Mesterséges beta-aminosav tartalmú önszerveződő polimerek térszerkezetének és stabilitásának vizsgálata = Study os space structure and stability of unnatural beta-amino acid containing foldamers

A projektben választ kaptunk a beta-peptidek másodlagos szerkezeteinek sztereokémiai szabályzásával, konformációs polimorfizmusával és a kontrolált önrendeződésével kapcsolatos legfontosabb kérdésekre. Megvalósítottuk a beta-peptid másodlagos szerkezetek gerinckonfigurációval történő szabályzását. Kimutattuk, hogy a csakúgy mint az alfa-peptidek, a beta-peptid hélixek is képesek a konformációs polimorfizmusra, ami igazolta valódi önrendeződési folyamatot a hélixképződéssel kapcsolatosan. Tervezett másodlagos szerkezetű beta-peptid oligomereket fehasználva nanostrukturált részecskéket (vezikulák és fibrillumok) hoztunk létre, melyeket harmadlagos szerkezeti elemek építenek fel (hélix-köteg, redőzött szendvics). A megfigyelések hozzájárulhatnak a beta-peptid alapvázzal rendelkező bioaktív molekulák tervezéséhez és bio-nanotechnológiai alkalmazások is elképzelhetők. | The project has shed light over the most important questions related to the stereochemical control of the secondary structure formation, the conformational polimorphism and controlled self-association of beta-peptides. The control of the secondary structure formation has been achieved by tuning beta-peptide backbone configuration. It has been shown, that similarly to the alpha-peptides, the conformational polimorhism is an inherent property of the beta-peptides, which proves the true self-organization in connection with helix formation. By using designed beta-peptide secondary structure units, nanostructured particles have been constructed (vesicles, fibrils), which are organized by the forces of the tertiary structure motifs (helix-bundle, pleated sheet sandwich). The observations may contribute to the design of bioactive substances having beta-peptide skeleton and their application in bio-nanotechnology is also possible

Characterization of Human Islet Amyloid Peptide via IM-MS and Infrared Spectroscopy

Protein-misfolding, aggregation and accumulation of insoluble deposits are the hallmark features of a variety of human diseases such as Parkinson's disease and diabetes mellitus type II. The latter is a systemic disorder characterized by insulin resistance, impaired insulin secretion, beta-cell apoptosis and islet amyloid formation. Fibrillar aggregates from the selfassembly of human islet amyloid polypeptide (hIAPP) are major component of islet amyloids. Accumulating evidence suggests that not the mature fibrils, but rather smaller, soluble, polymorphic and highly dynamic oligomers preceding the formation of the fibrils are the cytotoxic species. A detailed insight into the structures of the oligomeric intermediates is crucial for identification of potential targets and development of therapeutic strategies. However, the polydisperse nature of these peptides makes their structural characterization challenging. The traditional condensed-phase analytical techniques provide only averaged structural information on the dynamic ensemble. However, information on the structure of isolated species can be obtained by employing gas-phase techniques. Here, infrared action spectroscopy in combination with ion-mobility spectrometry is used to gain insight into the secondary structure of these isolated peptides. The orthogonal combination of these techniques allows to obtain fingerprint vibrational spectra of m/z- and conformer-selected species. The secondary structure of the full-length hIAPP, as well as of metal-associated hIAPP and hIAPP in heterogeneous co-assemblies with a fragment of the prion protein (PrP106-126) and the hexapeptide VEALYL were investigated. The obtained data suggests that the secondary structure of the monomeric hIAPP contains a significant fraction of alpha-helical motifs, which is seemingly maintained upon metal binding, self-assembly, or heterogeneous co-assembly with other peptides. These results provide solid evidence for the importance of helical intermediates in the formation of amyloids

Single-conformation IR and UV spectroscopy of a prototypical heterogeneous α/β-peptide: is it a mixed-helix former?

