Anil K. Mehta

Engineering metal ion coordination to regulate amyloid fibril assembly and toxicity

Protein and peptide assembly into amyloid has been implicated in functions that range from beneficial epigenetic controls to pathological etiologies. However, the exact structures of the assemblies that regulate biological activity remain poorly defined. We have previously used Zn2+ to modulate the assembly kinetics and morphology of congeners of the amyloid β peptide (Aβ) associated with Alzheimers disease. We now reveal a correlation among Aβ-Cu2+ coordination, peptide self-assembly, and neuronal viability. By using the central segment of Aβ, HHQKLVFFA or Aβ(13–21), which contains residues H13 and H14 implicated in Aβ-metal ion binding, we show that Cu2+ forms complexes with Aβ(13–21) and its K16A mutant and that the complexes, which do not self-assemble into fibrils, have structures similar to those found for the human prion protein, PrP. N-terminal acetylation and H14A substitution, Ac-Aβ(13–21)H14A, alters metal coordination, allowing Cu2+ to accelerate assembly into neurotoxic fibrils. These results establish that the N-terminal region of Aβ can access different metal-ion-coordination environments and that different complexes can lead to profound changes in Aβ self-assembly kinetics, morphology, and toxicity. Related metal-ion coordination may be critical to the etiology of other neurodegenerative diseases.

Rational Design of Helical Nanotubes from Self-assembly of Coiled-coil Lock Washers

Design of a structurally defined helical assembly is described that involves recoding of the amino acid sequence of peptide GCN4-pAA. In solution and the crystalline state, GCN4-pAA adopts a 7-helix bundle structure that resembles a supramolecular lock washer. Structurally informed mutagenesis of the sequence of GCN4-pAA afforded peptide 7HSAP1, which undergoes self-association into a nanotube via noncovalent interactions between complementary interfaces of the coiled-coil lock-washer structures. Biophysical measurements conducted in solution and the solid state over multiple length scales of structural hierarchy are consistent with self-assembly of nanotube structures derived from 7-helix bundle subunits. The dimensions of the supramolecular assemblies are similar to those observed in the crystal structure of GCN4-pAA. Fluorescence studies of the interaction of 7HSAP1 with the solvatochromic fluorophore PRODAN indicated that the nanotubes could encapsulate shape-appropriate small molecules with high binding affinity.

Templating Molecular Arrays in Amyloid’s Cross-β Grooves

Amyloid fibers, independent of primary amino acid sequence, share a common cross-beta structure and bind the histochemical dye Congo Red (CR). Despite extensive use of CR in amyloid diagnostics, remarkably little is known about the specific and characteristic binding interactions. Fibril insolubility, morphological inhomogeneity, and multiple possible ligand binding sites all conspire to limit characterization. Here, we have exploited the structure of cross-beta nanotubes, which limit the number of potential binding sites, to directly interrogate cross-beta laminate grooves. CR bound to cross-beta nanotubes displays the hallmark apple-green interference color, a broad red-shifted low energy transition, and a K(d) of 1.9 +/- 0.5 microM. Oriented electron diffraction and linear dichroism defines the orientation of CR as parallel to the amyloid long axis and colinear with laminate grooves. The broad red-shifted UV signature of CR bound to amyloid can be explained by semiempirical quantum calculations that support the existence of a precise network of J- and H-CR aggregates, illuminating the ability of the amyloid to organize molecules into extended arrays that underlie the remarkable diagnostic potential of CR.

Peptides Organized as Bilayer Membranes

From the organizing poten-tial of two-dimensional phospholipid membranes to theinformation-rich DNA helices, from the mechanical actinand tubulin cables to the structural collagen and elastinnetworks, these self-assembling asymmetric arrays define thearchitectures of all cells and tissues. Recent covalent hybridsof traditional biological macromolecular families (e.g.,nucleic acids with proteins

Kinetic Intermediates in Amyloid Assembly

In contrast to an expected Ostwald-like ripening of amyloid assemblies, the nucleating core of the Dutch mutant of the Aβ peptide of Alzheimers disease assembles through a series of conformational transitions. Structural characterization of the intermediate assemblies by isotope-edited IR and solid-state NMR reveals unexpected strand orientation intermediates and suggests new nucleation mechanisms in a progressive assembly pathway.

Peptide membranes in chemical evolution

Simple surfactants achieve remarkable long-range order in aqueous environments. This organizing potential is seen most dramatically in biological membranes where phospholipid assemblies both define cell boundaries and provide a ubiquitous structural scaffold for controlling cellular chemistry. Here we consider simple peptides that also spontaneously assemble into exceptionally ordered scaffolds, and review early data suggesting that these structures maintain the functional diversity of proteins. We argue that such scaffolds can achieve the required molecular order and catalytic agility for the emergence of chemical evolution.

