Article pubs.acs.org/Biomac
The Role of Protein Hydrophobicity in Conformation Change and Self-Assembly into Large Amyloid Fibers Devin M. Ridgley, Elizabeth C. Claunch, Parker W. Lee, and Justin R. Barone* Biological Systems Engineering Department, Virginia Tech, 301D HABB1, Blacksburg, Virginia 24061, United States S Supporting Information *
ABSTRACT: It has been found that a short hydrophobic “template” peptide and a larger α-helical “adder” protein cooperatively self-assemble into micrometer sized amyloid fibers. Here, a common template of trypsin hydrolyzed gliadin is combined with six adder proteins (α-casein, α-lactalbumin, amylase, hemoglobin, insulin, and myoglobin) to determine what properties of the adder protein drive amyloid selfassembly. Utilizing Fourier Transform-Infrared (FT-IR) spectroscopy, the Amide I absorbance reveals that the observed decrease in α-helix with time is approximately equal to the increase in high strand density β-sheet, which is indicative of amyloid formation. The results show that the hydrophobic moment is a good predictor of conformation change but the fraction of aliphatic amino acids within the α-helices is a better predictor. Upon drying, the protein mixtures form large amyloid fibers. The fiber twist is dependent on the aliphatic index and molecular weight of the adder protein. Here we demonstrate that it is possible to predict the propensity of an adder protein to unfold into an amyloid structure and to predict the fiber morphology, both from adder protein molecular features, which can be applied to the pragmatic engineering of large amyloid fibers.
■
nature.30−35 Escherichia coli and Streptomyces coelicolor bacteria produce amyloid fibrils for surface-to-surface adhesion to create a matrix that can lead to bacterial biofilms.36−38 Hydrophobin protein secreted from the fungus Magnaporthe grisea produces an amyloid hyphae that allows the fungus to attach to hydrophobic surfaces.39−41 Atomic force microscopy (AFM) can be used to measure nanofibril cross-section and topography to calculate bending rigidity and moment of inertia. Using this technique, Knowles et al. found elastic moduli of E = 2−14 GPa for nanofibrils of “regular” morphology and E = 0.14−0.40 GPa for nanofibrils of “irregular” morphology.42 AFM force spectroscopy yielded E = 3.3 ± 0.14 GPa for insulin amyloid fibrils.45 Peak force quantitative nanomechanical AFM found E = 3.7 ± 1.1 GPa for β-lactoglobulin amyloid fibrils.43,44 Molecular simulations predict amyloids to have E = 2.34 GPa at small strains and E = 12.43 GPa and 18.05 at large compressive and tensile strains, respectively.18,46 These values approach the moduli of silk fibers but silk requires spinning to form the fiber while amyloid fibrils form spontaneously.11,45,47−51 The functional role, high modulus, and easier production compared to other fibers like silk make amyloids an interesting motif for high performance biomaterials.19,52−54 Recent studies have focused on using the amyloid fibril as an engineered biomaterial. Adamcik et al. produced β-lactoglobulin amyloid tapes of approximately 20 nm wide with adjustable
INTRODUCTION Over the last few decades there has been extensive research focused on understanding the cause and mechanism of a debilitating class of neurodegenerative diseases termed “prion” diseases. 1−7 Prions are formed from the spontaneous misfolding of a protein into β-sheets that in some cases can further self-assemble into an amyloid fibril.1,6 Amyloid fibrils are characterized by a high strand density β-sheet secondary structure oriented perpendicular to the fibril axis.8 High strand density β-sheets typical of amyloids have a lower twist and more protein strands per β-sheet compared to native β-sheets found in silk or β-keratin.9 As a result, the Amide I absorbance of high strand density β-sheets as measured using Fourier transform infrared (FT-IR) spectroscopy appears at lower wavenumbers,
