E and M SARS‐CoV‐2 membrane protein expression and enrichment with plant lipid droplets
Résumé
Plants are gaining traction as a cost-effective and scalable platform for producing recombinant proteins. However, expressing integral membrane proteins in plants is challenging due to their hydrophobic nature. In our study, we used transient and stable expression systems in Nicotiana benthamiana and Camelina sativa respectively to express SARS-CoV-2 E and M integral proteins, and target them to lipid droplets (LDs). LDs offer an ideal environment for folding hydrophobic proteins and aid in their purification through flotation. We tested various protein fusions with different linkers and tags and used three dimensional structure predictions to assess their effects. E and M mostly localized in the ER in N. benthamiana leaves but E could be targeted to LDs in oil accumulating tobacco when fused with oleosin, a LD integral protein. In Camelina sativa seeds, E and M were however found associated with purified LDs. By enhancing the accumulation of E and M within LDs through oleosin, we enriched these proteins in the purified floating fraction. This strategy provides an alternative approach for efficiently producing and purifying hydrophobic pharmaceuticals and vaccines using plant systems.
Mots clés
Funding information Merck KGaA
frankfurter straße 250 64293 darmstadt Saclay Plant Sciences (SPS)
Grant/Award Number: ANR17-EUR-007 camelina
colabfold
Goldenbraid
lipid droplet
SARS-CoV-2
tobacco ACE-2
angiotensin-converting enzyme-2 E
envelope ER
endoplasmic reticulum GFP
green fluorescent protein LD
lipid droplet LDAP
lipid droplet associated protein M
membrane MDH
monodansylpentane mRFP1
monomeric red fluorescent protein 1 N
nucleocapsid S
spike TAG
triacylglycerol VLP
virus like particles
frankfurter straße 250 64293 darmstadt
Saclay Plant Sciences (SPS)
angiotensin-converting enzyme-2
E
envelope
ER
endoplasmic reticulum
GFP
green fluorescent protein
LD
LDAP
lipid droplet associated protein
M
membrane
MDH
monodansylpentane
mRFP1
monomeric red fluorescent protein 1
N
nucleocapsid
S
spike
TAG
triacylglycerol
VLP
Domaines
Sciences du Vivant [q-bio]
Origine : Fichiers éditeurs autorisés sur une archive ouverte
Licence : CC BY - Paternité
Licence : CC BY - Paternité