A Prototype for Modular Cell Engineering

Abstract

When aiming to produce a target chemical at high yield, titer, and productivity, various combinations of genetic parts available to build the target pathway can generate a large number of strains for characterization. This engineering approach will become increasingly laborious and expensive when seeking to develop desirable strains for optimal production of a large space of biochemicals due to extensive screening. Our recent theoretical development of modular cell (MODCELL) design principles can offer a promising solution for rapid generation of optimal strains by coupling a modular cell and exchangeable production modules in a plug-and-play fashion. In this study, we experimentally validated some designed properties of MODCELL by demonstrating: i) a modular (chassis) cell is required to couple with a production module, a heterologous ethanol pathway, as a testbed, ii) degree of coupling between the modular cell and production modules can be modulated to enhance growth and product synthesis, iii) a modular cell can be used as a host to select an optimal pyruvate decarboxylase (PDC) of the ethanol production module and to help identify a hypothetical PDC protein, and iv) adaptive laboratory evolution based on growth selection of the modular cell can enhance growth and product synthesis rates. We envision that the MODCELL design provides a powerful prototype for modular cell engineering to rapidly create optimal strains for synthesis of a large space of biochemicals.

Proteome reallocation enables the selective de novo biosynthesis of non-linear, branched-chain acetate esters

“…S4), suggesting the possible inhibition of fermentation by the cofactor imbalance under the oxygen limited conditions. Recent studies suggest that modular cell design principle can harness such electron redundancy for multi-objective strain design by coupling production pathways with cell growth (Garcia and Trinh, 2019;Wilbanks et al, 2018). Therefore, further optimization should include process engineering and chassis cell design for improved production of branched-chain acetate esters.…”

Controlling Selectivity of Modular Microbial Biosynthesis of Butyryl-CoA-Derived Designer Esters

“…Except for EcJWBA15, TCS083 △fadE (DE3) (Layton and Trinh, 2014) was used as a host strain. For EcJWBA15, TCS095 (DE3) (Wilbanks et al, 2018) was used as a host strain. A set of duet vectors including pACYCDuet-1, pETDuet-1, and pRSFDuet-1 was used as plasmid backbones for constructing a library of BA, EB, and BB production modules.…”

“…Except for EcJWBA15, TCS083 △fadE (DE3) (Layton and Trinh, 2014) was used as a host strain. For EcJWBA15, TCS095 (DE3) (Wilbanks et al, 2018) was used as a host strain. Key strains are in bold.…”

“…For instance, the endogenous bifunctional aldehyde/alcohol dehydrogenase adhE favors the formation of ethanol over butanol (Atsumi et al, 2008). To demonstrate the optimization of BA production, we replaced TCS083 ΔfadE (DE3) with TCS095 (DE3) that is an adhE-deficient chassis cell (Wilbanks et al, 2018).…”

“…To enable a systematic and rapid generation of microbial biocatalysts to produce various esters, a modular cell engineering framework has recently been developed (Garcia and Trinh, 2019a;Garcia and Trinh, 2019b;Garcia and Trinh, 2020;Trinh et al, 2015;Wilbanks et al, 2018).…”