Cell Fuelling and Metabolic Energy Conservation in Synthetic Cells

Abstract

We are aiming for a blue print for synthesizing (moderately complex) subcellular systems from molecular components and ultimately for constructing life. However, without comprehensive instructions and design principles, we rely on simple reaction routes to operate the essential functions of life. The first forms of synthetic life will not make every building block for polymers de novo according to complex pathways, rather they will be fed with amino acids, fatty acids and nucleotides. Controlled energy supply is crucial for any synthetic cell, no matter how complex. Herein, we describe the simplest pathways for the efficient generation of ATP and electrochemical ion gradients. We have estimated the demand for ATP by polymer synthesis and maintenance processes in small cell‐like systems, and we describe circuits to control the need for ATP. We also present fluorescence‐based sensors for pH, ionic strength, excluded volume, ATP/ADP, and viscosity, which allow the major physicochemical conditions inside cells to be monitored and tuned.

Experimental tests of functional molecular regeneration via a standard framework for coordinating synthetic cell building

“…For example, on a quantitative volumetric basis, we estimate that PURE is only capable of remaking ~1% of the peptide bonds needed to instantiate PURE itself; expression capacity will need to be increased ~87-fold to enable sustained self-reproduction (Materials and Methods). However, such quantitative challenges can likely be addressed via exogenous supply of resources directly from the environment, via booting up of a regenerative and auto-sustaining metabolism, or by addition of enzymes and mechanisms that increase the efficiency of protein expression by reducing the number of truncated peptides produced and decreasing the rate of ribosomal stalling [17,[21][22][23][24][25]. Whether PURE can satisfy the qualitative criteria for self-reproduction is less clear.…”

Multistimuli Sensing Adhesion Unit for the Self‐Positioning of Minimal Synthetic Cells

“…[ 2 ] Achieving analogous capabilities in synthetic cells, which are bottom‐up assembled systems from modular molecular building blocks with life‐like features, would be an important step forward in their autonomy. [ 3,4 ] This bottom‐up approach in synthetic biology builds on the idea of producing minimal living systems through the modular synthesis and integration [ 5,6 ] of well‐characterized units and modules capable of diverse functions (energy conversion, [ 7 ] metabolism, [ 8 ] division, [ 9 ] growth, [ 10 ] etc.). In this context, an adhesion module that responds to multiple environmental inputs would allow minimal synthetic cells to sense and position themselves in the right place to carry out their functions and would represent a key feature for potential applications in biotechnology like drug delivery, [ 11 ] nanoreactors, [ 12 ] biosensing, [ 13 ] and bioremediation.…”

Physicochemical considerations for bottom-up synthetic biology

“…Metabolic energy generation in living cells is complex and requires numerous enzymes and cofactors [35]. Nature offers alternative mechanisms to conserve metabolic energy through simple metabolic conversions (deamination of amino acids, oxidation of carboxylic acids) or the use of light, which has been recently reviewed [36]. Sustainable synthesis of ATP from the breakdown of arginine in a vesicle system and controlled import and export of reaction products has been demonstrated [37] (see Box I), yet more ways of generating metabolic energy should be explored, e.g.…”