A fluid material with high protein content (120-310 mg mL ) was formed through the ordered self-assembly of native proteins segregated from water. This material is instantly prepared by the simple mixing of a protein solution with anionic and cationic surfactants. By changing the ratio of the surfactants based on the electrostatic characteristics of the target protein, we observed that the surfactants could function as a versatile molecular glue for protein assembly. Moreover, these protein assemblies could be disassembled back into an aqueous solution depending on the salt conditions. Owing to the water-retaining properties of the hydrophilic part of surfactants, the proteins in this material are in a water-rich environment, which maintains their native structure and function. The inclusion of water also provides functional extensibility to this material, as demonstrated by the preparation of an enzymatically active gel. We anticipate that the unique features of this material will permit the use of proteins not only in solution but also as elements of integrated functionalized materials.

Despite the well-known thermal gelation property of egg white and its wide use in the food industry, the applications of gelled egg white in materials science are limited owing to the brittleness caused by the network structure of randomly associated denatured proteins. In this study, a gel material with high mechanical strength was fabricated from crude egg white proteins. Fluid materials containing orderly condensed proteins at regular intervals were prepared by addition of ionic surfactants to an egg white protein solution. The initiation of the heat denaturation of proteins from this ordered state with spatial confinement resulted in the formation of hydrogel materials with high mechanical strength. The maximum compressive fracture strength of the fabricated egg white-based hydrogel was 34.5 MPa, which is 150-fold higher than that of gelled egg white prepared by boiling and comparable to that of strong chemically synthesized hydrogels. Chemical modification of sulfhydryl groups of proteins and treatment with a denaturant revealed that denatured polypeptide chains were covalently linked by disulfide bonds and noncovalently associated; these findings showed that the high mechanical strength of our material is due to the synergic effects of both covalent and non-covalent networks.

A double-heptamer ring chaperonin GroEL binds denatured substrate protein, ATP, and GroES to the same heptamer ring and encapsulates substrate into the central cavity underneath GroES where productive folding occurs. GroES is a disk-shaped heptamer, and each subunit has a GroEL-binding loop. The residues of the GroEL subunit responsible for GroES binding largely overlap those involved in substrate binding, and the mechanism by which GroES can replace the substrate when GroES binds to GroEL/substrate complex remains to be clarified. To address this question, we generated single polypeptide GroES by fusing seven subunits with various combinations of active and GroEL binding-defective subunits. Functional tests of the fused GroES variants indicated that four active GroES subunits were required for efficient formation of the stable GroEL/ GroES complex and five subunits were required for the productive GroEL/substrate/GroES complex. An increase in the number of defective GroES subunits resulted in a slowing of encapsulation and folding. These results indicate the presence of an intermediate GroEL/substrate/GroES complex in which the substrate and GroES bind to GroEL by sharing seven common binding sites.

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