Designed Protein Cages as Scaffolds for Building Multienzyme Materials.,ACS Synthetic Biology

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Designed Protein Cages as Scaffolds for Building Multienzyme Materials.
ACS Synthetic Biology ( IF 3.7 ) Pub Date : 2020-01-17 , DOI: 10.1021/acssynbio.9b00407
Scott A McConnell _{1,

2,

3} , Kevin A Cannon _{1,

2} , Christian Morgan ₄ , Rachel McAllister ₂ , Brendan R Amer _{1,

2,

3} , Robert T Clubb _{1,

2,

3} , Todd O Yeates _{1,

2,

3}

Affiliation

The functions of enzymes can be strongly affected by their higher-order spatial arrangements. In this study we combine multiple new technologies-designer protein cages and sortase-based enzymatic attachments between proteins-as a novel platform for organizing multiple enzymes (of one or more types) in specified configurations. As a scaffold we employ a previously characterized 24-subunit designed protein cage whose termini are outwardly exposed for attachment. As a first-use case, we test the attachment of two cellulase enzymes known to act synergistically in cellulose degradation. We show that, after endowing the termini of the cage subunits with a short "sort-tag" sequence (LPXTG) and the opposing termini of the cellulase enzymes with a short polyglycine sequence tag, addition of sortase covalently attaches the enzymes to the cage with good reactivity and high copy number. The doubly modified cages show enhanced activity in a cellulose degradation assay compared to enzymes in solution, and compared to a combination of singly modified cages. These new engineering strategies could be broadly useful in the development of enzymatic material and synthetic biology applications.

中文翻译：

设计蛋白笼作为构建多酶材料的支架。

酶的功能可能受其高阶空间排列的强烈影响。在这项研究中，我们结合了多种新技术-设计者蛋白笼和蛋白质之间基于分选酶的酶促连接-作为用于以特定配置组织多种（一种或多种类型）酶的新型平台。作为支架，我们使用了以前表征的24个亚基设计的蛋白笼，其末端向外暴露以附着。作为首次使用的案例，我们测试了两种在纤维素降解中具有协同作用的纤维素酶的附着力。我们显示，赋予短的“ sort-tag”序列（LPXTG）笼子亚基的末端和带有短的聚甘氨酸序列标签的纤维素酶的相对末端，添加分选酶可将酶共价结合到笼子上，并具有良好的反应性和高拷贝数。与溶液中的酶相比，与单修饰笼的组合相比，双修饰笼在纤维素降解测定中显示出增强的活性。这些新的工程策略可能在酶材料和合成生物学应用的开发中广泛有用。

更新日期：2020-01-17

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