Conducting microhelices from self-assembly of protein fibrils.

Herein we utilize insulin to prepare amyloid based chiral helices with either right or left handed helicity. We demonstrate that the helices can be utilized as structural templates for the conducting polymer alkoxysulfonate poly(ethylenedioxythiophene) (PEDOT-S). The chirality of the helical assembly is transferred to PEDOT-S as demonstrated by polarized optical microscopy (POM) and Circular Dichroism (CD).

The emergence of superstructural order in insulin amyloid fibrils upon multiple rounds of self-seeding.

Typically, elongation of an amyloid fibril entails passing conformational details of the mother seed to daughter generations of fibrils with high fidelity. There are, however, several factors that can potentially prevent such transgenerational structural imprinting from perpetuating, for example heterogeneity of mother seeds or so-called conformational switching. Here, we examine phenotypic persistence of bovine insulin amyloid ([BI]) upon multiple rounds of self-seeding under quiescent conditions.

Thioflavin T: Electronic Circular Dichroism and Circularly Polarized Luminescence Induced by Amyloid Fibrils.

The circularly polarized luminescence (CPL) spectrum of thioflavin T (ThT) bound to insulin amyloid fibrils has been measured for the first time. It has been found that the samples exhibiting induced circular dichroism (CD) retain the optical activity in the CPL spectra, with the same sign of the rotatory strength. The fluorescence dissymmetry factor is substantial (of the order of magnitude 10(-2) ).

Dimethyl Sulfoxide Induced Destabilization and Disassembly of Various Structural Variants of Insulin Fibrils Monitored by Vibrational Circular Dichroism.

Dimethyl sulfoxide (DMSO) induced destabilization of insulin fibrils has been previously studied by Fourier transform infrared spectroscopy and interpreted in terms of secondary structural changes. The variation of this process for fibrils with different types of higher-order morphological structures remained unclear. Here, we utilize vibrational circular dichroism (VCD), which has been reported to provide a useful biophysical probe of the supramolecular chirality of amyloid fibrils, to characterize changes in the macroscopic chirality following DMSO-induced disassembly for two types of insulin fibrils formed under different conditions, at different reduced pH values with and without added salt and agitation.

On the function and fate of chloride ions in amyloidogenic self-assembly of insulin in an acidic environment: salt-induced condensation of fibrils.

Formation of amyloid fibrils is often facilitated in the presence of specific charge-compensating ions. Dissolved sodium chloride is known to accelerate insulin fibrillation at low pH that has been attributed to the shielding of electrostatic repulsion between positively charged insulin molecules by chloride ions. However, the subsequent fate of Cl(-) anions; that is, possible entrapment within elongating fibrils or escape into the bulk solvent, remains unclear.

Highly amyloidogenic two-chain peptide fragments are released upon partial digestion of insulin with pepsin.

Proteases play a well recognized role in the emergence of highly aggregation-prone protein fragments in vivo, whereas in vitro limited proteolysis is often employed to probe different phases of amyloidogenic pathways. Here, we show that addition of moderate amounts of pepsin to acidified bovine insulin at close to physiological temperature results in an abrupt self-assembly of amyloid-like fibrils from partially digested insulin fragments. Biochemical analysis of the pepsin-induced fibrils implicates peptide fragments (named H) consisting of the 13 or 15 N-terminal residues of the A-chain and 11 or 13 N-terminal residues of the B-chain linked by the disulfide bond between Cys-7A-Cys-7B as the main constituents.

Master and slave relationship between two types of self-propagating insulin amyloid fibrils.

Cross-seeding of fibrils of bovine insulin (BI) and Lys(B31)-Arg(B32) human insulin analog (KR) induces self-propagating amyloid variants with infrared features inherited from mother seeds. Here we report that when native insulin (BI or KR) is simultaneously seeded with mixture of equal amounts of both templates (i.e.

Supramolecular photochirogenesis with functional amyloid superstructures.

