Escherichia coli OmpA retains a folded structure in the presence of sodium dodecyl sulfate due to a high kinetic barrier to unfolding.
Escherichia coli OmpA can be solubilized by sodium dodecyl sulfate (SDS) in its folded structure, and it unfolds upon heating. Although the heat-denatured OmpA remains unfolded after lowering the temperature, the addition of a non-ionic surfactant, octyl glucoside results in refolding of unfolded OmpA. In the present study, we investigated the refolding kinetics of OmpA in a mixed surfactant system of SDS and octyl glucoside using far- and near-UV circular dichroism and fluorescence spectroscopies. We found four kinetic phases in the refolding reaction, which logarithmically depended on the weight fraction of octyl glucoside. We also examined the unfolding kinetics of OmpA upon heating in the presence of SDS by temperature jump experiments. A comparison of the rate constants for the refolding and the unfolding reactions in SDS-only solution at 30 degrees C revealed that the folded form of OmpA in SDS solution is less stable than the unfolding form, and that the unfolding is virtually unobservable near room temperature due to a high kinetic barrier.
Ohnishi
S
S
Institute for Protein Research, Osaka University, Yamadaoka 3-2, Suita, 565-0871, Osaka, Japan. satoshi_ohnishi@urmc.rochester.edu
Kameyama
K
K
D002942:Circular Dichroism; D004926:Escherichia coli / Q000737:chemistry*; D005960:Glucosides / Q000737:chemistry; D006358:Hot Temperature; D009943:Organophosphorus Compounds / Q000737:chemistry*; D017510:Protein Folding; D012967:Sodium Dodecyl Sulfate / Q000737:chemistry; D012996:Solutions; D013050:Spectrometry, Fluorescence; D013501:Surface-Active Agents / Q000737:chemistry
D005960:Glucosides; D009943:Organophosphorus Compounds; D012996:Solutions; D013501:Surface-Active Agents; C008705:octamethyl pyrophosphoramide; C018619:octyl-beta-D-glucoside; D012967:Sodium Dodecyl Sulfate