A lipid-based parallel processor for chemical signals

Cazimoglu, Idil; Mj, Booth; Bayley, Hagan

doi:10.1101/2021.05.05.442835

Cited by 3 publications

(3 citation statements)

References 41 publications

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“…Generation of αHL and αHL-4H Pores: Recombinant αHL was expressed in E. coli and purified as previously described. [22] In brief, αHL-D8H6 was expressed in E. coli BL21(DE3)pLysS cells (Agilent) and purified on Ni-NTA agarose, followed by size exclusion chromatography. Monomers were aliquoted and stored at −80 °C until use.…”

Section: Methodsmentioning

confidence: 99%

“…To demonstrate the formation of a functional bilayer interface between the two building blocks, we encapsulated the membrane pore-forming protein α-hemolysin (αHL) within the 3D-printed droplets. This pore-forming protein is known to self-assemble into lipid bilayers and allow the transfer of small molecules [19][20][21][22] and transmission of ionic currents [8,11,16] through lipid-bilayer-interconnected aqueous compartments. We then investigated the electrical properties of the assembled synthetic tissue (Note S1, Supporting Information).…”

Section: Connecting 3d-printed Building Blocksmentioning

confidence: 99%

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Modular Synthetic Tissues from 3D‐Printed Building Blocks

Alcinesio

Cazimoglu

Kimmerly

et al. 2021

Adv Funct Materials

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Biology employs modular organization at every scale: molecular building blocks make up living cells, specialized cells organize within tissues, and collections of tissues constitute organs. 3D-printed networks of picoliter-sized aqueous compartments interconnected by lipid bilayers form a powerful platform for building precisely patterned synthetic tissues. However, this technology has been limited to millimeter-sized networks, with slow fabrication times and lacking flexible design. Here, the authors apply modular design to construct modular synthetic tissues by assembling a wide range of 3D-printed building blocks. They use dedicated modules for storing and releasing reagents, performing logic operations, responding to magnetic fields, and encapsulating living cells. They build centimeter-sized synthetic tissues able to transmit electrical signals through thousands of interconnected compartments. They assemble hybrid tissues composed of both synthetic modules and modules containing living cells. Lastly, by incorporating mutant protein nanopores within the building blocks, they assemble modular synthetic tissues with electrical outputs that are modulated by the integration of chemical inputs.

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Section: Methodsmentioning

confidence: 99%

Section: Connecting 3d-printed Building Blocksmentioning

confidence: 99%

Modular Synthetic Tissues from 3D‐Printed Building Blocks

Alcinesio

Cazimoglu

Kimmerly

et al. 2021

Adv Funct Materials

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“…We functionalized the lipid bilayers between the compartments and between the structures and the external environment with the pore-forming membrane protein α-hemolysin (αHL), thereby allowing the selective diffusion of small molecules throughout the synthetic tissue and to or from the external solution (Figure 3a, d). [43] We first encapsulated 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl) amino]-2-deoxy-D-glucose (2-NBDG) within the synthetic tissue's compartments and monitored its release over time. We monitored the release of 2-NBDG using a laser scanning confocal microscope (Leica SP5) focused at the midpoint of the first compartment layer (at the bottom) of the synthetic tissue, to exclude the contribution of the upper layers from our analysis.…”

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confidence: 99%

Functional Multivesicular Structures with Controlled Architecture from 3D‐Printed Droplet Networks

2021

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Vesicles are versatile drug delivery systems and a fundamental chassis from which to build synthetic cells. Multivesicular structures, composed of multiple lipid‐bound compartments, could form systems with complex functionalities and recapitulate collective behaviour found in living tissues. However, control over number, composition, and arrangement of compartments within multivesicular structures has been limited. Here, we generate multivesicular structures composed of hundreds of lipid‐bound compartments arranged in precisely controlled geometries. Through a dewetting process, we transfer 3D‐printed droplet networks from oil to aqueous phase and obtain multivesicular structures with designed, complex architectures stable for weeks. We encapsulate membraneless coacervate sub‐compartments within the lipid‐bound compartments of the 3D‐printed multivesicular structures. Further, we use the multivesicular structures as a platform to build synthetic tissues that interact with the external aqueous environment by releasing or sensing molecules and ions. Finally, we build synthetic tissues that synthesize proteins encoded by encapsulated genes.

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Building programmable multicompartment artificial cells incorporating remotely activated protein channels using microfluidics and acoustic levitation

Jin

Jamieson

Dimitriou

et al. 2022

Preprint

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Intracellular compartments are functional units that support the metabolic processes within living cells, through spatiotemporal regulation of chemical reactions and biological processes. Consequently, as a step forward in the bottom-up creation of artificial cells, building analogous intracellular architectures is essential for the expansion of cell-mimicking functionality. Herein, we report the development of a droplet laboratory platform to engineer customised complex emulsion droplets as a multicompartment artificial cell chassis, using multiphase microfluidics and acoustic levitation. Such levitated constructs provide free-standing, dynamic, definable droplet networks for the encapsulation and organisation of chemical species. Equally, they can be remotely operated with pneumatic, heating, and magnetic elements for post-processing, including the incorporation of membrane proteins; alpha-hemolysin; and large-conductance mechanosensitive channel (MscL) and their activation. The assembly of droplet networks is three-dimensionally patterned with fluidic inputs configurations determining droplet contents and connectivity. Whilst acoustic manipulation can be harnessed to reconfigure the droplet network in situ. In addition, a mechanosensitive channel, MscL, can be repeatedly activated and deactivated in the levitated artificial cell by the application of acoustic and magnetic fields to modulate membrane tension on demand. This offers possibilities beyond one-time chemically mediated activation to provide repeated, non-contact control of membrane protein function. Collectively, this will expand our capability to program and operate increasingly sophisticated artificial cells as life-like materials.

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A lipid-based parallel processor for chemical signals

Cited by 3 publications

References 41 publications

Modular Synthetic Tissues from 3D‐Printed Building Blocks

Modular Synthetic Tissues from 3D‐Printed Building Blocks

Functional Multivesicular Structures with Controlled Architecture from 3D‐Printed Droplet Networks

Building programmable multicompartment artificial cells incorporating remotely activated protein channels using microfluidics and acoustic levitation

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