Giant vesicles functionally expressing membrane receptors for an insect pheromone

Abstract

To date, biochemical approaches to membrane receptors have been limited to the following methods: knockout or overexpression of membrane receptors by gene introduction and genome engineering or extraction of membrane receptor-surfactant complexes from innate cells and their introduction into model biomembranes. Here, we describe the development of a third method involving gene expression using cell-free in situ protein synthesis inside model biomembrane capsules. We verified this method by synthesizing olfactory receptors from the silkmoth Bombyx mori inside giant vesicles and found that they were excited in the presence of their ligand the Bombyx mori sex pheromone.

Synthetic Plants

“…The olfactory receptor complex from the silkmoth Bombyx mori has been also synthesised inside GVs prepared by the droplet transfer method (Hamada et al 2014). Interestingly, the receptor was able to respond to the presence of the ligand bombykol, suggesting that future developments of synthetic cell systems capable of communicating via chemical signals and surface receptors Rampioni et al 2014) are indeed possible.…”

“…The recently introduced 'droplet-transfer' method (Pautot et al 2003a,b), discussed by Walde et al (2010) and Stano et al (2011), is probably the most important recent development in the field, and most of the recent work is based on this method (Noireaux and Libchaber 2004;Pontani et al 2009;Nishimura et al 2012;Carrara et al 2012;Maeda et al 2012;Hamada et al 2014).…”

Advances in the Identification and Characterization of Olfactory Receptors in Insects

“…An insect cell-free expression system followed by integration into giant vesicles was used by a Japanese group. 121 Their proof-of-concept study utilized a sex-pheromone receptor of B. mori to demonstrate that a transmembrane receptor could be functionally produced in a cell-free manner. Similarly, expression of insect ORs has been achieved using the wheat-germ cell-free expression system, as well as in insect cells using the baculovirus expression system to produce relatively large quantities of membrane-bound insect ORs which can then be purified, for subsequent structural studies.…”

Communicating artificial cells

“…One solution would be to incorporate membrane protein sensors and transporters within the artificial cells. Advances in cellfree expression have led to the reconstitution of the secYEG translocon, ATP synthase [59], multidrug transporter [60], pheromone sensor [61], and ion channels [62] in phospholipid vesicles. The surface of vesicles can also be decorated with antibodies in order to sense specific target cells.…”