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Abstract
BackgroundRecently, we reported a unique approach to preserve the activity of some proteins in the presence of the denaturing agent, Sodium Dodecyl Sulfate (SDS). This was made possible by addition of the amphipathic solvent 2,4-Methyl-2-PentaneDiol (MPD), used as protecting but also as refolding agent for these proteins. Although the persistence of the protein activity in the SDS/MPD mixture was clearly established, preservation of their structure was only speculative until now.ResultsIn this paper, a detailed X-ray study addresses the pending question. Crystals of hen egg-white lysozyme were grown for the first time in the presence of MPD and denaturing concentrations of SDS. Depending on crystallization conditions, tetragonal crystals in complex with either SDS or MPD were collected. The conformation of both structures was very similar to the native lysozyme and the obtained complexes of SDS-lysozyme and MPD-lysozyme give some insights in the interplay of protein-SDS and protein-MPD interactions.ConclusionThis study clearly established the preservation of the enzyme structure in a SDS/MPD mixture. It is hypothesized that high concentrations of MPD would change the properties of SDS and lower or avoid interactions between the denaturant and the protein. These structural data therefore support the hypothesis that MPD avoids disruption of the enzyme structure by SDS and can protect proteins from SDS denaturation.
Highlights
We reported a unique approach to preserve the activity of some proteins in the presence of the denaturing agent, Sodium Dodecyl Sulfate (SDS)
Our previous work has shown that amphipathic solvents, like 2-methyl-2,4pentanediol (MPD), a commonly used precipitant for crystallization studies [3], can protect proteins from SDS denaturation, and in several cases can drive the transition from the SDS-denatured state to a functional folded state [4]
It is worth noting that this study presents one of the very few structures of proteins co-crystallized with a denaturing concentration of SDS in its native form
Summary
We reported a unique approach to preserve the activity of some proteins in the presence of the denaturing agent, Sodium Dodecyl Sulfate (SDS) This was made possible by addition of the amphipathic solvent 2,4-Methyl-2-PentaneDiol (MPD), used as protecting and as refolding agent for these proteins. Our previous work has shown that amphipathic solvents, like 2-methyl-2,4pentanediol (MPD), a commonly used precipitant for crystallization studies [3], can protect proteins from SDS denaturation, and in several cases can drive the transition from the SDS-denatured state to a functional folded state [4] This protecting effect of MPD is observed with a wide range of proteins including membrane proteins and soluble enzymes, and is not applicable if proteins are denatured in guanidine or urea, two other denaturant agents. When the enzyme was first denatured with SDS for 24 hours prior to adding MPD, full enzymatic activity was recov-
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