Change In Net Charge Research Articles

Relative Binding Affinity Prediction of Charge-Changing Sequence Mutations with FEP in Protein–Protein Interfaces

Building on the substantial progress that has been made in using free energy perturbation (FEP) methods to predict the relative binding affinities of small molecule ligands to proteins, we have previously shown that results of similar quality can be obtained in predicting the effect of mutations on the binding affinity of protein–protein complexes. However, these results were restricted to mutations which did not change the net charge of the side chains due to known difficulties with modeling perturbations involving a change in charge in FEP. Various methods have been proposed to address this problem. Here we apply the co-alchemical water approach to study the efficacy of FEP calculations of charge changing mutations at the protein–protein interface for the antibody–gp120 system investigated previously and three additional complexes. We achieve an overall root mean square error of 1.2 kcal/mol on a set of 106 cases involving a change in net charge selected by a simple suitability filter using side-chain predictions and solvent accessible surface area to be relevant to a biologic optimization project. Reasonable, although less precise, results are also obtained for the 44 more challenging mutations that involve buried residues, which may in some cases require substantial reorganization of the local protein structure, which can extend beyond the scope of a typical FEP simulation. We believe that the proposed prediction protocol will be of sufficient efficiency and accuracy to guide protein engineering projects for which optimization and/or maintenance of a high degree of binding affinity is a key objective.

Heat-induced aggregation and gelation of proteins from edible honey bee brood (Apis mellifera) as a function of temperature and pH

Edible insects offer a huge potential as a new source of proteins for food and feed and as a promising functional agent. In this regard, our study aimed to evaluate the effect of pH and temperature on aggregation and gelation of honey bee brood (Apis mellifera). Aggregation of soluble proteins was dependent on temperature and pH value and the highest coagulation was reached at 85 °C at pH 5 and 7 (73.7 and 68.4%, respectively). Changes in protein surface hydrophobicity, net charge and amount of available and buried sulfhydryl groups demonstrated the contribution of either covalent or non-covalent intermolecular interactions resulted in aggregation at various pH. The least gelation concentration of raw powder from honey bee brood varied from 5 to 11% on raw powder basis at pH 7 and 3, respectively. Moreover, the effect of pH on rheological and textural properties, as well as gel microstructure from honey bee brood is discussed. Therefore, freeze dried raw powder of honey bee brood has rich nutritional value and demonstrated gelation concentration comparable to conventional protein sources that reveal its potential as an ingredient for insect-based gel product.

Dependence of interfacial properties of p-Cu2O/electrolyte and p-Cu2O/Au junctions on the electrodeposition bath pH of p-Cu2O films

Improvements in electrodeposited cuprous oxide (Cu2O) junction devices could contribute to development of a wider range of affordable and environmentally friendly electronic devices. In this regard, ability of controlling interfacial properties of contacts of Cu2O thin films with other suitable materials will be a very important property. A study on interfacial properties of p-Cu2O/Au and p-Cu2O/electrolyte junctions was conducted using capacitance-voltage and current-voltage measurements. It was found that pH of p-Cu2O film deposition bath has a significant effect on junction properties of Cu2O. Particularly, flat band potential at p-Cu2O/electrolyte interface and built-in potential at p-Cu2O/Au interface were shifted positively by the increase of deposition bath pH of p-Cu2O films. Also, it was revealed that the shift was more pronounce at p-Cu2O/Au interface compared to that at p-Cu2O/electrolyte interface. The observed shifts in flat band potential and built in potentials were correlated with the shifts in relative band edge positions at the interfaces and with the change in net surface charge on the interfacial layers. It was observed, for the first time, that built in potential at p-Cu2O/Au junction can be controlled with bath pH of p-Cu2O films so that the junction can make either ohmic or non ohmic. Further, it was found that surface modification of p-Cu2O films with sulphidation reduces barrier height at p-Cu2O/Au junction and make the contact ohmic irrespective of bath pH of p-Cu2O films. We report here, for the first time, that relative band edge position at interfaces of electrodeposited p-Cu2O junctions can be controlled with pH of film deposition bath and surface modification. This will be very useful in application of electrodeposited p-Cu2O thin films in solar cells and other junction devices.

