Membraneless Compartmentalization Facilitates Enzymatic Cascade Reactions and Reduces Substrate Inhibition.
Membraneless Compartmentalization Facilitates Enzymatic Cascade Reactions and Reduces Substrate Inhibition.
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DOI：
10.1021/acsami.8b07573
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发表时间：
2018-09-26
影响因子：
9.5
通讯作者：
Takayama S
中科院分区：
文献类型：
--
作者：
Kojima T;Takayama S
活细胞具有由液-液分离形成的无膜细胞器。为了更好地了解无膜微室的一般功能，本文利用多水相体系构建了无细胞多室反应系统。无膜凝聚液滴被放置在分子拥挤的环境中，较大的葡聚糖(DEX)液滴被淹没在聚乙二醇(PEG)溶液中。凝聚液滴具有很长的滞留时间，能够隔离试剂和酶，并通过液-液界面示出多步级联反应。改变相动力学的能力也通过盐介导的凝聚液滴的溶解来证明，这导致单独隔离的试剂和酶的释放和混合。最后，由于无膜细胞器中的相分离物质通常是细胞器中隔离或排除的酶的底物和底物类似物，因此本文探讨了DEX与DEX的水解酶--葡聚糖酶之间的相互作用。结果表明，当葡聚糖酶直接分配到DEX相中时，会受到底物抑制，但这种抑制可以通过在与DEX相相邻但相分离的凝聚液滴中的葡聚糖酶的区隔来减轻和极大地加速反应。酶的区隔可以通过减轻底物抑制来加速反应的见解是特别新颖的，这是人工无膜细胞器样系统可能为生理细胞功能提供新见解的一个例子。
Living cells possess membraneless-organelles formed by liquid-liquid phase separation. With the aim of better understanding the general functions of membrane-less microcompartments, this paper constructs acellular multi-compartment reaction systems using an aqueous multi-phase system. Membrane-less coacervate droplets are placed within a molecularly crowded environment where a larger dextran (DEX) droplet is submerged in a polyethylene glycol (PEG) solution. The coacervate droplets are capable of sequestering reagents and enzymes with a long retention time, and demonstrate multi-step cascading reactions through the liquid-liquid interfaces. The ability to change phase dynamics is also demonstrated through salt-mediated dissolution of coacervate droplets, which leads to release and mixing of separately sequestered reagents and enzymes. Finally, as phase-separated materials in membraneless organelles are often substrates and substrate analogues for the enzymes sequestered or excluded in the organelles, this paper explores the interaction between DEX and dextranase, an enzyme that hydrolyzes DEX. Results revealed that dextranase suffers from substrate inhibition when partitioned directly in a DEX phase but that this inhibition can be mitigated and reactions greatly accelerated by compartmentalization of dextranase inside a coacervate droplet that is adjacent to, but phase-separated from, the DEX phase. The insight that compartmentalization of enzymes can accelerate reactions by mitigating substrate inhibition is particularly novel and is an example where artificial membrane-less organelle-like systems may provide new insights into physiological cell functions.
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影响因子：
4.3
作者：
Audas, Timothy E.;Jacob, Mathieu D.;Lee, Stephen
通讯作者：
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影响因子：
56.9
作者：
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DOI：
10.1073/pnas.1509317112
发表时间：
2015-09-22
影响因子：
11.1
作者：
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通讯作者：
Brangwynne, Clifford P.
影响因子：
21.8
作者：
Aumiller, William M., Jr.;Keating, Christine D.
通讯作者：
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影响因子：
64.5
作者：
Feric M;Vaidya N;Harmon TS;Mitrea DM;Zhu L;Richardson TM;Kriwacki RW;Pappu RV;Brangwynne CP
通讯作者：
Brangwynne CP