Synthetic Nanochaperones Facilitate Refolding of Denatured Proteins

Abstract

The folding process of a protein is inherently error-prone, owing to the large number of possible conformations that a protein chain can adopt. Partially folded or misfolded proteins typically expose hydrophobic surfaces and tend to form dysfunctional protein aggregates. Therefore, materials that can stabilize unfolded proteins and then efficiently assist them refolding to its bioactive form are of significant interest. Inspired by natural chaperonins, we have synthesized a series of polymeric nanochaperones that can facilitate the refolding of denatured proteins with a high recovery efficiency (up to 97%). Such nanochaperones possess phase-separated structure with hydrophobic microdomains on the surface. This structure allows nanochaperones to stabilize denatured proteins by binding them to the hydrophobic microdomains. We have also investigated the mechanism by which nanochaperones assist the protein refolding and established the design principles of nanochaperones in order to achieve effective recovery of a certain protein from their denatured forms. With a carefully designed composition of the microdomains according to the surface properties of the client proteins, the binding affinity between the hydrophobic microdomain and the denatured protein molecules can be tuned precisely, which enables the self-sorting of the polypeptides and the refolding of the proteins into their bioactive states. This work provides a feasible and effective strategy to recover inclusion bodies to their bioactive forms, which has potential to reduce the cost of the manufacture of recombinant proteins significantly.

A Balance Between Capture and Release: How Nanochaperones Regulate Refolding of Thermally Denatured Proteins

“…Four nanochaperones, pos‐nChap‐1, pos‐nChap‐2, neg‐nChap‐1, and neg‐nChap‐2, as listed in Table S1 (see Supporting Information, SI), were prepared according to our previous report [10] . The chemical structure of each polymer is shown in Figure S1 (SI).…”

“…However, it is still unclear whether client protein refolding is affected by the release from the nanochaperones. On this subject, we designed and fabricated a nanochaperone system that could facilitate de novo protein folding [10] . We further found a new working mechanism of nanochaperones, similar to that of adenosine triphosphate (ATP)‐independent chaperones.…”

Neurotransmitter-derived lipidoids (NT-lipidoids) for enhanced brain delivery through intravenous injection

“…The lipid structure was confirmed using ESI-MS. The NT1-Neu was synthesized according to the reported procedure ( 36 ). Briefly, into a dry flask, palmitic-acid-5-hydroxy-stearic-acid and dichloromethane (DCM) were added, and then dropwise addition of (COCl) 2 that was dissolved in DCM.…”

Nanochaperone‐Based Strategies to Control Protein Aggregation Linked to Conformational Diseases

“…[41][42][43] Such artificial chaperones could stabilize the denatured state of lysozyme and assist its refolding and release upon stimuli-induced transformation of the hydrophobic domains into hydrophilic chains. [44,45] Reversible protein adsorption during up to seven heating/cooling cycles was achieved. [46] Such mixed-shell polymeric micelles have also been co-assembled with the g-secretase inhibitor DAPT to simultaneously reduce Ab overproduction in AD.…”

“…Micellar NPs that mimic HSP70s have been designed that could suppress Aβ aggregation and facilitate its clearance [41–43] . Such artificial chaperones could stabilize the denatured state of lysozyme and assist its refolding and release upon stimuli‐induced transformation of the hydrophobic domains into hydrophilic chains [44, 45] . Reversible protein adsorption during up to seven heating/cooling cycles was achieved [46] .…”