Metal-catalyzed oxidation of human serum albumin does not alter the interactive binding to the two principal drug binding sites

It is well known that various physiological factors such as pH, endogenous substances or post-translational modifications can affect the conformational state of human serum albumin (HSA). In a previous study, we reported that both pH- and long chain fatty acid-induced conformational changes can alter the interactive binding of ligands to the two principal binding sites of HSA, namely, site I and site II. In the present study, the effect of metal-catalyzed oxidation (MCO) caused by ascorbate/oxygen/trace metals on HSA structure and the interactive binding between dansyl-L-asparagine (DNSA; a site I ligand) and ibuprofen (a site II ligand) at pH 6.5 was investigated. MCO was accompanied by a time-dependent increase in carbonyl content in HSA, suggesting that the HSA was being oxidized. In addition, The MCO of HSA was accompanied by a change in net charge to a more negative charge and a decrease in thermal stability. SDS-PAGE patterns and α-helical contents of the oxidized HSAs were similar to those of native HSA, indicating that the HSA had not been extensively structurally modified by MCO. MCO also caused a selective decrease in ibuprofen binding. In spite of the changes in the HSA structure and ligand that bind to site II, no change in the interactive binding between DNSA and ibuprofen was observed. These data indicated that amino acid residues in site II are preferentially oxidized by MCO, whereas the spatial relationship between sites I and II (e.g. the distance between sites), the flexibility or space of each binding site are not altered. The present findings provide insights into the structural characteristics of oxidized HSA, and drug binding and drug-drug interactions on oxidized HSA.

Effect of Bath pH on Interfacial Properties of Electrodeposited n‐Cu2O Films

For developing semiconductor junction devices with low cost electrodeposited cuprous oxide (Cu2O), it will be very useful to investigate possibilities to improve the interface properties of Cu2O with suitable junction materials. We have investigated the effect of electrodeposition bath pH on the interfacial properties of n‐Cu2O/electrolyte, n‐Cu2O/Au, and n‐Cu2O/p‐Cu2O junctions by exploiting capacitance–voltage and photoresponse measurements. In addition, XRD and SEM measurements were also employed for thin film characterization. We have observed that for an increase in bath pH from 5.7 to 6.5 the resulted n‐Cu2O films produced a shift in the extrapolated flat band potential at the aqueous electrolyte interface by 300 mV. In addition, we have observed that extrapolated built‐in potentials at n‐Cu2O/Au and n‐Cu2O/p‐Cu2O interfaces are also affected by the bath pH. With the change in bath pH, the shift in the flat band potential at n‐Cu2O/electrolyte interface is completely opposite to the built‐in potential shift at n‐Cu2O/Au interface. We explained the observation as a result of the change in net charge at the interfacial surface layers of the interfaces, due to the change in bath pH of the films. Evidence in support of the explanation was further established by the observation that the built‐in potential at n‐Cu2O/Au interface is increased due to the surface modification of n‐Cu2O films with S2− ions. In addition, bath pH could be optimized in the fabrication process of Cu2O homojunctions to yield high photoactivity. In comparison with the best n‐Cu2O/Au Schottky junction, we could fabricate a Cu2O homojunction with a 590% increase in Voc and an 800% increase in Isc. All the experimental evidences directed toward the idea that the relative band edge position of n‐Cu2O at the interfaces is affected by the bath pH. The results reported in this investigation will be very useful in the applications of electrodeposited Cu2O films in photocataytic, solar cell, and other electronic devices.

Cationic (Charge Shift) Exciplexes.

Of the many known examples of exciplexes, those formed from bimolecular encounter between a cationic, excited state electron acceptor and a neutral donor in fluid media have not been previously reported. We now show that emissive exciplexes formed from excited N-methyl isoquinolinium cation (NMiQ+) with alkyl benzene donors are readily detected in acetonitrile. These cationic exciplexes result in a charge shift (A+* + D → A•D•+) with no net change in charge, which differs fundamentally from the charge-generation of conventional exciplex formation (A* + D → A•-D•+). We find that cationic and conventional exciplexes show similar trends, e.g., bathochromic shifts and decreases in fluorescence quantum yields with decreasing oxidation potentials of the donors. In the presented examples of NMiQ+ exciplexes, the fluorescence quantum yield decreases by a factor of 30 and the radiative rate constant by 6.6 as the fractional CT character of the exciplex increases from ∼0.79 to ∼0.95. Interestingly, the electronic coupling matrix elements for the NMiQ+ exciplexes, derived from a correlation of the radiative rate constants with the average emission frequencies, are similar to those of related conventional exciplexes, in spite of the absence of Coulombic stabilization in the cationic exciplexes.

