May 03, 2024
Database-driven protein solubilization
- Caroline Brown1,2,3,
- Snehasish Ghosh1,2,3,
- Kallol Gupta1,2,3
- 1Nanobiology Institute, Yale University, West Haven, CT, USA;
- 2Department of Cell Biology, Yale University School of Medicine, New Haven, CT, USA;
- 3Aligning Science Across Parkinson’s (ASAP) Collaborative Research Network, Chevy Chase, MD, 20815
Protocol Citation: Caroline Brown, Snehasish Ghosh, Kallol Gupta 2024. Database-driven protein solubilization. protocols.io https://dx.doi.org/10.17504/protocols.io.36wgqn735gk5/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: May 03, 2024
Last Modified: May 03, 2024
Protocol Integer ID: 99218
Abstract
This is a protocol for using the extraction database found at www.polymerscreen.yale.edu for optimal extraction of membrane proteins into native nanodiscs.
Choose your protein of interest.
Visit www.polymerscreen.yale.edu to search for your protein in the extraction database. This will provide you with the polymer to use for optimal protein extraction.
As an example, below are representative organellar markers and the optimal polymer for extraction:
TMEM192 --> AASTY650
TGN46 --> ChloroSMA60
OMP25 --> SMA200
VAPA --> AASTY650
KRas --> ChloroSMA80
Once the optimal polymer has been determined, express the protein of interest, or harvest cells for endogenous solubilization.
Resuspend cellular membranes in the chosen polymer and incubate at 4 °C with rotation for 02:00:00 hours.
2h
Ultracentrifuge samples ay 200,000xg for 01:00:00 hours to remove insoluble material.
1h