Synthetic foldamers are non-natural polymers designed to fold into unique secondary structures that either mimic nature’s preferred secondary structures, or expand their possibilities. Among the most studied synthetic foldamers are $\beta$-peptides, which lengthen the distance between amide groups from the single substituted carbon spacer in $\alpha$-peptides by one additional carbon. We present data on a mixed $\alpha$/$\beta$ tri-peptide in which a single $\beta$-residue with a conformationally constrained cis-2-aminocyclohexanecarboxylic acid (cis-ACHC) substitution is inserted in an $\alpha$-peptide backbone to form Ac-Ala-$\beta$-ACHC-Ala-NHBn. This $\alpha$$\beta$$\alpha$ structure is known in longer sequences to prefer formation of a 9/11 mixed helix. Under isolated, jet cooled conditions, four unique conformers were observed in the expansion. The dominant conformer is configured in a tetramer cycle with every amide carbonyl and amine group involved in hydrogen bonding, giving rise to a tightly folded C12/C7/C8/C7 structure reminiscent of a $\beta$-turn. This talk will describe the conformation specific IR and UV spectroscopy methods used to study this mixed peptide, as well as its experimentally observed conformational preferences

Influence of Nanoparticle Size and Shape on Oligomer Formation of an Amyloidogenic Peptide

Understanding the influence of macromolecular crowding and nanoparticles on the formation of in-register $\beta$-sheets, the primary structural component of amyloid fibrils, is a first step towards describing \emph{in vivo} protein aggregation and interactions between synthetic materials and proteins. Using all atom molecular simulations in implicit solvent we illustrate the effects of nanoparticle size, shape, and volume fraction on oligomer formation of an amyloidogenic peptide from the transthyretin protein. Surprisingly, we find that inert spherical crowding particles destabilize in-register $\beta$-sheets formed by dimers while stabilizing $\beta$-sheets comprised of trimers and tetramers. As the radius of the nanoparticle increases crowding effects decrease, implying smaller crowding particles have the largest influence on the earliest amyloid species. We explain these results using a theory based on the depletion effect. Finally, we show that spherocylindrical crowders destabilize the ordered $\beta$-sheet dimer to a greater extent than spherical crowders, which underscores the influence of nanoparticle shape on protein aggregation

Dimensionality of Carbon Nanomaterials Determines the Binding and Dynamics of Amyloidogenic Peptides: Multiscale Theoretical Simulations

Experimental studies have demonstrated that nanoparticles can affect the rate of protein self-assembly, possibly interfering with the development of protein misfolding diseases such as Alzheimer's, Parkinson's and prion disease caused by aggregation and fibril formation of amyloid-prone proteins. We employ classical molecular dynamics simulations and large-scale density functional theory calculations to investigate the effects of nanomaterials on the structure, dynamics and binding of an amyloidogenic peptide apoC-II(60-70). We show that the binding affinity of this peptide to carbonaceous nanomaterials such as C60, nanotubes and graphene decreases with increasing nanoparticle curvature. Strong binding is facilitated by the large contact area available for π-stacking between the aromatic residues of the peptide and the extended surfaces of graphene and the nanotube. The highly curved fullerene surface exhibits reduced efficiency for π-stacking but promotes increased peptide dynamics. We postulate that the increase in conformational dynamics of the amyloid peptide can be unfavorable for the formation of fibril competent structures. In contrast, extended fibril forming peptide conformations are promoted by the nanotube and graphene surfaces which can provide a template for fibril-growth

Substrate control in stereoselective lanthionine biosynthesis.

Enzymes are typically highly stereoselective catalysts that enforce a reactive conformation on their native substrates. We report here a rare example in which the substrate controls the stereoselectivity of an enzyme-catalysed Michael-type addition during the biosynthesis of lanthipeptides. These natural products contain thioether crosslinks formed by a cysteine attack on dehydrated Ser and Thr residues. We demonstrate that several lanthionine synthetases catalyse highly selective anti-additions in which the substrate (and not the enzyme) determines whether the addition occurs from the re or si face. A single point mutation in the peptide substrate completely inverted the stereochemical outcome of the enzymatic modification. Quantum mechanical calculations reproduced the experimentally observed selectivity and suggest that conformational restraints imposed by the amino-acid sequence on the transition states determine the face selectivity of the Michael-type cyclization