Abstract
Peptides—short chains of amino acids—can self-assemble into ordered nanostructures with applications ranging from skin repair to neurotransmitters. Here we examine a minimalist, alternating sequence composed of aspartic acid (D) and tyrosine (Y). Three structures—DY 6x2, DY 12x2, and DY 24x2—were built and simulated with YASARA to examine how chain length controls aggregation behaviour. Our computer simulations showed that the 24‑amino‑acid structure quickly locks into strong, criss‑crossed Beta‑sheets, while the shorter structure stays floppy and unstable. In conclusion, a chain of approximately 24 residues is required for the DY peptide to form a stable structure.
Introduction
Peptides are short polymers that are typically thought to have fewer than fifty amino acid residues. Thus, they hold a strategic midpoint between small molecules and highly folded proteins. They are sufficiently large to display several precisely positioned functional groups but small enough to be synthesized quickly by automated solid‑phase chemistry or by ribosomal expression systems. This dual nature gives scientists excellent control: by modifying a single residue, they can modulate charge distribution, hydrophobic balance, stereochemistry, or the presence of recognition motifs with atomic precision. Such functional diversity underlies their extraordinary diversity of natural functions. Hormonal peptides such as insulin and glucagon regulate blood-glucose levels; neuropeptides such as oxytocin and vasopressin control behaviour and water balance; antimicrobial peptides lyse bacterial membranes on mucosal surfaces; and tripeptides such as glutathione maintain intracellular red-ox homeostasis. Reflected in this value, the global peptide-therapeutics market surpassed US $50 billion in 2024 and is expanding at a double-digit compound rate.
Peptide chemistry advances have also driven a bio-nanotechnology revolution. Every peptide backbone is punctuated by regularly positioned donors and acceptors for hydrogen bonding, and the twenty standard side chains provide a combinatorial palette of charges, aromatic rings, aliphatic groups, and heteroatoms. This chemical grammar permits even very short sequences to spontaneously fold into higher-order structures such as α‑helices, antiparallel β‑sheets, nanotubes, twisted ribbons, and liquid coacervates. These supramolecular scaffolds translate molecular recognition into macroscale function, forming shear‑thinning injectable hydrogels for wound healing and 3‑D cell culture, piezoelectric fibres for soft robotics, ion‑conductive channels for bioelectronics, and enzyme‑mimetic catalysts that operate in water. A thematic design strategy is sequence economy—the use of simple, repetitive motifs that reduce synthetic cost while enhancing the predictability of the final architecture.
Within this minimalist ethos, alternating charge aromatic dipeptides have emerged as workhorses. Motifs such as FKFE, DFEFK, and KLVFF succeed because Coulombic attraction (or repulsion) and π–π stacking interact to nucleate together parallel or antiparallel β‑sheets that develop further into fibrils. All of this notwithstanding, such advances and many possible dyads remain to be investigated. Here, we report on the aspartic‑acid/tyrosine couple, DY. Aspartic acid offers a negatively charged β‑carboxylate at physiological pH, and tyrosine offers an electron-rich phenolic ring that can participate in hydrogen bonding and aromatic stacking. Positioning these residues next to one another juxtaposes long-range electrostatic interaction with short-range π‑stacking, a combination likely to induce β‑sheet formation.
The DY motif also offers advantages. Chemically, its two side chains are orthogonal handles: tyrosines phosphorylate, iodinate or photo-cross-link, and aspartates chelate divalent metals and nucleate calcium phosphate, of value for osteogenic scaffolds. Physically, the significant discrepancy of their pKₐ values (≈ 3.9 for Asp and ≈ 10 for Tyr) renders them any DY assembly environmentally pH-sensitive, with reversible sol–gel transitions, which can be used for drug release. Catalytically, the proximity of an acidic carboxylate to an aromatic π‑system can template mineral phases or facilitate electron transfer, suggesting redox or photoactive material functions.
Yet even with these desirable properties, the minimum chain length required for a DY sequence to nucleate an ordered β‑sheet is not known. Establishing this value is more than a matter of intellectual curiosity: it defines whether a DY‑based formulation is a free‑flowing solution, a self-supporting hydrogel, or an insoluble precipitate, and hence its usefulness in injectable therapeutics, surface coating, or nano‑electronic devices.
Objective. The present study consequently raises a simple but unresolved question: How short can a DY peptide be and still fold into a structured β‑sheet in neat water? To address it, we designed and simulated three antiparallel dimers—(DY)₆, (DY)₁₂, and (DY)₂₄—spanning the 6‑ to 24‑residue range most relevant to functional biomaterials, and we examined their conformational pathways with a battery of structural metrics.
Computational Details
All Molecular Dynamics (MD) simulations were conducted using GROMACS 2022 program with the force field used was CHARMM36. MD simulations for all sequences of peptides were done at 200 ns. Initially energy was minimized on all simulations to ensure the stability of each structure. Then natural volume and temperature conditions were used for all simulations. After running MD, the representative structure of the simulation, using gcluster technique, clustering the structure with respect to root mean square mean square deviation (RMSD).
