Research

Millennium Nucleus in
NanoBioPhysics

Research for a responsible development of nanotechnology

N²BP

Responsible development of nanotechnology requires understanding nano-bio interactions at a fundamental level. Our research team combines materials science, molecular simulation and biophysics with an emphasis on statistical mechanics, quantitative experiments and simulation. We study nanoscale systems, in particular, the interaction between nanoparticles, membrane channels, small polar solvents and proteins. The overall goal of N2BP is to understand how structure and function emerge in nano-confined geometries, unravel the nanobio interaction and its effects on membrane channels.

Carbon nanotubes

The thermodynamics of nanoconfinement will be determined using carbon nanotubes (CNTs) as simple models of extreme confinement.

The smallest systems in which we have observed the appearance of structures are those of water within narrow nanotubes. Water loading is due to confinement and cooperative dipole-dipole interactions, resulting in spatial correlations and structure. Few studies have explored and generalized this phenomenon to other polar molecules. Through quantum and classical molecular simulations we investigated this process for small alcohols inside nanotubes.

Our models are calibrated by comparing with contact angle (CA) measurements of graphene-water and graphene-alcohol interfaces. Different CA values are reported in the literature, so we grow pristine graphene for accurate CA measurements.

In addition to the principal investigators, our team includes 2 post-docs and undergraduate and graduate students from different programs. Their research at N2BP not only trains them as experts in their fields, but also allows them to develop multidisciplinary research, which is essential for solving complex problems.

Aquaporins

Aquaporins (AQP) serve as an experimental measure of confined water and alcohols.

AQPs are membrane proteins that form nanopores that transport water through cells. We have molecular dynamic (MD) simulation evidence indicating that depolarization produces a structural change in the Arginine-Histidine pair in the selectivity filter (SF) in h-AQP4, suggesting that this change could be measured as an activation current. Through patch clamp assays we sought to test this prediction and explore whether this is a general mechanism in AQP from different kingdoms, given the high conservation of SF.

In particular, we will study the dipolar alignment of SF. We already have preliminary data on AQP activation currents in humans and plants and have observed different activation kinetics exhibiting different water permeabilities. We then hypothesize that the movement of water through the SF is affecting the current kinetic gating, essentially making the gating currents a measure of nanoconfined water.

AQP can also transport small alcohols, so we will explore whether gating currents can be used to study small alcohol confinements. In general, we seek to determine voltage sensing in AQP and explore its use as a tool for measuring nanoconfined solvents.

Hv1 proton channels

Intracellular pH-sensitive, voltage-gated proton channels (Hv1) extrude protons when their intracellular concentration increases. The structure of Hv1 is similar to the voltage-sensing domain (VSD) of other ion channels, but lacks the pore domain. The Hv1 channel has 4 transmembrane segments and it has been proposed that the S4 segment is the VSD that upon depolarization triggers structural changes that allow H+ conduction. We sought to unravel the molecular mechanism of the electromechanical coupling between S4 and H+ transport. We have molecular dynamic (MD) simulation results showing that, for active states, the channel increases its permeability to water, suggesting that water and H+ transport are intimately coupled. We then hypothesized that activation currents should also be coupled to osmotic pressure differences, similar to the delta pH dependence of voltage sensing.

MD studies predict that water structuring also influences H+ transport so we will further explore this aspect through quantum and classical molecular simulation.

Thus, we sought to establish a functional role for nanoconfined water in the Hv1 channel.

Amino acid interaction with carbon nanostructures

Carbon nanostructures (CBN) such as nanotubes, graphene and fullerenes have been proposed for various applications. The general consensus is that, given their size and nonspecificity, CBNs interact with proteins and membranes via noncovalent interactions with aromatic and hydrophobic groups.
The interaction of amino acids with CBN is essential for understanding the protein-CBN interface. We have current molecular dynamic (MD) simulation data on the free energy (DG) of physisorption of the 20 amino acids in graphene. We will extend these studies to the quantum molecular (QM) simulation level, since the polarization of pi electron clouds is not included in classical models.

Arginines (R) have been predicted to interact strongly with graphene, a non-trivial result due to the positive charge of R. We sought to validate these studies by atomic force microscopy physisorption assays and chromatography. These experiments will be performed at different temperatures to estimate the entropic and enthalpic contributions of DG physisorption.

In this way we seek to unravel the amino acid-CBN interaction, in particular the R-CBN pair, and to gain insight into the DG and atomistic forces that determine these interactions.

Effects of nanotubes on voltage sensors

Arginines, which in many cases are the molecular determinants of the voltage sensing domain (VSD), interact strongly with carbon nanostructures, so we sought to determine the arginine-nanotube interaction and its consequences on ion channel voltage sensors such as Hv1.

The VSDs are placed inside the membrane core, making them accessible to the hydrophobic carbon nanotube (CBN). Consequently, it is plausible that CBNs interact with VSDs, through arginines (R), and perturb voltage sensing. Thus, the HV1 channel becomes the ideal test system, since it is a VSD that conducts protons with R in it. Through activation currents of HV1 preincubated with CBN, in combination with binding physisorption free energy (DG) calculations, we explored whether CBNs can bind VSD through R and modify voltage sensing.

The selectivity filter (SF) in AQP also contains an R, therefore, through the use of CBN, we will unravel the role of arginine in voltage sensing of h-AQP4. Through this objective, we hope to pave the way towards the modulation of membrane channels by CBN.

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We use the facilities of the Surface Physics Laboratory, located in the Physics Department of the Universidad Técnica Federico Santa María, Valparaíso. This allows us to have equipment for the growth and characterization of nanomaterials through different experimental techniques.

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