Proteolytic Maturation of the Outer Membrane c-Type Cytochrome OmcZ by a Subtilisin-Like Serine Protease Is Essential for Optimal Current Production by Geobacter sulfurreducens
Proteolytic Maturation of the Outer Membrane c-Type Cytochrome OmcZ by a Subtilisin-Like Serine Protease Is Essential for Optimal Current Production by Geobacter sulfurreducens
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枯草杆菌蛋白酶样丝氨酸蛋白酶对外膜 c 型细胞色素 OmcZ 的蛋白水解成熟对于硫还原地杆菌的最佳电流生产至关重要
DOI：
10.1128/aem.02617-20
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发表时间：
2021
影响因子：
4.4
通讯作者：
Kengo Inoue
中科院分区：
文献类型：
--
作者：
Ayako kai;Takahiro Tokuishi;Takashi Fujikawa;Yoshihiro Kawano;Toshiyuki Ueki;Miyuki Nagamine;Yoichi Sakakibara;Masahito Suiko;Kengo Inoue
硫还原地杆菌的外膜型细胞色素（OmcZ）对微生物燃料电池的最佳电流生产至关重要。OmcZ有大小两种形式，分别命名为OmcZL和OmcZL。然而，这两种结构是如何形成的尚不清楚。编码枯草菌素样丝氨酸蛋白酶的GSU2075基因被破坏的突变体，位于omcz的下游，产生与omcz缺陷突变株相似的低电流。生化分析表明，积累的ozpamutant不会产生OmcZS，而OmcZS被认为是细胞外电子转移到电极所必需的成熟形式。从产生OzpA裂解OmcZLand的大肠杆菌菌株的异源表达系统细胞裂解物产生蛋白水解产物OmcZSas。在野生型G.硫还原菌的培养上清、松散结合的外表面和细胞内蛋白部分中，只有培养上清蛋白部分显示出OmcZ裂解活性，表明在细胞外可以产生成熟形式的OmcZ，即OmcZS。这些结果表明，OzpA是通过OmcZ成熟产生电流的必需蛋白酶，而omczz是细胞外电子向电极转移的关键。这种OmcZ的蛋白水解成熟是硫还原G.中已知类型细胞色素的独特调控。微生物燃料电池是一种很有前途的从各种废物中发电的技术。然而，微生物胞外电子向电极转移的分子机制还有待阐明。G.硫还原菌是混合培养微生物燃料电池系统中发电的关键角色，也是研究细胞外电子转移的模式微生物。外膜型细胞色素OmcZ对G.硫还原菌的最佳电流生产至关重要。OmcZ蛋白水解裂解发生在成熟过程中，但潜在的机制尚不清楚。本研究鉴定了一种类似枯草菌素的蛋白酶OzpA，它在切割OmcZ和产生成熟形式的OmcZ （OmcZS）中起作用。OzpA对于当前的生产和OmcZ的蛋白水解成熟是必不可少的。这是c型细胞色素调控硫还原g细胞外电子转移的新发现。该研究还为微生物胞外电子转移的设计策略和发展提供了新的见解，以实现从化学能到电能的有效转换。
An outer membranec-type cytochrome (OmcZ) in Geobacter sulfurreducens is essential for optimal current production in microbial fuel cells. OmcZ exists in two forms, small and large, designated OmcZSand OmcZL, respectively. However, it is still not known how these two structures are formed. A mutant with a disruption of the GSU2075 gene encoding a subtilisin-like serine protease (designatedozpAfor theOmcZprotease), which is located downstream ofomcZ, produced low currents at a level similar to that of theomcZ-deficient mutant strain. Biochemical analyses revealed that theozpAmutant accumulated OmcZLand did not produce OmcZS, which is thought to be a mature form that is essential for the extracellular electron transfer to the electrode. A heterologous expression system cell lysate from an Escherichia coli strain producing OzpA cleaved OmcZLand generated OmcZSas the proteolytic product. Among the culture supernatant, loosely bound outer surface, and intracellular protein fractions from wild-type G. sulfurreducens, only the culture supernatant protein fraction showed OmcZLcleavage activity, indicating that the mature form of OmcZ, OmcZS, can be produced outside the cells. These results indicate that OzpA is an essential protease for current production via the maturation of OmcZ, and OmcZSis the key to the extracellular electron transfer to electrodes. This proteolytic maturation of OmcZ is a unique regulation among knownc-type cytochromes in G. sulfurreducens.IMPORTANCEMicrobial fuel cells are a promising technology for energy generation from various waste types. However, the molecular mechanisms of microbial extracellular electron transfer to the electrode need to be elucidated. G. sulfurreducens is a common key player in electricity generation in mixed-culture microbial fuel cell systems and a model microorganism for the study of extracellular electron transfer. Outer membranec-type cytochrome OmcZ is essential for an optimal current production by G. sulfurreducens. OmcZ proteolytic cleavage occurs during maturation, but the underlying mechanism is unknown. This study identifies a subtilisin-like protease, OzpA, which plays a role in cleaving OmcZ and generating the mature form of OmcZ (OmcZS). OzpA is essential for current production and, thus, the proteolytic maturation of OmcZ. This is a novel regulation of thec-type cytochrome for G. sulfurreducens extracellular electron transfer. This study also provides new insights into the design strategy and development of microbial extracellular electron transfer for an efficient energy conversion from chemical energy to electricity.
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