Stimuli responsive charge-switchable lipids: Capture and release of nucleic acids

Stimuli responsive lipids, which enable control over the formation, transformation, and disruption of supramolecular assemblies, are of interest for biosensing, diagnostics, drug delivery, and basic transmembrane protein studies. In particular, spatiotemporal control over a supramolecular structure can be achieved using light activated compounds to induce significant supramolecular rearrangements. As such, a family of cationic lipids are described which undergo a permanent switch in charge upon exposure to 365nm ultraviolet (UV) light to enable the capture of negatively charged nucleic acids within the self-assembled supramolecular structure of the lipids and subsequent release of these macromolecules upon exposure to UV light and disruption of the assemblies. The lipids are composed of either two different tripeptide head groups, Lysine–Glycine–Glycine (KGG) and Glycine–Glycine–Glycine (GGG) and three different hydrocarbon chain lengths (C6, C10, or C14) terminated by a UV light responsive 1-(2-nitrophenyl)ethanol (NPE) protected carboxylic acid. The photolysis of the NPE protected lipid is measured as a function of time, and the resulting changes in net molecular charge are observed using zeta potential analysis for each head group and chain length combination. A proof of concept study for the capture and release of both linear DNA (calf thymus) and siRNA is presented using an ethidium bromide quenching assay where a balance between binding affinity and supramolecular stability are found to be the key to optimal nucleic acid capture and release.

Partial oxidation of step-bound water leads to anomalous pH effects on metal electrode step-edges.

The design of better heterogeneous catalysts for applications such as fuel cells and electrolyzers requires a mechanistic understanding of electrocatalytic reactions and the dependence of their activity on operating conditions such as pH. A satisfactory explanation for the unexpected pH dependence of electrochemical properties of platinum surfaces has so far remained elusive, with previous explanations resorting to complex co-adsorption of multiple species and resulting in limited predictive power. This knowledge gap suggests that the fundamental properties of these catalysts are not yet understood, limiting systematic improvement. Here, we analyze the change in charge and free energies upon adsorption using density-functional theory (DFT) to establish that water adsorbs on platinum step edges across a wide voltage range, including the double-layer region, with a loss of approximately 0.2 electrons upon adsorption. We show how this as-yet unreported change in net surface charge due to this water explains the anomalous pH variations of the hydrogen underpotential deposition (Hupd) and the potentials of zero total charge (PZTC) observed in published experimental data. This partial oxidation of water is not limited to platinum metal step edges, and we report the charge of the water on metal step edges of commonly used catalytic metals, including copper, silver, iridium, and palladium, illustrating that this partial oxidation of water broadly influences the reactivity of metal electrodes.

Characterization of the Dielectric Constant in the Trichoderma reesei Cel7B Active Site.

An attempt is made to evaluate the dielectric constant of the Trichoderma reesei Cel7B active site. Through kinetic measurements, the pKa value of the catalytic acid E201 is determined. Mutations (away from E201) with net charge changes are introduced to perturb the E201 pKa. It is shown that the mutation with a +1 charge change (including G225R, G230R, and A335R) decreases the pKa of E201, whereas the mutation with a -1 charge change (including Q149E, A222D, G225D, and G230D) increases the pKa. This effect is consistent with the electrostatic interaction between the changed charge and the E201 side chain. The fitting of the experimental data yields an apparent dielectric constant of 25-80. Molecular dynamics simulations with explicit water molecules indicate that the high solvent accessibility of the active site contributes largely to the high dielectric constant. ONIOM calculations show that high dielectric constant benefits the catalysis through decreasing the energy of the transition state relative to that of the enzyme substrate complex.

Novel silicon-doped, silicon and nitrogen-codoped carbon nanomaterials with high activity for the oxygen reduction reaction in alkaline medium

Si-doped and Si, N-codoped carbon nanomaterials exhibited much higher electrocatalytic activity and longer-term stability for the oxygen reduction reaction than non-doped carbon nanomaterials, due to the changes in net charges and the energy characteristics of the Si-containing carbon framework and the adsorption mode of oxygen.

Determination of the second virial coefficient of bovine serum albumin under varying pH and ionic strength by composition-gradient multi-angle static light scattering.

