Self-assembled monolayers from a designed combinatorial library of de novo β-sheet proteins

Abstract

A variety of naturally occurring biomaterials owe their unusual structural and mechanical properties to layers of ␤-sheet proteins laminated between layers of inorganic mineral. To explore the possibility of fabricating novel two-dimensional protein layers, we studied the self-assembly properties of de novo proteins from a designed combinatorial library. Each protein in the library has a distinct 63 amino acid sequence, yet they all share an identical binary pattern of polar and nonpolar residues, which was designed to favor the formation of six-stranded amphiphilic ␤-sheets. Characterization of proteins isolated from the library demonstrates that (i) they self assemble into monolayers at an air͞water interface; (ii) the monolayers are dominated by ␤-sheet secondary structure, as shown by both circular dichroism and infrared spectroscopies; and (iii) the measured areas (500-600 Å 2 ) of individual protein molecules in the monolayers match those expected for proteins folded into amphiphilic ␤-sheets. The finding that similar structures are formed by distinctly different protein sequences suggests that assembly into ␤-sheet monolayers can be encoded by binary patterning of polar and nonpolar amino acids. Moreover, because the designed binary pattern is compatible with a wide variety of different sequences, it may be possible to fabricate ␤-sheet monolayers by using combinations of side chains that are explicitly designed to favor particular applications of novel biomaterials. P roteins play key roles in controlling the self-assembly of biological materials (1-5). In recent years, interest has grown in mimicking biological self assembly (2-11) with peptides or proteins designed de novo. This interest is motivated both by a desire to test our understanding of self assembly in natural systems and by the drive to lay foundations for the fabrication of novel protein-based biomaterials.Self-assembled two-dimensional layers of proteins are of particular interest for the fabrication of biomaterials. Such layers play important roles in biological mineralization by controlling the size, orientation, and morphology of inorganic crystals at the surface of the protein layer (2,5,(11)(12)(13)(14). A well-studied example of protein͞inorganic layering occurs in the nacre of mollusk shells (''mother of pearl'') (4, 5, 11-17). These laminated structures are composed of alternating layers of a protein-rich matrix and aragonite (a crystal form of calcium carbonate). The proteins in the protein-rich layer are dominated by ␤-sheet secondary structure (1,5,(11)(12)(13)(14).To explore the possibility of fabricating de novo ␤-sheet layers, we studied the self-assembly properties of a collection of de novo proteins derived from a combinatorial library of sequences designed to favor the formation of amphiphilic ␤-strands punctuated by reverse turns (18,19). Each sequence in the library is distinct. However, all sequences are the same length (63 residues), and they all share the identical binary patterning of polar and nonpolar residues. Her...

Template-Directed Assembly of a de Novo Designed Protein

“…3 Previous studies of proteins from this library demonstrated that they self-assemble into (i) amyloid-like fibrils in a homogeneous aqueous environment 3 and (ii) -sheet monolayers at an air/water interface. 4 Given the tendency of these de noVo proteins to selfassemble in the absence of an external template, we sought to assess whether the presence of a highly ordered surface (template) would induce structures that recapitulate the order and symmetry of the template.To assess the propensity of our de noVo proteins to undergo template-directed assembly, 6 we focused on one protein, 17-6 (Figure 1), from our -sheet library. When probing the ability of a surface to template protein assembly, it is important to present the template with a protein that is not preassembled.…”

“…3 Previous studies of proteins from this library demonstrated that they self-assemble into (i) amyloid-like fibrils in a homogeneous aqueous environment 3 and (ii) -sheet monolayers at an air/water interface. 4 Given the tendency of these de noVo proteins to selfassemble in the absence of an external template, we sought to assess whether the presence of a highly ordered surface (template) would induce structures that recapitulate the order and symmetry of the template.…”

Peptide Self-Assembled Nanostructures for Drug Delivery Applications

“…The -sheet consists of alternating hydrophilic and hydrophobic amino acids in the peptide sequence, which can provide amphiphilic property to the peptide that drives the self-assembly ofsheets [96]. The -sheet peptides also could be utilized to form many different nanostructures including nanotubes, monolayers in nanoscale order, and nanoribbons [97][98][99][100][101]. For example, -sheet peptide QQRFEWEFEQQ can selfassemble into a pH responsive hydrogel using peptides' ionizable side chains from Glu and Arg amino acids.…”

Peptide based amphiphiles

“…The above mentioned ideas were elaborated by Xu et al 9 who prepared small proteins designed to form six b-strands, each one consisting of seven alternating polar and non-polar residues, which were connected by four amino acids to induce reverse turns. In this case b-sheet monolayers were formed.…”