The C-Terminal Zwitterionic Sequence of CotB1 Is Essential for Biosilicification of the Bacillus cereus Spore Coat
The C-Terminal Zwitterionic Sequence of CotB1 Is Essential for Biosilicification of the Bacillus cereus Spore Coat
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DOI：
10.1128/jb.00447-15
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发表时间：
2015-10
影响因子：
3.2
通讯作者：
K. Motomura;Takeshi Ikeda;S. Matsuyama;Mohamed A. A. Abdelhamid-Mohamed-A.-A.-Abdelhamid-47118162;Tatsuya Tanaka;Takenori Ishida;R. Hirota;A. Kuroda
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文献类型：
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作者：
K. Motomura;Takeshi Ikeda;S. Matsuyama;Mohamed A. A. Abdelhamid-Mohamed-A.-A.-Abdelhamid-47118162;Tatsuya Tanaka;Takenori Ishida;R. Hirota;A. Kuroda
摘要 二氧化硅沉积在蜡状芽孢杆菌的孢子衣层及其周围，增强了孢子的耐酸性。多种肽和蛋白质，包括硅藻硅蜡和硅酸素肽，参与真核二氧化硅生物矿化（生物硅化）。同源序列搜索揭示了蜡状芽胞杆菌孢子外壳蛋白 CotB1 的 C 末端区域有一个类似 silacidin 的序列。带负电的硅酸丁样序列后面是带正电的富含精氨酸的 14 个氨基酸序列，这与硅蜡非常相似。这些序列赋予 CotB1 的 C 末端两性离子特征。有趣的是，cotB1 基因似乎与其旁系同源物 cotB2 形成双顺反子操纵子，但其产物缺乏 C 端两性离子序列。 ΔcotB1B2 突变菌株与野生型细菌生长速度和形成孢子的速度相同，但没有表现出生物硅化。互补分析表明，CotB1（但 CotB2 和 CotB1 C 端截短突变体）都不能恢复 ΔcotB1B2 突变体的生物硅化活性，这表明 CotB1 C 端两性离子序列对于该过程至关重要。我们发现 CotB1 表达的动力学及其定位与生物硅化的时间进程和沉积二氧化硅的位置密切相关。据我们所知，这是直接参与原核生物硅化的蛋白质的首次报道。重要性 生物硅化是生物体以不溶性二氧化硅形式吸收可溶性硅酸盐的过程。尽管真核生物硅化的机制已被深入研究，但原核生物硅化直到最近才被研究。我们之前证明了蜡样芽孢杆菌及其近亲中发生生物硅化，并且二氧化硅沉积在孢子包衣层内和周围作为抗酸的保护涂层。本研究表明，蜡样芽胞杆菌孢子外壳蛋白 CotB1 携带 C 端两性离子序列，对于生物硅化至关重要。我们的结果首次深入了解原核生物生物硅化所需的机制。
ABSTRACT Silica is deposited in and around the spore coat layer of Bacillus cereus, and enhances the spore's acid resistance. Several peptides and proteins, including diatom silaffin and silacidin peptides, are involved in eukaryotic silica biomineralization (biosilicification). Homologous sequence search revealed a silacidin-like sequence in the C-terminal region of CotB1, a spore coat protein of B. cereus. The negatively charged silacidin-like sequence is followed by a positively charged arginine-rich sequence of 14 amino acids, which is remarkably similar to the silaffins. These sequences impart a zwitterionic character to the C terminus of CotB1. Interestingly, the cotB1 gene appears to form a bicistronic operon with its paralog, cotB2, the product of which, however, lacks the C-terminal zwitterionic sequence. A ΔcotB1B2 mutant strain grew as fast and formed spores at the same rate as wild-type bacteria but did not show biosilicification. Complementation analysis showed that CotB1, but neither CotB2 nor C-terminally truncated mutants of CotB1, could restore the biosilicification activity in the ΔcotB1B2 mutant, suggesting that the C-terminal zwitterionic sequence of CotB1 is essential for the process. We found that the kinetics of CotB1 expression, as well as its localization, correlated well with the time course of biosilicification and the location of the deposited silica. To our knowledge, this is the first report of a protein directly involved in prokaryotic biosilicification. IMPORTANCE Biosilicification is the process by which organisms incorporate soluble silicate in the form of insoluble silica. Although the mechanisms underlying eukaryotic biosilicification have been intensively investigated, prokaryotic biosilicification was not studied until recently. We previously demonstrated that biosilicification occurs in Bacillus cereus and its close relatives, and that silica is deposited in and around a spore coat layer as a protective coating against acid. The present study reveals that a B. cereus spore coat protein, CotB1, which carried a C-terminal zwitterionic sequence, is essential for biosilicification. Our results provide the first insight into mechanisms required for biosilicification in prokaryotes.