Composition-gradient multi-angle static light scattering (CG-MALS) is an emerging technique for the determination of intermolecular interactions via the second virial coefficient B22. With CG-MALS, detailed studies of the second virial coefficient can be carried out more accurately and effectively than with traditional methods. In addition, automated mixing, delivery and measurement enable high speed, continuous, fluctuation-free sample delivery and accurate results. Using CG-MALS we measure the second virial coefficient of bovine serum albumin (BSA) in aqueous solutions at various values of pH and ionic strength of a univalent salt (NaCl). The systematic variation of the second virial coefficient as a function of pH and NaCl strength reveals the net charge change and the isoelectric point of BSA under different solution conditions. The magnitude of the second virial coefficient decreases to 1.13 x 10(-5) ml*mol/g(2) near the isoelectric point of pH 4.6 and 25 mM NaCl. These results illuminate the role of fundamental long-range electrostatic and van der Waals forces in protein-protein interactions, specifically their dependence on pH and ionic strength.

Interaction of C1 Molecules with a Pt Electrode at Open Circuit Potential: A Combined Infrared and Mass Spectroscopic Study

The interaction of CO, HCOOH, HCHO, and CH3OH molecules with a Pt surface initially covered with a layer of oxide under open circuit potential (OCP) is studied by combined infrared and mass spectroscopy. We found that after switching to the fuel containing solution and concomitantly switching off the electrode potential control at 1.2 V, (i) the OCP decays from 1.2 V down to values of 0.58 V (CO), 0.12 V (HCOOH), 0.08 V(HCHO) and 0.24 V(CH3OH); (ii) CO is the only adsorbate formed at the Pt surface from the fuels; (iii) the rates for the decay of OCP and for the buildup of COad adlayer decrease in the order of CO > HCHO > HCOOH > CH3OH; (iv) the rate of CO2 production and the total amount of CO2 produced decreases in the order of CO > HCOOH > HCHO > CH3OH; and (v) a significant amount of HCOOH is formed for the case with HCHO and the main by product from CH3OH is HCHO. Our results indicate that (i) the change in OCP is determined by the change of net charge at the electrode/electrolyte interface due to the production of electrons from fuel oxidation and the consumption of electrons by Oad/OHad reduction; and (ii) even without an externally potential control, the reactions occurring at the interface are controlled by the electrochemical potential of the respective reactants and products. The implication from OCP transient to the local potential distribution and local activity in practical fuel cells are discussed.

Mapping disease‐related missense mutations in the immunoglobulin‐like fold domain of lamin A/C reveals novel genotype–phenotype associations for laminopathies

Mutations in A-type nuclear lamins cause laminopathies. However, genotype-phenotype correlations using the 340 missense mutations within the LMNA gene are unclear: partially due to the limited availability of three-dimensional structure. The immunoglobulin (Ig)-like fold domain has been solved, and using bioinformatics tools (including Polyphen-2, Fold X, Parameter OPtimized Surfaces, and PocketPicker) we characterized 56 missense mutations for position, surface exposure, change in charge and effect on Ig-like fold stability. We find that 21 of the 27 mutations associated with a skeletal muscle phenotype are distributed throughout the Ig-like fold, are nonsurface exposed and predicted to disrupt overall stability of the Ig-like fold domain. Intriguingly, the remaining 6 mutations clustered, had higher surface exposure, and did not affect stability. The majority of 9 lipodystrophy or 10 premature aging syndrome mutations also did not disrupt Ig-like fold domain stability and were surface exposed and clustered in distinct regions that overlap predicted binding pockets. Although buried, the 10 cardiac mutations had no other consistent properties. Finally, most lipodystrophy and premature aging mutations resulted in a -1 net charge change, whereas skeletal muscle mutations caused no consistent net charge changes. Since premature aging, lipodystrophy and the subset of 6 skeletal muscle mutations cluster tightly in distinct, charged regions, they likely affect lamin A/C -protein/DNA/RNA interactions: providing a consistent genotype-phenotype relationship for mutations in this domain. Thus, this subgroup of skeletal muscle laminopathies that we term the 'Skeletal muscle cluster', may have a distinct pathological mechanism. These novel associations refine the ability to predict clinical features caused by certain LMNA missense mutations.

Net charge changes in the calculation of relative ligand-binding free energies via classical atomistic molecular dynamics simulation.

