Abstract
In 2008, I reviewed and proposed a model for our discovery in 2005 that unrefoldable and insoluble proteins could in fact be solubilized in unsalted water. Since then, this discovery has offered us and other groups a powerful tool to characterize insoluble proteins, and we have further addressed several fundamental and disease-relevant issues associated with this discovery. Here I review these results, which are conceptualized into several novel scenarios. 1) Unlike 'misfolded proteins', which still retain the capacity to fold into well-defined structures but are misled to 'off-pathway' aggregation, unrefoldable and insoluble proteins completely lack this ability and will unavoidably aggregate in vivo with ~150 mM ions, thus designated as 'intrinsically insoluble proteins (IIPs)' here. IIPs may largely account for the 'wastefully synthesized' DRiPs identified in human cells. 2) The fact that IIPs including membrane proteins are all soluble in unsalted water, but get aggregated upon being exposed to ions, logically suggests that ions existing in the background play a central role in mediating protein aggregation, thus acting as 'dark mediators'. Our study with 14 salts confirms that IIPs lack the capacity to fold into any well-defined structures. We uncover that salts modulate protein dynamics and anions bind proteins with high selectivity and affinity, which is surprisingly masked by pre-existing ions. Accordingly, I modified my previous model. 3) Insoluble proteins interact with lipids to different degrees. Remarkably, an ALS-causing P56S mutation transforms the β-sandwich MSP domain into a helical integral membrane protein. Consequently, the number of membrane-interacting proteins might be much larger than currently recognized. To attack biological membranes may represent a common mechanism by which aggregated proteins initiate human diseases. 4) Our discovery also implies a solution to the 'chicken-and-egg paradox' for the origin of primitive membranes embedded with integral membrane proteins, if proteins originally emerged in unsalted prebiotic media.
Highlights
Self-assembly and protein folding Life represents extremely unique systems that supposedly emerged from inanimate nature, but its origin remains a great mystery
Previously we have addressed this question by titrating unrefoldable and insoluble proteins solubilized in unsalted water with NaCl, and demonstrated that the addition of NaCl triggered no formation of well-folded structures, NaCl, is generally considered to be neutral in the Hofmeister series and it is of fundamental interest to evaluate the effects of other salts
Our results provide a mechanism to rationalize a recent report that the P56S mutant is able to trigger the formation of a novel form of organized smooth endoplasmic reticulum (ER) with stacked cisternae[104,105], despite failing to detect significant formation of aggregated inclusions in motor neurons derived from induced pluripotent stem cells of patients carrying the P56S mutation[106]
Summary
Self-assembly and protein folding Life represents extremely unique systems that supposedly emerged from inanimate nature, but its origin remains a great mystery. Due to the previous absence of a general method to solubilize these insoluble proteins in the aqueous solution without adding denaturants and detergents, the solution conformations of these unrefoldable and insoluble proteins remained completely unknown The lack of such knowledge leads to the inability to answer a fundamental question: whether these mutants still possess the intrinsic capacity to fold into well-defined structures, and their aggregation is only due to ‘off-pathway’ misfolding, or whether they have lost this ability; and they aggregate following the mechanism underlying ‘intrinsical insoluble proteins’. Previously we have addressed this question by titrating unrefoldable and insoluble proteins solubilized in unsalted water with NaCl, and demonstrated that the addition of NaCl triggered no formation of well-folded structures, NaCl, is generally considered to be neutral in the Hofmeister series and it is of fundamental interest to evaluate the effects of other salts. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript
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