Methodology
Among the 190 conceivable dipeptide pairs, we selected the alternating aspartic‑acid/tyrosine motif—written (DY)ₙ—for three tightly linked reasons. Complementary chemistry: at physiological pH, Aspartate carries a negatively charged β‑carboxylate, whereas Tyrosine offers an electron‑rich aromatic ring capped by a mildly acidic phenolic hydroxyl; bringing these side chains into proximity marries long‑range Coulombic attraction with π–π stacking, a synergy that stabilises β‑sheet assemblies. pH responsiveness: the markedly different pKₐ values of the Asp carboxylate (~3.9) and the Tyr phenol (~10) render the sequence sensitive to external pH, enabling reversible switching between soluble and aggregated states. Versatile functional handles: Tyrosine residues can be phosphorylated, iodinated, or photo‑cross‑linked, while Asp can chelate metals or nucleate mineral phases—orthogonal chemistries that ease post‑assembly functionalisation for tissue‑engineering scaffolds and soft‑electronics devices.
Despite the rich literature on charge‑aromatic peptides such as FKFE or DFEFK, the minimal DY repeat remains underexplored. We therefore constructed three dimers differing only in chain length: (DY)₆ – 6 residues per chain (12 total), (DY)₁₂ – 12 residues per chain (24 total), and (DY)₂₄ – 24 residues per chain (48 total)
All dimers were placed in explicit water and simulated for 100 ns using the YASARA molecular‑dynamics engine.
Results and Discussion
The trajectories reveal a clear length threshold. The twenty‑four‑residue dimer snaps into a compact, criss‑crossed β‑sheet within the first twenty‑five nanoseconds and retains roughly sixty percent β‑sheet content thereafter. The twelve‑residue version forms smaller β‑patches that fluctuate in and out of register, averaging about thirty percent sheet, while the six‑residue construct remains disordered, displaying little more than transient turns. Root‑mean‑square deviation stabilises after roughly twenty nanoseconds for all systems, but the radius of gyration shrinks markedly with length, indicating progressively tighter packing. Hydrogen‑bond analysis supports the picture: the short dimer shares fewer than one inter‑chain hydrogen bond per snapshot, the intermediate length averages three, and the longest strand maintains more than seven, well above the five‑bond threshold usually cited for persistent β‑sheets.
Still images extracted from the simulations illustrate this behaviour. A single (DY)₆ strand, numbered for clarity, shows the alternating acidic and aromatic side chains that motivate the design. Side‑by‑side panels juxtapose the fully extended starting dimers with their final conformations: the (DY)₆ dimer stays largely intact; the (DY)₁₂ pair frays by two hundred nanoseconds; and the (DY)₂₄ assembly, although it folds rapidly, eventually splits into smaller sheet clusters after nearly five hundred nanoseconds.
We found that the 24‑amino‑acid structure quickly locks into strong, criss‑crossed Beta‑sheets, while the shorter structure stays floppy and unstable. This is an important finding since this information provides a rational guideline for choosing sequence length depending on the intended application: Drug‑delivery hydrogels require rapid gelation and thus benefit from a length above the β‑sheet threshold. Soluble peptide carriers for imaging probes may require lengths below the threshold to avoid premature aggregation. Bioelectronic films and wires may exploit the aromatic stacking of tyrosine once a stable sheet is formed. These long trajectories hint that repeat length is necessary but not sufficient for permanent integrity—mechanical strain accumulated in very long strands can drive partial disassembly over extended times. From a materials‑engineering standpoint, an intermediate window of roughly twelve to eighteen residues may offer the best compromise between fast nucleation and long‑term stability: long enough to form β‑sheets quickly, yet short enough to avoid brittle fracture or aggregation into insoluble masses.
Conclusion
This work identifies roughly twenty‑four residues as the critical length at which DY peptides jump from a floppy coil to a robust β‑sheet. Chemically, the alternating Asp–Tyr motif provides two orthogonal functional handles—carboxylates and phenolic rings—that can be phosphorylated, metal-chelated, iodinated, or photo-cross-linked, enabling precise post-assembly modification. Physically, the length‑triggered sheet formation translates directly into higher mechanical stiffness, lower radius of gyration, and a densely hydrogen‑bonded core, providing a tunable lever for tailoring hydrogel rigidity or film conductivity. Catalytically, the juxtaposed acidic and aromatic side chains create microenvironments capable of templating mineral phases, binding redox‑active metals, and potentially accelerating electron‑transfer reactions in soft‑electronic devices. Together, these chemical, physical, and catalytic insights furnish a clear design rule for crafting DY‑based hydrogels, soluble carriers, and bioelectronic films, while charting an experimental roadmap for tuning peptide length to match the desired material behavior.
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