Examining roles of varying synthesis and assembly conditions on the incorporation of complex multi-lipid extracts into membrane scaffold protein nanodiscs
- 1Department of Chemistry
- 2University of Michigan
- 3Department of Internal Medicine
Abstract
Nanodiscs are tunable and widely used membrane mimetics that can allow for the in vitro study of membrane proteins in a native-like lipid bilayer environment. The lipids incorporated into nanodiscs can be modulated to represent specific biophysical environments, but many nanodisc applications rely on simplified lipid compositions that inadequately represent native membrane complexity. Natural lipid extracts offer more physiological relevance; however, recent work has shown that (1) lipid composition of a nanodisc does not stoichiometrically reflect the initial mixture and that (2) assembly conditions such as temperature and belt size may influence lipid incorporation. Here, we add to this body of work by implementing an untargeted LC-MS/MS lipidomics workflow to profile the incorporation of multiple, combined natural lipid extracts into membrane scaffold protein nanodiscs, examining not only assembly temperature and belt size, but also the effects of detergent and synthetic lipid additives on lipid incorporation. Species level lipidomic analysis, including trends in acyl chain length and degree of unsaturation were evaluated. We observe that assembly temperatures have minimal impact on lipid incorporation, whereas belt size effects vary across extracts, as smaller scaffolds likely enrich lipids that promote a more tightly packed membrane. Furthermore, we found that lipid solubilization and incorporation into nanodiscs is broadly preserved regardless of detergent identity. Conversely, synthetic lipid additives influence incorporated lipid class composition more strongly than detergent, temperature, or MSP belt size, with unsaturated additives favoring glycerolipid enrichment. Taken together, this data demonstrates the interplay of nanodisc assembly conditions in shaping the incorporated lipid environment.
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BibTeX
@misc{sarcinella2026examining,
title = {{Examining roles of varying synthesis and assembly conditions on the incorporation of complex multi-lipid extracts into membrane scaffold protein nanodiscs}},
author = {Marina C. Sarcinella and Anusha Vajrala and Samantha A. Edgcombe and Joshua D. Jones and Brady G. Anderson and Charles R. Evans and Brandon T. Ruotolo and Robert T. Kennedy and Ryan C. Bailey},
howpublished = {ChemRxiv},
year = {2026},
month = sep,
doi = {10.26434/chemrxiv.15009419/v1},
url = {https://chemrxiv.org/doi/full/10.26434/chemrxiv.15009419/v1},
note = {CC BY},
}
RIS
TY - UNPB
TI - Examining roles of varying synthesis and assembly conditions on the incorporation of complex multi-lipid extracts into membrane scaffold protein nanodiscs
AU - Marina C. Sarcinella
AU - Anusha Vajrala
AU - Samantha A. Edgcombe
AU - Joshua D. Jones
AU - Brady G. Anderson
AU - Charles R. Evans
AU - Brandon T. Ruotolo
AU - Robert T. Kennedy
AU - Ryan C. Bailey
PY - 2026
DA - 2026/09/24
PB - ChemRxiv
DO - 10.26434/chemrxiv.15009419/v1
UR - https://chemrxiv.org/doi/full/10.26434/chemrxiv.15009419/v1
AB - Nanodiscs are tunable and widely used membrane mimetics that can allow for the in vitro study of membrane proteins in a native-like lipid bilayer environment. The lipids incorporated into nanodiscs can be modulated to represent specific biophysical environments, but many nanodisc applications rely on simplified lipid compositions that inadequately represent native membrane complexity. Natural lipid extracts offer more physiological relevance; however, recent work has shown that (1) lipid composition of a nanodisc does not stoichiometrically reflect the initial mixture and that (2) assembly conditions such as temperature and belt size may influence lipid incorporation. Here, we add to this body of work by implementing an untargeted LC-MS/MS lipidomics workflow to profile the incorporation of multiple, combined natural lipid extracts into membrane scaffold protein nanodiscs, examining not only assembly temperature and belt size, but also the effects of detergent and synthetic lipid additives on lipid incorporation. Species level lipidomic analysis, including trends in acyl chain length and degree of unsaturation were evaluated. We observe that assembly temperatures have minimal impact on lipid incorporation, whereas belt size effects vary across extracts, as smaller scaffolds likely enrich lipids that promote a more tightly packed membrane. Furthermore, we found that lipid solubilization and incorporation into nanodiscs is broadly preserved regardless of detergent identity. Conversely, synthetic lipid additives influence incorporated lipid class composition more strongly than detergent, temperature, or MSP belt size, with unsaturated additives favoring glycerolipid enrichment. Taken together, this data demonstrates the interplay of nanodisc assembly conditions in shaping the incorporated lipid environment.
ER -