Endoproteolysis of Oligopeptide-Based Coacervates for Enzymatic Modeling

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Endoproteolysis of Oligopeptide-Based Coacervates for Enzymatic Modeling
Title:
Endoproteolysis of Oligopeptide-Based Coacervates for Enzymatic Modeling
Journal Title:
ACS Nano
Publication Date:
14 August 2023
Citation:
Jin, Z., Ling, C., Yim, W., Chang, Y.-C., He, T., Li, K., Zhou, J., Cheng, Y., Li, Y., Yeung, J., Wang, R., Fajtová, P., Amer, L., Mattoussi, H., O’Donoghue, A. J., & Jokerst, J. V. (2023). Endoproteolysis of Oligopeptide-Based Coacervates for Enzymatic Modeling. ACS Nano, 17(17), 16980–16992. https://doi.org/10.1021/acsnano.3c04259
Abstract:
Better insights into the fate of membraneless organelles could strengthen the understanding of the transition from prebiotic components to multicellular organisms. Compartmentalized enzyme reactions in a synthetic coacervate have been investigated, yet there remains a gap in understanding the enzyme interactions with coacervate as a substrate hub. Here, we study how the molecularly crowded nature of the coacervate affects the interactions of the embedded substrate with a protease. We design oligopeptide-based coacervates that comprise an anionic Asp-peptide (D10) and a cationic Arg-peptide (R5R5) with a proteolytic cleavage site. The coacervates dissolve in the presence of the main protease (Mpro) implicated in the coronavirus lifecycle. We capitalize on the condensed structure, introduce a self-quenching mechanism, and model the enzyme kinetics by using Cy5.5-labeled peptides. The determined specificity constant (kcat/KM) is 5817 M–1 s–1 and is similar to that of the free substrate. We further show that the enzyme kinetics depend on the type and quantity of dye incorporated into the coacervates. Our work presents a simple design for enzyme-responsive coacervates and provides insights into the interactions between the enzyme and coacervates as a whole.
License type:
Publisher Copyright
Funding Info:
The authors thank the National Institutes of Health (R01 DE031114; R21 AG065776-S1; R21 AI157957) for financial support. We also thank NSF under grant #1845683. This work was also supported by National Science Foundation (DMR-2011924) via equipment in the UC San Diego Materials Research Science and Engineering Center (UCSD MRSEC). The electron microscopy work was performed in part at the San Diego Nanotechnology Infrastructure (SDNI) of University of California San Diego, a member of the National Nanotechnology Coordinated Infrastructure (NNCI), which is supported by the National Science Foundation (Grant ECCS-1542148).
Description:
This document is the Accepted Manuscript version of a Published Work that appeared in final form in ACS Nano, copyright © American Chemical Society after peer review and technical editing by the publisher. To access the final edited and published work see doi.org/10.1021/acsnano.3c04259
ISSN:
1936-086X
1936-0851
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