The calculation of binding free energies of charged species to a target molecule is a frequently encountered problem in molecular dynamics studies of (bio-)chemical thermodynamics. Many important endogenous receptor-binding molecules, enzyme substrates, or drug molecules have a nonzero net charge. Absolute binding free energies, as well as binding free energies relative to another molecule with a different net charge will be affected by artifacts due to the used effective electrostatic interaction function and associated parameters (e.g., size of the computational box). In the present study, charging contributions to binding free energies of small oligoatomic ions to a series of model host cavities functionalized with different chemical groups are calculated with classical atomistic molecular dynamics simulation. Electrostatic interactions are treated using a lattice-summation scheme or a cutoff-truncation scheme with Barker–Watts reaction-field correction, and the simulations are conducted in boxes of different edge lengths. It is illustrated that the charging free energies of the guest molecules in water and in the host strongly depend on the applied methodology and that neglect of correction terms for the artifacts introduced by the finite size of the simulated system and the use of an effective electrostatic interaction function considerably impairs the thermodynamic interpretation of guest-host interactions. Application of correction terms for the various artifacts yields consistent results for the charging contribution to binding free energies and is thus a prerequisite for the valid interpretation or prediction of experimental data via molecular dynamics simulation. Analysis and correction of electrostatic artifacts according to the scheme proposed in the present study should therefore be considered an integral part of careful free-energy calculation studies if changes in the net charge are involved. © 2013 The Authors Journal of Computational Chemistry Published by Wiley Periodicals, Inc.

Changes in helical content or net charge of apolipoprotein C-I alter its affinity for lipid/water interfaces

Amphipathic α-helices mediate binding of exchangeable apolipoproteins to lipoproteins. To probe the role of α-helical structure in protein-lipid interactions, we used oil-drop tensiometry to characterize the interfacial behavior of apolipoprotein C-I (apoC-I) variants at triolein/water (TO/W) and 1-palmitoyl-2-oleoylphosphatidylcholine/triolein/water (POPC/TO/W) interfaces. ApoC-I, the smallest apolipoprotein, has two amphipathic α-helices. Mutants had single Pro or Ala substitutions that resulted in large differences in helical content in solution and on phospholipids. The ability of apoC-I to bind TO/W and POPC/TO/W interfaces correlated strongly with α-helical propensity. On binding these interfaces, peptides with higher helical propensity increased surface pressure to a greater extent. Likewise, peptide exclusion pressure at POPC/TO/W interfaces increased with greater helical propensity. ApoC-I retention on TO/W and POPC/TO/W interfaces correlated strongly with phospholipid-bound helical content. On compression of these interfaces, peptides with higher helical content were ejected at higher pressures. Substitution of Arg for Pro in the N-terminal α-helix altered net charge and reduced apoC-I affinity for POPC/TO/W interfaces. Our results suggest that peptide-lipid interactions drive α-helix binding to and retention on lipoproteins. Point mutations in small apolipoproteins could significantly change α-helical propensity or charge, thereby disrupting protein-lipid interactions and preventing the proteins from regulating lipoprotein catabolism at high surface pressures.

Comparison on effect of hydrophobicity on the antibacterial and antifungal activities of α-helical antimicrobial peptides

HPRP-A1, a 15-mer α-helical cationic peptide, was derived from N-terminus of ribosomal protein L1 (RpL1) of Helicobacter pylori. In this study, HPRP-A1 was used as a framework to obtain a series of peptide analogs with different hydrophobicity by single amino acid substitutions in the center of nonpolar face of the amphipathic helix in order to systematically study the effect of hydrophobicity on biological activities of α-helical antimicrobial peptides. Hydrophobicity and net charge of peptides played key roles in the biological activities of these peptide analogs; HPRP-A1 and peptide analogs with relative higher hydrophobicity exerted broad spectrum antimicrobial activity against Gram-negative bacteria, Gram-positive bacteria and pathogenic fungi, but also showed stronger hemolytic activity; the change of hydrophobicity and net charge of peptides had similar effects with close trend and extent on antibacterial activities and antifungal activities. This indicated that there were certain correlations between the antibacterial mode of action and the antifungal mode of action of these peptides in this study. The peptides exhibited antimicrobial specificity for bacteria and fungi, which provided potentials to develop new antimicrobial drugs for clinical practices.

