Abstract
The thermodynamic stability of proteins is typically measured at high denaturant concentrations and then extrapolated back to zero denaturant conditions to obtain unfolding free energies under native conditions. For membrane proteins, the extrapolations are fraught with considerable uncertainty as the denaturants may have complex effects on the membrane or micellar structure. We therefore sought to measure stability under native conditions, using a method that does not perturb the properties of the membrane or membrane mimetics. We use a technique called steric trapping to measure the thermodynamic stability of bacteriorhodopsin in bicelles and micelles. We find that bacteriorhodopsin has a high thermodynamic stability, with an unfolding free energy of ~11 kcal/mol in dimyristoyl phosphatidylcholine bicelles. Nevertheless, the stability is much lower than predicted by extrapolation of measurements made at high denaturant concentrations. We investigated the discrepancy and found that unfolding free energy is not linear with denaturant concentration. Apparently, long extrapolations of helical membrane protein unfolding free energies must be treated with caution. Steric trapping, however, provides a method for making these measurements.
| Original language | English |
|---|---|
| Pages (from-to) | 219-224 |
| Number of pages | 6 |
| Journal | Proceedings of the National Academy of Sciences of the United States of America |
| Volume | 111 |
| Issue number | 1 |
| DOIs | |
| Publication status | Published - 2014 |
| Externally published | Yes |
Fingerprint
Keywords
- Membrane protein folding
- Steric trap
ASJC Scopus subject areas
- General
Cite this
Measuring membrane protein stability under native conditions. / Chang, Yu Chu; Bowie, James U.
In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 111, No. 1, 2014, p. 219-224.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Measuring membrane protein stability under native conditions
AU - Chang, Yu Chu
AU - Bowie, James U.
PY - 2014
Y1 - 2014
N2 - The thermodynamic stability of proteins is typically measured at high denaturant concentrations and then extrapolated back to zero denaturant conditions to obtain unfolding free energies under native conditions. For membrane proteins, the extrapolations are fraught with considerable uncertainty as the denaturants may have complex effects on the membrane or micellar structure. We therefore sought to measure stability under native conditions, using a method that does not perturb the properties of the membrane or membrane mimetics. We use a technique called steric trapping to measure the thermodynamic stability of bacteriorhodopsin in bicelles and micelles. We find that bacteriorhodopsin has a high thermodynamic stability, with an unfolding free energy of ~11 kcal/mol in dimyristoyl phosphatidylcholine bicelles. Nevertheless, the stability is much lower than predicted by extrapolation of measurements made at high denaturant concentrations. We investigated the discrepancy and found that unfolding free energy is not linear with denaturant concentration. Apparently, long extrapolations of helical membrane protein unfolding free energies must be treated with caution. Steric trapping, however, provides a method for making these measurements.
AB - The thermodynamic stability of proteins is typically measured at high denaturant concentrations and then extrapolated back to zero denaturant conditions to obtain unfolding free energies under native conditions. For membrane proteins, the extrapolations are fraught with considerable uncertainty as the denaturants may have complex effects on the membrane or micellar structure. We therefore sought to measure stability under native conditions, using a method that does not perturb the properties of the membrane or membrane mimetics. We use a technique called steric trapping to measure the thermodynamic stability of bacteriorhodopsin in bicelles and micelles. We find that bacteriorhodopsin has a high thermodynamic stability, with an unfolding free energy of ~11 kcal/mol in dimyristoyl phosphatidylcholine bicelles. Nevertheless, the stability is much lower than predicted by extrapolation of measurements made at high denaturant concentrations. We investigated the discrepancy and found that unfolding free energy is not linear with denaturant concentration. Apparently, long extrapolations of helical membrane protein unfolding free energies must be treated with caution. Steric trapping, however, provides a method for making these measurements.
KW - Membrane protein folding
KW - Steric trap
UR - http://www.scopus.com/inward/record.url?scp=84891917719&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84891917719&partnerID=8YFLogxK
U2 - 10.1073/pnas.1318576111
DO - 10.1073/pnas.1318576111
M3 - Article
C2 - 24367094
AN - SCOPUS:84891917719
VL - 111
SP - 219
EP - 224
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
SN - 0027-8424
IS - 1
ER -