Lipid and Protein Transfer between Nanolipoprotein Particles and Supported Lipid Bilayers

Abstract

A nanolipoprotein particle (NLP) is a lipid bilayer disc stabilized by two amphipathic "scaffold" apolipoproteins. It has been most notably utilized as a tool for solubilizing a variety of membrane proteins while preserving structural and functional properties. Transfer of functional proteins from NLPs into model membrane systems such as supported lipid bilayers (SLBs) would enable new opportunities for example: two-dimensional protein crystallization and studies on protein-protein interactions. This work used fluorescence microscopy (FM) and atomic force microscopy (AFM) to investigate the interaction between NLPs and SLBs. When incubated with SLBs, NLPs were found to spontaneously deliver lipid and protein cargo. The impact of membrane composition on lipid exchange was explored, revealing a positive correlation between the magnitude of lipid transfer and concentration of defects in the target SLB. Incorporation of lipids capable of binding specifically to polyhistidine tags encoded into the apolipoproteins also boosted transfer of NLP cargo. Optimal conditions for lipid and protein delivery from NLPs to SLBs are proposed based on interaction mechanisms.

Transfer mechanism of cell-free synthesized membrane proteins into mammalian cells

“…With CHO cells, most of the transferred MPs are localized in the cytoplasm. It has been shown that the composition of supported lipid bilayers affects the transfer of MPs from NDs (Dang et al, 2019). Different membrane compositions of the analyzed cell lines might therefore contribute to the observed preference in MP localization.…”

“…According to the fusion pore model for the transfer of cargo from NDs to bicelles, the NDs are suggested to align in a planar orientation on top of the target bilayer (Lai et al, 2015). The MPs may then spontaneously transfer between nanoparticle and cell membranes (Dang et al, 2019). Two contact sites of the MP/ND particles with the cell membrane are possible.…”

“…A significant advantage for MP transfer was the implementation of NDs as vectors (Denisov and Sligar, 2017). If NDs are mixed with other lipid bilayers such as micelles, supported bilayers, or even lipid cubic phases, lipids and inserted MPs can be transferred (Lai et al, 2015;Henrich et al, 2017a;Nikolaev et al, 2017;Dang et al, 2019). Furthermore, NDs have been used to transfer a variety of other biomolecules such as RNA, optical probes, or drugs into living cells (Murakami, 2012;Carney et al, 2015;He et al, 2020).…”

Nanolipoprotein-Mediated Her2 Protein Transfection Induces Malignant Transformation in Human Breast Acinar Cultures

“…Furthermore, NLP-mediated protein delivery is believed to be a spontaneous exchange of proteins between NLPs and plasma membrane, thus minimizing the potential off-target effects of the delivery system. 18 Given the facile synthesis of NLPs and the simplicity of this protein transfection system, we sought to determine if this system could be used to rapidly engineer new oncoproteindriven cancers. Half of all drugs that have been approved to treat cancer in the past decade target cell surface receptors.…”

“…In comparison to existing protein delivery systems that risk denaturing the proteins they carry, either by harsh packaging or endosomal lytic processes, − NLPs maintain correct folding of mammalian cell surface receptors. ,, NLPs, in contrast to other protein delivery systems, do not require cell membrane disruption ,− and diffuse readily due to their small size relative to silica or poly­(lactic- co -glycolic acid) (PLGA) nanoparticles, ,,− potentially resulting in a more efficient transfer of proteins into cells. Furthermore, NLP-mediated protein delivery is believed to be a spontaneous exchange of proteins between NLPs and plasma membrane, thus minimizing the potential off-target effects of the delivery system …”