DWI-Leibniz Institute for Interactive Materials, Aachen 52074, Germany.
Institute of Technical and Macromolecular Chemistry, RWTH Aachen University, Aachen 52074, Germany.
J Phys Chem B. 2024 Nov 7;128(44):10861-10869. doi: 10.1021/acs.jpcb.4c05990. Epub 2024 Oct 30.
Weak nonspecific interactions between biomacromolecules determine the cytoplasmic organization. Despite their importance, it is challenging to determine these interactions in the intracellular dense and heterogeneous mixture of biomacromolecules. Here, we develop a method to indicate electrostatic and hydrophobic associative interactions and map these interactions. The method relies on a genetically encoded probe containing a sensing peptide and a circularly permuted green fluorescent protein that provides a ratiometric readout. Inside bacterial and mammalian cells, we see that the cytoplasmic components interact strongly with cationic and hydrophobic probes but not with neutral hydrophilic probes, which remain inert. The cytoplasm interacts strongly with highly negatively charged hydrophilic probes, but the HEK293T cytoplasm does not. These associative interactions are modulated by ATP depletion. Hence, the nonspecific associative interaction profile in cells is condition- and species-dependent.
生物大分子之间的弱非特异性相互作用决定了细胞质的组织。尽管这些相互作用非常重要，但在细胞内生物大分子密集且不均匀的混合物中确定这些相互作用具有挑战性。在这里，我们开发了一种方法来指示静电和疏水相互作用，并对这些相互作用进行作图。该方法依赖于一种遗传编码的探针，其中包含一个感应肽和一个环状排列的绿色荧光蛋白，提供了一种比率读数。在细菌和哺乳动物细胞内，我们发现细胞质成分与阳离子和疏水探针强烈相互作用，但与中性亲水探针不相互作用，后者保持惰性。细胞质与带高负电荷的亲水探针强烈相互作用，但 HEK293T 细胞质则没有。这些缔合相互作用可被 ATP 耗竭调节。因此，细胞中非特异性缔合相互作用的模式取决于条件和物种。
