New Method for Measuring the Anchoring Energy of Strongly-Bound Membrane-Associated Proteins [Method for measuring the anchoring energy of strongly-bound membrane-associated proteins].

Journal Article · Mon Feb 01 00:00:00 EST 2016 · Biophysical Journal

DOI:https://doi.org/10.1016/j.bpj.2015.11.3091· OSTI ID:1340234

Kent, Michael S. ^[1]; La Bauve, Elisa ^[1]; Vernon, Briana C. ^[2]; Ye, Dongmei ^[3]; Rogers, David M. ^[4]; Siegrist, Cathryn M. ^[2]; Carson, Bryan ^[1]; Rempe, Susan L. ^[3]; Zheng, Aihua ^[5]; Kielian, Margaret ^[5]; Schreve, Andrew P. ^[6]

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1. Sandia National Lab. (SNL-NM), Albuquerque, NM (United States). Bioenergy and Defense Technology
2. Sandia National Lab. (SNL-NM), Albuquerque, NM (United States)
3. Sandia National Lab. (SNL-NM), Albuquerque, NM (United States). Nanobiology
4. Univ. of South Florida, Tampa, FL (United States). Chemistry
5. Albert Einstein College of Medicine, Bronx, NY (United States). Dept. of Cell Biology
6. Univ. of New Mexico, Albuquerque, NM (United States). Center for Biomedical Engineering and Dept. of Chemical and Biological Engineering

Here, we describe a new method to measure the activation energy required to remove a strongly-bound membrane-associated protein from a lipid membrane (anchoring energy). It is based on measuring the rate of release of a liposome-bound protein during centrifugation on a sucrose gradient as a function of time and temperature. The method was used to determine anchoring energy for the soluble dengue virus envelope protein (sE) strongly bound to 80:20 POPC:POPG liposomes at pH 5.5. We also measured the binding energy of sE at the same pH for the initial, predominantly reversible, phase of binding to a 70:30 PC:PG lipid bilayer. The anchoring energy (37 +/- 1.7 kcal/mol, 20% PG) was found to be much larger than the binding energy (7.8 +/- 0.3 kcal/mol for 30% PG, or est. 7.0 kcal/mol for 20% PG). This is consistent with data showing that free sE is a monomer at pH 5.5, but assembles into trimers after associating with membranes. But, trimerization alone is insufficient to account for the observed difference in energies, and we conclude that some energy dissipation occurs during the release process. This new method to determine anchoring energy should be useful to understand the complex interactions of integral monotopic proteins and strongly-bound peripheral membrane proteins with lipid membranes.

• Research Organization: Sandia National Lab. (SNL-NM), Albuquerque, NM (United States)
• Sponsoring Organization: USDOE National Nuclear Security Administration (NNSA)
• Grant/Contract Number: AC04-94AL85000
• OSTI ID: 1340234
• Report Number(s): SAND2016-2840J; PII: S0006349515042745
• Journal Information: Biophysical Journal, Vol. 110, Issue S1; ISSN 0006-3495
• Publisher: Elsevier
• Country of Publication: United States
• Language: English

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59 BASIC BIOLOGICAL SCIENCES