Cytochrome C with peroxidase-like activity encapsulated inside the small DPS protein nanocage
Cytochrome C with peroxidase-like activity encapsulated inside the small DPS protein nanocage
复制标题
具有过氧化物酶样活性的细胞色素 C 封装在小型 DPS 蛋白纳米笼内
DOI：
10.1039/d1tb00234a
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发表时间：
2021
影响因子：
7
通讯作者：
Douglas, Trevor
中科院分区：
文献类型：
--
作者：
Waghwani, Hitesh Kumar;Douglas, Trevor
大自然利用自组装的蛋白质结构作为原核生物的亚细胞区室，为特殊的生化反应隔离催化剂。这些蛋白质笼结构为被封装的酶提供了独特的隔离环境。了解这些系统对合成催化细胞器样纳米材料的生物启发设计是有用的。从solfataricus中分离的饥饿细胞DNA结合蛋白（Dps）是一个9 nm的十二聚体蛋白笼，使其成为已知最小的天然蛋白质笼。它在氧化应激反应中自然过度表达。在体内实验中，小体积、自然生物分布到肾脏以及能够穿过肾小球滤过屏障的能力突出了其作为合成抗氧化剂的潜力。细胞色素C （CytC）是一种小的血红素蛋白，具有过氧化物酶样活性，参与电子传递链，在细胞凋亡中起关键作用。在这里，我们报道了CytC在Dps的5nm内腔内的封装，并证明了所得到的Dps纳米腔的催化活性和增强的抗氧化行为。小腔体可以容纳单个CytC，这是通过由Dps亚基和CytC融合的Dps亚基组成的嵌合笼的自组装实现的。为了选择性分离含有Dps的CytC，我们使用了仅存在于酶融合Dps亚基（6His-Dps-CytC）上的工程多组氨酸标签。以愈创木酚和3,3 ',5,5 ' -四甲基联苯胺（TMB）为底物，研究了包封CytC的催化活性，并与游离（未包封）CytC的活性进行了比较。与游离酶相比，封装的CytC表现出更好的pH依赖性催化活性，并为设计这些小蛋白质笼作为催化纳米反应器的潜力提供了概念验证模型。
Nature utilizes self-assembled protein-based structures as subcellular compartments in prokaryotes to sequester catalysts for specialized biochemical reactions. These protein cage structures provide unique isolated environments for the encapsulated enzymes. Understanding these systems is useful in the bioinspired design of synthetic catalytic organelle-like nanomaterials. The DNA binding protein from starved cells (Dps), isolated from Sulfolobus solfataricus, is a 9 nm dodecameric protein cage making it the smallest known naturally occurring protein cage. It is naturally over-expressed in response to oxidative stress. The small size, natural biodistribution to the kidney, and ability to cross the glomerular filtration barrier in in vivo experiments highlight its potential as a synthetic antioxidant. Cytochrome C (CytC) is a small heme protein with peroxidase-like activity involved in the electron transport chain and also plays a critical role in cellular apoptosis. Here we report the encapsulation of CytC inside the 5 nm interior cavity of Dps and demonstrate the catalytic activity of the resultant Dps nanocage with enhanced antioxidant behavior. The small cavity can accommodate a single CytC and this was achieved through self-assembly of chimeric cages comprising Dps subunits and a Dps subunit to which the CytC was fused. For selective isolation of CytC containing Dps cages, we utilized engineered polyhistidine tag present only on the enzyme fused Dps subunits (6His-Dps-CytC). The catalytic activity of encapsulated CytC was studied using guaiacol and 3,3′,5,5′-tetramethylbenzidine (TMB) as two different peroxidase substrates and compared to the free (unencapsulated) CytC activity. The encapsulated CytC showed better pH dependent catalytic activity compared to free enzyme and provides a proof-of-concept model to engineer these small protein cages for their potential as catalytic nanoreactors.