An anopore-based Cu II-sensings ystem is reported that allows for an ultrasensitive and selective detection of Cu II with the possibility for ab road rangeo fa pplications, for example in medical diagnostics. Af luorescent ATCUN-like peptide5 /6-FAM-Dap-b-Ala-His is employedt o selectively bind Cu II ions in the presence of Ni II and Zn II and was crafted into ion track-etched nanopores. Upon Cu II binding the fluorescence of the peptides ensor is quenched, permitting the detection of Cu II in solution. The ion transport characteristics of peptide-modified nanopore are shown to be extremelys ensitivea nd selective towards Cu II allowing to sense femtomolarC u II concentrations in human urine mimics. Washing with EDTA fully restores the Cu II-binding properties of the sensor, enabling multipler epetitive measurements. The robustness of the system clearly has the potentialt ob ef urtherd eveloped into an easy-to-use, lab-on-chip Cu II-sensing device, which will be of great importance for bedside diagnosis and monitor of Cu II levelsi np atients with copper-dysfunctional homeostasis. Copper is an essential trace element that is inevitable in biological systems and can be found in many enzymes such as amine oxidases and ferroxidases but is also required for infant growth, the iron metabolism and brain developmenti nh uman organisms. [1] It is also widely used in agriculturals ystems, and therefore belongst oamajor metal pollutant in our environment. [2] There are also diseases linked to at oxic copper con-[a] Dr.

Nanopores comprise a versatile class of membrane proteins that carry out a range of key physiological functions and are increasingly developed for different biotechnological applications. Yet, a capacity to study and engineer protein nanopores by combinatorial means has so far been hampered by a lack of suitable assays that combine sufficient experimental resolution with throughput. Addressing this technological gap, the functional nanopore (FuN) screen now provides a quantitative and dynamic readout of nanopore assembly and function in the context of the inner membrane of Escherichia coli. The assay is based on genetically encoded fluorescent protein sensors that resolve the nanopore-dependent influx of Ca 2+ across the inner membrane of E. coli. Illustrating its versatile capacity, the FuN screen is first applied to dissect the molecular features that underlie the assembly and stability of nanopores formed by the S 21 68 holin. In a subsequent step, nanopores are engineered by recombining the transmembrane module of S 21 68 with different ring-shaped oligomeric protein structures that feature defined hexa-, hepta-, and octameric geometries. Library screening highlights substantial plasticity in the ability of the S 21 68 transmembrane module to oligomerize in alternative geometries, while the functional properties of the resultant nanopores can be fine-tuned through the identity of the connecting linkers. Overall, the FuN screen is anticipated to facilitate both fundamental studies and complex nanopore engineering endeavors with many potential applications in biomedicine, biotechnology, and synthetic biology.

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The development of flexible and reconfigurable sensors that can be readily tailored toward different molecular analytes constitutes a key goal and formidable challenge in biosensing. In this regard, synthetic nanopores have emerged as potent physical transducers to convert molecular interactions into electrical signals. Yet, systematic strategies to functionalize their surfaces with receptor proteins for the selective detection of molecular analytes remain scarce. Addressing these limitations, a general strategy is presented to immobilize nanobodies in a directional fashion onto the surface of track‐etched nanopores exploiting copper‐free click reactions and site‐specific protein conjugation systems. The functional immobilization of three different nanobodies is demonstrated in ligand binding experiments with green fluorescent protein, mCherry, and α‐amylase (α‐Amy) serving as molecular analytes. Ligand binding is resolved using a combination of optical and electrical recordings displaying quantitative dose–response curves. Furthermore, a change in surface charge density is identified as the predominant molecular factor that underlies quantitative dose–responses for the three different protein analytes in nanoconfined geometries. The devised strategy should pave the way for the systematic functionalization of nanopore surfaces with biological receptors and their ability to detect a variety of analytes for diagnostic purposes.

Cells of multicellular organisms are surrounded by and attached to a matrix of fibrous polysaccharides and proteins known as the extracellular matrix. This fibrous network not only serves as a structural support to cells and tissues but also plays an integral part in the process as important as proliferation, differentiation, or defense. While at first sight, the extracellular matrices of plant and animals do not have much in common, a closer look reveals remarkable similarities. In particular, the proteins involved in the adhesion of the cell to the extracellular matrix share many functional properties. At the sequence level, however, a surprising lack of homology is found between adhesion-related proteins of plants and animals. Both protein machineries only reveal similarities between small subdomains and motifs, which further underlines their functional relationship. In this review, we provide an overview on the similarities between motifs in proteins known to be located at the plant cell wall-plasma membrane-cytoskeleton interface to proteins of the animal adhesome. We also show that by comparing the proteome of both adhesion machineries at the level of motifs, we are also able to identify potentially new candidate proteins that functionally contribute to the adhesion of the plant plasma membrane to the cell wall.