Genomic Science Program
U.S. Department of Energy | Office of Science | Biological and Environmental Research Program

Encapsulin Nanocompartment Systems in Rhodococcus opacus for Compartmentalized Biosynthesis Applications

Authors:

Daniel J. Wackelin* (wackelin1@llnl.gov), Tiffany M. Halvorsen, Mimi C. Yung

Institutions:

Biosciences and Biotechnology Division, Lawrence Livermore National Laboratory

Goals

This project is focused on understanding how encapsulin nanocompartment systems can be used to enhance the biosynthesis of next-generation biomaterials in Rhodococcus species. The project seeks (1) to probe the mechanistic basis for how these compartments are regulated, biosynthesized, and maintained and (2) to engineer these systems to achieve new biosynthetic functions.

Abstract

With recent innovations in synthetic biology, engineered microbes now have the potential to produce a wide variety of bioproducts from renewable sources (e.g., biomass) to support the U.S. bioeconomy. Biosynthetic pathways leading to these products are often hindered by poor reaction efficiencies and toxicity, however, resulting in low yields and impure products. Compartmentalization of these pathways has the potential to overcome these challenges through co-localization, concentration, and sequestration. The goal of this early career research is to identify mechanisms for engineering compartmentalized biosynthesis in the emerging model bioproduction bacterium, Rhodococcus opacus PD630, using its native encapsulin nanocompartment system (herein called encapsulins). Toward this goal, we have integrated a fluorescent reporter into the R. opacus genome under the control of the native encapsulin promoter. With this, we are investigating the regulation, biosynthesis, and maintenance of the native encapsulin system using growth assays and transposon mutagenesis.

These studies will uncover the native pathways that govern encapsulin synthesis with the goal of ultimately harnessing these pathways for improved recombinant encapsulin formation and yield. We are also developing novel, high-throughput methods for the engineering of encapsulins to systematically identify optimal insertion locations and sequences, as well as easily modulate their properties. This will enable us to quickly tailor the properties of encapsulins to process requirements, greatly enhancing the utility of this system. As a case study, the R. opacus encapsulin system will be redirected to support and control the biosynthesis of cadmium sulfide nanoparticles, semi-conducting materials used in optical and electronic applications. Ultimately, this work will establish encapsulin compartmentalization systems as a means of improving yields and enabling new biosynthetic routes toward next generation bioproducts and biomaterials, in support of the DOE’s mission to build a strong bioeconomy.

Funding Information

Work at Lawrence Livermore National Laboratory (LLNL) is performed under the auspices of the U.S. DOE at LLNL under Contract DE-AC52-07NA27344 (LLNL-ABS-860402). This program is supported by the U.S. DOE, Office of Science, BER program, GSP, under FWP SCW1770 (Early Career Award).