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Title: Repurposing the Saccharomyces cerevisiae peroxisome for compartmentalizing multi-enzyme pathways

Abstract

The peroxisome of Saccharomyces cerevisiae was targeted for repurposing in order to create a synthetic organelle that provides a generalizable compartment for engineered metabolic pathways. Compartmentalization of enzymes into organelles is a promising strategy for limiting metabolic crosstalk, improving pathway efficiency, and ultimately modifying the chemical environment to be distinct from that of the cytoplasm. We focused on the Saccharomyces cerevisiae peroxisome, as this organelle is not required for viability when grown on conventional media. We identified an enhanced peroxisomal targeting signal type 1 (PTS1) for rapidly importing non-native cargo proteins. Additionally, we performed the first systematic in vivo measurements of nonspecific metabolite permeability across the peroxisomal membrane using a polymer exclusion assay and characterized the size dependency of metabolite trafficking. Finally, we applied these new insights to compartmentalize a two-enzyme pathway in the peroxisome and characterize the expression regimes where compartmentalization leads to improved product titer. This work builds a foundation for using the peroxisome as a synthetic organelle, highlighting both promise and future challenges on the way to realizing this goal.

Authors:
 [1];  [1];  [1];  [1]
  1. Univ. of California, Berkeley, CA (United States)
Publication Date:
Research Org.:
Univ. of California, Berkeley, CA (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Biological and Environmental Research (BER) (SC-23)
OSTI Identifier:
1394729
Report Number(s):
DOE-BERKELEY-0008084
DOE Contract Number:
SC0008084
Resource Type:
Technical Report
Resource Relation:
Related Information: additional data are described in the supplementary information from the Nature Communication paper (DOI: 10.1038/ncomms11152).
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES; 59 BASIC BIOLOGICAL SCIENCES; 10 SYNTHETIC FUELS; synthetic biology; organelle engineering; peroxisome; compartmentalization; metabolic engineering

Citation Formats

DeLoache, William, Russ, Zachary, Samson, Jennifer, and Dueber, John. Repurposing the Saccharomyces cerevisiae peroxisome for compartmentalizing multi-enzyme pathways. United States: N. p., 2017. Web. doi:10.2172/1394729.
DeLoache, William, Russ, Zachary, Samson, Jennifer, & Dueber, John. Repurposing the Saccharomyces cerevisiae peroxisome for compartmentalizing multi-enzyme pathways. United States. doi:10.2172/1394729.
DeLoache, William, Russ, Zachary, Samson, Jennifer, and Dueber, John. Mon . "Repurposing the Saccharomyces cerevisiae peroxisome for compartmentalizing multi-enzyme pathways". United States. doi:10.2172/1394729. https://www.osti.gov/servlets/purl/1394729.
@article{osti_1394729,
title = {Repurposing the Saccharomyces cerevisiae peroxisome for compartmentalizing multi-enzyme pathways},
author = {DeLoache, William and Russ, Zachary and Samson, Jennifer and Dueber, John},
abstractNote = {The peroxisome of Saccharomyces cerevisiae was targeted for repurposing in order to create a synthetic organelle that provides a generalizable compartment for engineered metabolic pathways. Compartmentalization of enzymes into organelles is a promising strategy for limiting metabolic crosstalk, improving pathway efficiency, and ultimately modifying the chemical environment to be distinct from that of the cytoplasm. We focused on the Saccharomyces cerevisiae peroxisome, as this organelle is not required for viability when grown on conventional media. We identified an enhanced peroxisomal targeting signal type 1 (PTS1) for rapidly importing non-native cargo proteins. Additionally, we performed the first systematic in vivo measurements of nonspecific metabolite permeability across the peroxisomal membrane using a polymer exclusion assay and characterized the size dependency of metabolite trafficking. Finally, we applied these new insights to compartmentalize a two-enzyme pathway in the peroxisome and characterize the expression regimes where compartmentalization leads to improved product titer. This work builds a foundation for using the peroxisome as a synthetic organelle, highlighting both promise and future challenges on the way to realizing this goal.},
doi = {10.2172/1394729},
journal = {},
number = ,
volume = ,
place = {United States},
year = {Mon Sep 25 00:00:00 EDT 2017},
month = {Mon Sep 25 00:00:00 EDT 2017}
}

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