Julián Valero

Binding to protein surfaces by supramolecular multivalent scaffolds.

Multivalency plays a pivotal role in biological recognition, particularly at protein-protein and protein-carbohydrate interaction sites. Scaffolds of diverse structure, flexibility, and valency are gaining increasing biomedical importance in the development of artificial multivalent ligands for these interfaces. Relevant examples range from small C(4) symmetric calix[4]arenes and porphyrin ligands, which may achieve nanomolar affinity for protein surfaces of pharmaceutical interest, to large-sized dendrimers that provide promising adherence-inhibition for toxins and other relevant lectins. In addition, highly flexible supramolecular platforms like rotaxanes and polymers have been proposed as challenging alternatives to more rigid designs. Finally, nanoparticles are being exploited for this aim as they present important advantages from the biological and synthetic points of view.

A tetraguanidinium macrocycle for the recognition and cavity expansion of calix[4]arene tetraoxoanions

Molecular containers have raised an increased interest over the last decade. These supramolecular architectures have found applications in catalysis, molecular sensing or as insulators for key intermediates, among others. In this study, we describe the synthesis and binding properties of a tetraguanidinium macrocycle which forms robust complexes with diverse calix[4]arene tetraoxoanions through hydrogen bonding and electrostatic interactions. The binding behaviour and affinity strength of these constructs have been measured by NMR and isothermal titration calorimetry. Besides, VT NMR experiments show that this novel cyclic tetracation is able to stabilise the cone conformation of these calix[4]arenes. Preliminary NMR-binding experiments between a tetraguanidinium calix[4]arenate and quinolinium or isoquinolinium salts suggest an effective increase in the cavity volume of these supramolecular constructs.

A bio-hybrid DNA rotor–stator nanoengine that moves along predefined tracks

Biological motors are highly complex protein assemblies that generate linear or rotary motion, powered by chemical energy. Synthetic motors based on DNA nanostructures, bio-hybrid designs or synthetic organic chemistry have been assembled. However, unidirectionally rotating biomimetic wheel motors with rotor–stator units that consume chemical energy are elusive. Here, we report a bio-hybrid nanoengine consisting of a catalytic stator that unidirectionally rotates an interlocked DNA wheel, powered by NTP hydrolysis. The engine consists of an engineered T7 RNA polymerase (T7RNAP-ZIF) attached to a dsDNA nanoring that is catenated to a rigid rotating dsDNA wheel. The wheel motor produces long, repetitive RNA transcripts that remain attached to the engine and are used to guide its movement along predefined ssDNA tracks arranged on a DNA nanotube. The simplicity of the design renders this walking nanoengine adaptable to other biological nanoarchitectures, facilitating the construction of complex bio-hybrid structures that achieve NTP-driven locomotion.The DNA nanoengine constitutes a chemical energy-driven unidirectional rotor that directionally walks along a predefined multistep track.

Single-Stranded Tile Stoppers for Interlocked DNA Architectures

Interlocked DNA architectures are useful for DNA nanotechnology because of their mechanically bonded components, which can move relative to one another without disassembling. We describe the design, synthesis, and characterization of novel single‐stranded tile (SST) stoppers for the assembly of interlocked DNA architectures. SST stoppers are shown to self‐assemble into a square‐shaped rigid structure upon mixing 97 oligodeoxynucleotide (ODN) strands. The structures are equipped with a sticky end that is designed for hybridization with the sticky ends of a dsDNA axle of a DNA rotaxane. Because the diameter of the macrocycle threaded onto the axle is 14 nm, the dimension of the square‐shaped stopper was designed to be bulky enough to prevent the dethreading of the macrocycle. An asymmetric rotaxane with a SST‐ and a ring‐shaped stopper featuring two stations for hybridization of the macrocycle to the axle was assembled. The macrocycle can be directed towards one or the other station upon triggering with fuel ODNs.

Cellular Antisense Activity of PNA–Oligo(bicycloguanidinium) Conjugates Forming Self-Assembled Nanoaggregates

A series of peptide nucleic acid–oligo(bicycloguanidinium) (PNA–BGn) conjugates were synthesized and characterized in terms of cellular antisense activity by using the pLuc750HeLa cell splice correction assay. PNA–BG4 conjugates exhibited low micromolar antisense activity, and their cellular activity required the presence of a hydrophobic silyl terminal protecting group on the oligo(BG) ligand and a minimum of four guanidinium units. Surprisingly, a nonlinear dose–response with an activity threshold around 3–4 μM, indicative of large cooperativity, was observed. Supported by light scattering and electron microscopy analyses, we propose that the activity, and thus cellular delivery, of these lipo‐PNA–BG4 conjugates is dependent on self‐assembled nanoaggregates. Finally, cellular activity was enhanced by the presence of serum. Therefore we conclude that the lipo‐BG‐PNA conjugates exhibit an unexpected mechanism for cell delivery and are of interest for further in vivo studies.