Rationally Designed Mutations Convert de novo Amyloid-like Fibrils into Monomeric β-Sheet Proteins

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 99; no. 5; pp. 2760 - 2765
• Main Authors: Wang, Weixun, Hecht, Michael H.
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences 05.03.2002; National Acad Sciences; The National Academy of Sciences
• Subjects: Amino Acid Sequence; Amino acids; Amyloid - chemistry; Amyloid - genetics; Amyloids; Binary codes; Biological Sciences; Inclusion bodies; Libraries; Models, Molecular; Molecular Sequence Data; Monomers; Mutagenesis; Oligomers; Oligopeptides - chemistry; Periodicity; Protein Structure, Secondary; Proteins; Solar fibrils; Solubility
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.052706199

Amyloid fibrils are associated with a variety of neurodegenerative maladies including Alzheimer's disease and the prion diseases. The structures of amyloid fibrils are composed of β-strands oriented orthogonal to the fibril axis ("cross β" structure). We previously reported the design and characterization of a combinatorial library of de novo β-sheet proteins that self-assemble into fibrillar structures resembling amyloid. The libraries were designed by using a "binary code" strategy, in which the locations of polar and nonpolar residues are specified explicitly, but the identities of these residues are not specified and are varied combinatorially. The initial libraries were designed to encode proteins containing amphiphilic β-strands separated by reverse turns. Each β-strand was designed to be seven residues long, with polar (○) and nonpolar (●) amino acids arranged with an alternating periodicity (○●○●○●○). The initial design specified the identical polar/nonpolar pattern for all of the β-strands; no strand was explicitly designated to form the edges of the resulting β-sheets. With all β-strands preferring to occupy interior (as opposed to edge) locations, intermolecular oligomerization was favored, and the proteins assembled into amyloid-like fibrils. To assess whether explicit design of edge-favoring strands might tip the balance in favor of monomeric β-sheet proteins, we have now redesigned the first and/or last β-strands of several sequences from the original library. In the redesigned β-strands, the binary pattern is changed from ○●○●○●○ to ○●○K○●○ (K denotes lysine). The presence of a lysine on the nonpolar face of a β-strand should disfavor fibrillar structures because such structures would bury an uncompensated charge. The nonpolar → lysine mutations, therefore, would be expected to favor monomeric structures in which the ○●○K○●○ sequences form edge strands with the charged lysine side chain accessible to solvent. To test this hypothesis, we constructed several second generation sequences in which the central nonpolar residue of either the N-terminal β-strand or the C-terminal β-strand (or both) is changed to lysine. Characterization of the redesigned proteins shows that they form monomeric β-sheet proteins.