Asbestos increases mitochondrial Ca2+ and H2O2 generation in THP1 cells

Asbestosis is an important cause of pulmonary fibrosis. We have shown that hydrogen peroxide (H2O2) released from macrophages contributes to the genesis of fibrosis. Because mitochondrial calcium (Ca2+) influx can increase mitochondrial H2O2 production, we examined if Ca2+ influx and H2O2 generation could be altered in macrophages exposed to asbestos. Using THP1 cells loaded with the Ca2+‐sensitive fluorescent dye Fura‐2, we found that intracellular Ca2+ increased by ~2 fold within 4–8 min after asbestos exposure and remained elevated for 60 min. Cells loaded with MitoTracker Red plus Ca2+‐sensitive Fluo3 (or Ca2+‐indicator Rhod2) showed that mitochondrial Ca2+levels were elevated within min after asbestos exposure. Since mitochondrial Ca2+ uptake is driven by a negative mitochondrial membrane potential (ΔΨ), we evaluated ΔΨ in cells loaded with JC‐1. We found that ΔΨ was reduced by ~25–50% in cells exposed to asbestos, a finding consistent with expected net charge changes associated with mitochondrial Ca2+ entry. Finally, to evaluate the role of mitochondrial Ca2+ entry on H2O2 generation, we treated cells with Ru360, an inhibitor of mitochondrial Ca2+ uniporter (MCU), and found that Ru360 reduced H2O2 generation in cells exposed to asbestos. These findings suggest that asbestos rapidly increases mitochondrial Ca2+ via the MCU, which contributes to mitochondrial H2O2 generation in macrophages. (Supported by 2R01ES015981–06, R01ES014871, VA Merit Review 1BX001135 to A.B.C.)

Engineering Myosin V Velocity by Tuning Nanocomposite Morphology

Devices which utilize biological components such as molecular motors may be capable of detecting or separating analytes with speed and resolution rivaling macroscopic instruments. The design and manufacturing of such devices requires detailed understanding of self-assembly and interactions of biological materials with synthetic environments. We demonstrated that nano-scale roughness modulates surface binding of F-actin (Langmuir 28:12216; 2012). By controllably converting a thin film of polystyrene-random-tert butyl acrylate copolymer to polyacrylic acid (PAA), we grafted amine-functionalized nanoparticles at high coverage to the PAA hydrogel, creating a surface with tunable roughness. Extending that approach here, we explore the effect of nanocomposite morphology on myosin V processive motility. We find that tuning the size and density of the surface grafted nanoparticles controls the velocity and run length of myosin V. Velocity decreases significantly as the nanoparticle size is increased independent of the concentration of MgATP. This effect of particle size may be due to changes in net charge, nanoparticle curvature at the interaction sites, or tortuosity of the actin imposed by the surface features. Experiments to distinguish these hypotheses are underway. Further understanding of the relationship between nanoparticle surface morphology and activity of actin in support of motility may permit the engineering of stops or stalls with high spacial precision into biomolecular devices. Supported by NSF/NSEC grant DMR08-32802.

A protein kinase assay based on FRET between quantum dots and fluorescently-labeled peptides

A novel protein kinase assay was developed, based on FRET between QDs and fluorescently-labeled substrate peptides. The negatively charged QDs recognize the change in net charge of the peptide upon phosphorylation. Despite its simple mechanism, this assay is sensitive and robust enough to be applied to the evaluation of protein kinase inhibitors.

Detection of real-time dynamics of drug–target interactions by ultralong nanowalls

Detecting drug-target interactions in real-time is a powerful approach for drug discovery and analytics. We show here for the first time the ultra fast electrical real-time detection and quantification of antibiotics using a novel biohybrid nanosensor. The biomolecular sensing is performed on ultralong (mm range) high aspect ratio nanowall (50 nm width) surfaces functionalized with operator DNA tetO which is specifically bound by the sensor protein TetR. This sensor protein is released from the operator DNA in a dose dependent manner by exposing the device functionalized with this bound DNA-protein complex to tetracycline antibiotics. As a result, the electrical conductance is accordingly modulated by these surface net charge changes. The switching mechanism of sensor proteins attached at the functionalized surfaces and releasing them again by antibiotics is demonstrated. With the here presented device the detection limit is below the limits of prevailing detection methods. Moreover, the study is extended to detect antibiotic residues in spiked organic milk from cows far below the maximum residual level of the European Union. In spiked milk samples a detection limit for tetracycline concentrations in the 100 fM level was achieved. The nanowall devices are fabricated by atomic layer deposition-based spacer lithography on full wafer scale which is a simple approach capable for mass production.