing” conditions for disassembly. We conclude that in vivo, CsxA self-organizes into a highly resilient, disulfide cross-linked array decorated with additional protein appendages enveloping the forespore. This pattern is remarkably similar to that in Bacillus spores, despite a lack of protein homology. In both cases, intracellular disulfide formation is favored by the high lattice symmetry. We have identified cysteine-rich proteins in many distantly related spore formers and propose that they may adopt a similar strategy for intracellular assembly of robust protective structures.

中文翻译：

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梭状芽孢杆菌和肉毒梭菌孢子表面的结构和自组装说明了孢子形成者的一般保护策略。

孢子是许多厚壁菌门的传染因子，是非常有弹性的细胞形式。甚至远亲也可能具有相似的孢子结构，尽管有些具有独特的特征；它们都包含保护性蛋白质包膜。我们以前发现芽孢杆菌孢子可以通过自组装的富含半胱氨酸的蛋白质阵列的广泛二硫键交联来实现这些保护特性。我们预测这可能是孢子形成者普遍采用的一种机制，甚至是来自其他属的孢子形成者。在这里，我们通过在纳米细节上揭示了产孢梭菌（肉毒梭菌的替代物）中的外膜（外孢子组 I) 和其他梭菌亲属中，形成六边形对称的半透阵列。富含半胱氨酸的蛋白质 CsxA 在大肠杆菌中表达时，会自组装成与天然外孢子相同的高度热稳定结构。与外孢子一样，CsxA 阵列需要苛刻的“还原”条件进行拆卸。我们得出的结论是，在体内，CsxA 自组织成一个高弹性的二硫化物交联阵列，该阵列装饰有包裹前孢子的额外蛋白质附属物。这种模式与芽孢杆菌中的模式非常相似孢子，尽管缺乏蛋白质同源性。在这两种情况下，高晶格对称性有利于细胞内二硫化物的形成。我们已经在许多远缘孢子形成者中鉴定了富含半胱氨酸的蛋白质，并提出它们可能采用类似的策略来组装强大的保护结构。