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Self-Assembling Metabolon Enables the Cell Free Conversion of Glycerol to 1,3-Propanediol

Xu, Qi and Alahuhta, Markus and Hewitt, Patrick and Sarai, Nicholas S. and Wei, Hui and Hengge, Neal N. and Mittal, Ashutosh and Himmel, Michael E. and Bomble, Yannick J. (2021) Self-Assembling Metabolon Enables the Cell Free Conversion of Glycerol to 1,3-Propanediol. Frontiers in Energy Research, 9 . Art. No. 680313. ISSN 2296-598X. doi:10.3389/fenrg.2021.680313.

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Cell free biocatalysis is showing promise as a replacement or complement to conventional microbial biocatalysts due to the potential for achieving high yields, titers, and productivities. However, there exist several challenges that need to be addressed before its broader industrial adoption is achieved. New paradigms and innovative solutions are needed to overcome these challenges. In this study we demonstrate high levels of glycerol conversion to 1,3-propanediol using a self-assembling metabolic pathway leveraging the arraying strategy (protein scaffolds) used by thermophilic cellulolytic bacteria to assemble their biomass degrading enzymes. These synthetic metabolons were capable of producing 1,3-PDO at a yield more than 95% at lower glycerol concentration and close to 70% at higher concentrations at a higher productivity rate than the equivalent microbial strain. One of the benefits of our approach is the fact that no enzyme purification is required, and that the assembly of the complex is accomplished in vivo before immobilization, while product formation is conducted in vitro. We also report the recovery of enzymatic activity upon fusion enzymes binding to these protein scaffolds, which could have broader applications when assembling arrayed protein complexes.

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Additional Information:© 2021 Xu, Alahuhta, Hewitt, Sarai, Wei, Hengge, Mittal, Himmel and Bomble. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. Received: 14 March 2021; Accepted: 09 June 2021; Published: 15 July 2021. This work was authored by Alliance for Sustainable Energy, LLC, the manager and operator of the National Renewable Energy Laboratory for the U.S. Department of Energy (DOE) under Contract No. DE-AC36-08GO28308. Funding provided by U.S. Department of Energy Office of Energy Efficiency and Renewable Energy Bioenergy Technologies Office under Contract DE-AC36-08GO28308 with the National Renewable Energy Laboratory. The publisher, by accepting the article for publication, acknowledges that the U. S. Government retains a nonexclusive, paid-up, irrevocable, worldwide license to publish or reproduce the published form of this work, or allow others to do so, for U. S. Government purposes. The views expressed in the article do not necessarily represent the views of the U.S. Department of Energy or the United States Government. We would also like to thank Professor Edward Bayer for sharing the original PCR templates of cohesin and dockerin modules. Data Availability Statement: The original contributions presented in the study are included in the article/Supplementary Material, further inquiries can be directed to the corresponding author. Author Contributions: QX and YB designed the study, QX built the constructs and tested 1,3 PDO production in vivo. MA, PH, and NS conducted protein purification and in vitro assays. HW and NH conducted transformations and protein expression. AM conducted substrate and product analysis. MH helped analyze the results and reviewed the manuscript. YB supervised the research and provided the funding. QX and YB wrote the manuscript and all authors reviewed and approved of the manuscript. The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
Funding AgencyGrant Number
Department of Energy (DOE)DE-AC36-08GO28308
Subject Keywords:glycerol, biocatalysis, metabolic pathway, biochemical, cellulose, cellulosome assembly
Record Number:CaltechAUTHORS:20210915-150220508
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Official Citation:Xu Q, Alahuhta M, Hewitt P, Sarai NS, Wei H, Hengge NN, Mittal A, Himmel ME and Bomble YJ (2021) Self-Assembling Metabolon Enables the Cell Free Conversion of Glycerol to 1,3-Propanediol. Front. Energy Res. 9:680313. doi: 10.3389/fenrg.2021.680313
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:110908
Deposited By: Tony Diaz
Deposited On:15 Sep 2021 18:01
Last Modified:15 Sep 2021 18:01

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