Shape selection and multi-stability in helical ribbons

Helical structures, almost ubiquitous in biological systems, have inspired the design and manufacturing of helical devices with applications in nanoelecromechanical systems, morphing structures, optoelectronics, micro-robotics, and drug delivery devices. Meanwhile, multi-stable structures, represented by the Venus flytrap and slap bracelet, have attracted increasing attention due to their applications in making artificial muscles, bio-inspired robots, deployable aerospace components, and energy harvesting devices. Here we show that the mechanical anisotropy pertinent to helical deformation, together with geometric nonlinearity associated with multi-stability, can lead to a selection principle of the geometric shape and multi-stability in spontaneous helical ribbons. Simple table-top experiments were also performed to illustrate the working principle. Our work will promote understanding of spontaneous curling, twisting, wrinkling of thin objects, and their instabilities. The proposed theoretical framework can also serve as a tool for developing functional structures and devices featuring tunable, morphing geometries and smart actuation mechanisms that can be applied in a spectrum of areas.

Nucleobase-Directed Amyloid Nanotube Assembly

Cytosine nucleobases were successfully incorporated into the side chain of the self-assembling amyloid peptide fragment HHQALVFFA to give ccAQLVFFA. At a pH range of 3-4, where cytosine is expected to be partially protonated, small-angle X-ray scattering analyses revealed the nucleobase peptide assembles to be well-defined nanotubes with an outer diameter of 24.8 nm and wall thicknesses of 3.3 nm. FT-IR and X-ray diffraction confirmed beta-sheet-rich assembly with the characteristic cross-beta architecture of amyloid. The beta-sheet registry, determined by measuring (13)CO-(13)CO backbone distances with solid-state NMR and linear dichroism, placed the cytosine bases roughly perpendicular to the nanotube axis, resulting in a model where the complementary interactions between the cytosine bases increases beta-sheet stacking to give the nanotube architecture. These scaffolds then extend the templates used to encode biological information beyond the nucleic acid duplexes and into covalent networks whose self-assembly is still defined by a precise complementarity of the side-chain registry.

Digital and analog chemical evolution.

Living matter is the most elaborate, elegant, and complex hierarchical material known and is consequently the natural target for an ever-expanding scientific and technological effort to unlock and deconvolute its marvelous forms and functions. Our current understanding suggests that biological materials are derived from a bottom-up process, a spontaneous emergence of molecular networks in the course of chemical evolution. Polymer cooperation, so beautifully manifested in the ribosome, appeared in these dynamic networks, and the special physicochemical properties of the nucleic and amino acid polymers made possible the critical threshold for the emergence of extant cellular life. These properties include the precise and geometrically discrete hydrogen bonding patterns that dominate the complementary interactions of nucleic acid base-pairing that guide replication and ensure replication fidelity. In contrast, complex and highly context-dependent sets of intra- and intermolecular interactions guide protein folding. These diverse interactions allow the more analog environmental chemical potential fluctuations to dictate conformational template-directed propagation. When these two different strategies converged in the remarkable synergistic ribonucleoprotein that is the ribosome, this resulting molecular digital-to-analog converter achieved the capacity for both persistent information storage and adaptive responses to an ever-changing environment. The ancestral chemical networks that preceded the Central Dogma of Earths biology must reflect the dynamic chemical evolutionary landscapes that allowed for selection, propagation, and diversification and ultimately the demarcation and specialization of function that modern biopolymers manifest. Not only should modern biopolymers contain molecular fossils of this earlier age, but it should be possible to use this information to reinvent these dynamic functional networks. In this Account, we review the first dynamic network created by modification of a nucleic acid backbone and show how it has exploited the digital-like base pairing for reversible polymer construction and information transfer. We further review how these lessons have been extended to the complex folding landscapes of templated peptide assembly. These insights have allowed for the construction of molecular hybrids of each biopolymer class and made possible the reimagining of chemical evolution. Such elaboration of biopolymer chimeras has already led to applications in therapeutics and diagnostics, to the construction of novel nanostructured materials, and toward orthogonal biochemical pathways that expand the evolution of existing biochemical systems. The ability to look beyond the primordial emergence of the ribosome may allow us to better define the origins of chemical evolution, to extend its horizons beyond the biology of today and ask whether evolution is an inherent property of matter unbounded by physical limitations imposed by our planets diverse environments.

Copper(II)-bis-Histidine Coordination Structure in a Fibrillar Amyloid β-Peptide Fragment and Model Complexes Revealed by Electron Spin Echo Envelope Modulation Spectroscopy

Truncated and mutated amyloid‐β (Aβ) peptides are models for systematic study—in homogeneous preparations—of the molecular origins of metal ion effects on Aβ aggregation rates, types of aggregate structures formed, and cytotoxicity. The 3D geometry of bis‐histidine imidazole coordination of CuII in fibrils of the nonapetide acetyl‐Aβ(13–21)H14A has been determined by powder 14N electron spin echo envelope modulation (ESEEM) spectroscopy. The method of simulation of the anisotropic combination modulation is described and benchmarked for a CuII‐bis‐cis‐imidazole complex of known structure. The revealed bis‐cis coordination mode, and the mutual orientation of the imidazole rings, for CuII in Ac‐Aβ(13–21)H14A fibrils are consistent with the proposed β‐sheet structural model and pairwise peptide interaction with CuII, with an alternating [‐metal‐vacancy‐]n pattern, along the N‐terminal edge. Metal coordination does not significantly distort the intra‐β‐strand peptide interactions, which provides a possible explanation for the acceleration of Ac‐Aβ(13–21)H14A fibrillization by CuII, through stabilization of the associated state and low‐reorganization integration of β‐strand peptide pair precursors.