Biology subjects

Grawe, A.

Publications and source records attributed to Grawe, A..

6 recordsLinked to original sources

bioRxiv2026

DIALECT: DNA-Programmable Synthetic Cells Communicate with Mammalian Cells through Synthetic Receptors

Synthetic cells can regulate mammalian signalling through the controlled release of diffusible protein ligands, but coupling programmable synthetic-cell senders to orthogonal, engineered mammalian receptors remains challenging. Here we present DIALECT (DNA-Instructed Assembly of Ligands for Engineered Cellular Transduction), a platform in which proteinosome-based synthetic cells translate sequence-defined DNA inputs into coiled-coil signalling ligand outputs for engineered mammalian receptors. DNA-ligand conjugates are localized within semipermeable proteinosomes through DNA hybridization and released by toehold-mediated DNA strand displacement (TMSD). The released ligands diffuse to neighbouring mammalian cells and activate cognate CC-GEMS receptors, inducing JAK/STAT3 signalling and reporter activation, with a higher response in cognate than non-cognate receiver cells under activation conditions. We further implement distributed Boolean AND logic using two independently programmed proteinosome populations that respond to distinct DNA inputs and release complementary ligand components. Simultaneous addition of both inputs produces substantially higher reporter activity than the respective single-input conditions, consistent with extracellular assembly of an active receptor agonist. DIALECT thus establishes a modular interface linking DNA-based molecular computation in synthetic cells to orthogonal signalling pathways in mammalian cells, providing a foundation for hybrid artificial-biological communication networks.

synthetic biology↗

bioRxiv2026

Laccase-mediated biotransformation potential for fluorinated compounds by geographically diverse human gut microbiota

The growing prevalence of synthetic organofluorine substances in agrochemicals, food packaging, and consumer products has led to increasing gastrointestinal exposure, with potential consequences for human health. Despite the extreme stability of fluorinated compounds, several microbial pathways for their transformation are known, including by laccases, a type of multicopper oxidase. However, the functionality of laccases in the gut microbiome, a natural contact point between food-associated chemicals and microbial biotransformation pathways, is poorly defined. Through a multi-study analysis of 1578 human gut metagenomes spanning a global gradient from hunter-gatherer societies to industrialized urban populations, we found that laccase-coding gene homologs are widely distributed in the human gut microbiome. We identified a significant association between both the abundance and phylogenetic diversity of laccase homologs with the degree of urbanization. As human gut microbial laccase activity has not been experimentally demonstrated, eight gut metagenome-derived laccases were heterologously expressed and screened for activity with a redox mediator system. Six of the eight laccases demonstrated activity. One of these gut microbial laccases and three previously characterized laccases were then tested for their capacity to deplete 11 different food-associated chemicals. All four effectively depleted the agrochemicals cyflumetofen and fluazinam, as well as the industrial chemical bisphenol AF to a lesser extent. By linking global gut metagenomes with activity assays, this work demonstrates the untapped potential of mining human gut metagenomes for laccases and other microbial enzymes that can actively modify various agricultural and industrial chemicals. ImportanceAs adverse effects of fluorinated compounds on human health are emerging, the responsible enzymes from the human gut microbiome of geographically diverse human cohorts mediating interactions with fluorinated compounds in the gastrointestinal tract remain poorly characterized. In a multi-study analysis of publicly available microbiome sequencing data, we linked the abundance, diversity, and phylogeny of laccases within the gut microbiome to the degree of urbanization of human cohorts, and experimentally demonstrated the ability of these laccases to deplete a range of food-associated fluorochemicals. Another significant contribution of our study is in the integration of rural catchment areas as a quantitative metric of urbanization in gut microbiome metagenomics and enzyme activity surveys.

microbiology↗

bioRxiv2025

Reversible Sandwich-Based Particle Nanoswitch for Continuous Protein Monitoring at Picomolar Concentrations with Automated Calibration