Chiral variants of amyloid fibrils prepared by agitating acidified solutions of bovine insulin at 45 °C not only induced quasi-mirror-imaged circular dichroism spectra upon complexation with 2-anthracenecarboxylate but also gave anti-head-to-head-cyclodimers of the opposite absolute configurations upon photoirradiation.

Vortex-induced amyloid superstructures of insulin and its component A and B chains.

Insulin is an amyloid-forming polypeptide built of two disulfide-linked chains (A and B), both themselves amyloidogenic. An interesting property of insulin is that agitation strongly influences the course of its aggregation, resulting in characteristic chiral superstructures of amyloid fibrils. Here, we investigate the self-assembly of these superstructures by comparing the quiescent and vortex-assisted aggregation of insulin and its individual A and B chains in the presence or absence of reducing agent tris(2-carboxyethyl)phosphine (TCEP).

Amino acid sequence determinants in self-assembly of insulin chiral amyloid superstructures: role of C-terminus of B-chain in association of fibrils.

Formation of chiral amyloid superstructures is a newly recognised phenomenon observed upon agitation-assisted fibrillation of bovine insulin. Here, by surveying several amyloidogenic precursors we examine whether formation of such entities is unique to bovine insulin. Our results indicate that only bovine, human, and porcine insulins are capable of chiral superstructural self-assembly.

Conformational memory effect reverses chirality of vortex-induced insulin amyloid superstructures.

Formation of amyloid fibrils is often associated with intriguing far-from-equilibrium phenomena such as conformational memory effects or flow-driven self-assembly. Insulin is a model amyloidogenic polypeptide forming distinct structural variants of fibrils, which self-propagate through seeding. According to infrared absorption, fibrils from bovine insulin ([BI]) and Lys(B31)-Arg(B32) human insulin analogue ([KR]) cross-seed each other and imprint distinct structural features in daughter fibrils.

On the DMSO-dissolved state of insulin: a vibrational spectroscopic study of structural disorder.

Upon dissolving in dimethyl sulfoxide (DMSO), native insulin and insulin amyloid fibrils convert into an identical disordered structural state based on IR spectral characteristics. Here, we investigate the DMSO-denatured state of insulin using a number of spectroscopic methods: near-UV circular dichroism, infrared absorption spectroscopy, vibrational circular dichroism (VCD), Raman scattering, and Raman optical activity (ROA), as well as by carrying out 140-ns-long molecular dynamics (MD) simulations of DMSO-dissolved native insulin monomers. According to this work, the DMSO-solvated state of insulin is an ensemble of conformations including polyproline II-type helix and possibly a residual α-helical structure.

Thioflavin T forms a non-fluorescent complex with α-helical poly-L-glutamic acid.

Thioflavin T (ThT) is a molecular-rotor-type fluorophore reputed for the selective binding to amyloid fibrils. Using induced circular dichroism, here we show that ThT binds in an orderly manner to α-helical poly-L-glutamic acid (PLGA) implying that neither stacked β-sheets nor π-π stacking interactions are necessary for the binding between the dye and proteins.

Chiral superstructures of insulin amyloid fibrils.

Hydrodynamic forces are capable of inducing structural order in dispersed solid phases, and of causing symmetry-breaking when chiral crystals precipitate from an achiral liquid phase. Until it was observed upon vortex-assisted fibrillation of insulin, such behavior had been thought to be confined to few unbiological systems. In this paper we are discussing chiroptical properties of two chiral variants of insulin amyloid, termed +ICD and -ICD, which form during the process of chiral bifurcation in vortexed solutions of aggregating insulin.

Insulin amyloid superstructures as templates for surface enhanced Raman scattering.

Nanostructuring of noble metal surfaces with biomorphic and biological templates facilitates a variety of applications of surface enhanced Raman scattering (SERS). Here we show that the newly reported insulin amyloid superstructures may be employed as stable nanoscaffolds for metallic Au films providing an effective substrate for SERS on covalently bound molecules of 4-mercaptobenzoic acid (4-MBA). The vortex-aligned insulin fibrils are capable of templating nanopatterns in sputtered Au layers without overlapping the SERS spectra of 4-MBA with vibrational bands stemming from the protein.