Continuous monitoring of specific proteins is essential for understanding the dynamics of biological systems and for enabling real-time measurement-and-control strategies in bioprocesses. Ideally, sensors for continuous monitoring should be intrinsically reversible and able to perform accurate measurements over long time spans. Here, we present a particle nanoswitch sensor containing two different antibody fragments that bind reversibly to a protein of interest and thus form transient sandwich complexes. The antibody fragments are incorporated into the sensor using site-specific conjugation strategies to achieve optimal antibody orientation. Short-lived sandwich complexes are detected with single-molecule resolution, by tracking the motion of tethered particles. The sensing concept is demonstrated for lactoferrin, an iron-binding and immune-modulating protein. We show continuous measurements of picomolar concentrations with a response time of [~]10 min over periods of 12-15 h. Automated calibration strategies are described that result in a mean absolute relative difference below 10% compared to reference measurements. These results demonstrate how continuous fast protein sensing at picomolar concentrations can be achieved using reversible sandwich-based particle nanoswitches, enabling long-term monitoring of dynamic bioprocesses.

bioengineering↗

bioRxiv2024

Direct and ultrasensitive bioluminescent detection of intact respiratory viruses

Respiratory viruses such as SARS-CoV-2, influenza, and respiratory syncytial virus (RSV) represent pressing health risks. Rapid diagnostic tests for these viruses detect single antigens or nucleic acids, which do not necessarily correlate with the amount of intact virus. Instead, specific detection of intact respiratory virus particles may better assess the contagiousness of a patient. Here, we report GLOVID, a modular biosensor platform to detect intact virions against a background of free viral proteins in solution. Our approach harnesses the multivalent display of distinct proteins on the surface of a viral particle to template the reconstitution of a split luciferase, allowing specific, single-step detection of intact influenza A and RSV virions corresponding to 0.1 - 0.3 fM of genomic units. The protein ligation system used to assemble GLOVID sensors is compatible with a broad range of binding domains, including nanobodies, scFv fragments and cyclic peptides, which allows straightforward adjustment of the sensor platform to target different viruses.

bioengineering↗

bioRxiv2023

Bioluminescent detection of viral surface proteins using branched multivalent protein switches

Fast and reliable virus diagnostics is key to prevent the spread of viruses in populations. A hallmark of viruses is the presence of multivalent surface proteins, a property that can be harnessed to control conformational switching in sensor proteins. Here, we introduce a new sensor platform (dark-LUX) for the detection of viral surface proteins consisting of a general bioluminescent framework that can be post-translationally functionalized with separately expressed binding domains. The platform relies on 1) plug-and-play bioconjugation of different binding proteins via SpyTag/SpyCatcher technology to create branched protein structures, 2) an optimized turn-on bioluminescent switch based on complementation of the split-luciferase NanoBiT upon target binding and 3) straightforward exploration of the protein linker space. The influenza A virus (IAV) surface proteins hemagglutinin (HA) and neuraminidase (NA) were used as relevant multivalent targets to establish proof of principle and optimize relevant parameters such as linker properties, choice of target binding domains and the optimal combination of the competing NanoBiT components SmBiT and DarkBiT. The sensor framework allows rapid conjugation and exchange of various binding domains including scFvs, nanobodies and de novo designed binders for a variety of targets, including the construction of a heterobivalent switch that targets the head and stem region of hemagglutinin. The modularity of the platform thus allows straightforward optimization of binding domains and scaffold properties for existing viral targets, and is well suited to quickly adapt bioluminescent sensor proteins to effectively detect newly evolving viral epitopes.

biochemistry↗

bioRxiv2020

Functional specialization of human salivary glands and origins of proteins intrinsic to human saliva

Salivary proteins are essential for maintaining health in the oral cavity and proximal digestive tract and serve as a diagnostic window into human disease. However, their precise organ origins remain unclear. Through transcriptomic analysis of major adult and fetal salivary glands, and integration with the saliva proteome and transcriptomes of 28+ organs, we linked human saliva proteins to their source, identified salivary gland-specific genes, and uncovered fetal- and adult-specific gene repertoires. Our results also provide new insights into the degree of gene retention during maturation and suggest that functional diversity between adult gland-types is driven by specific dosage combinations of hundreds of transcriptional regulators rather than a few gland-specific factors. Finally, we demonstrate the hitherto unrecognized heterogeneity of the human acinar cell lineage. Our results pave the way for future investigations into glandular biology and pathology, as well as salivas use as a diagnostic fluid.

developmental biology↗