Detection of protein oligomers with nanopores
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Upcycling of polyamides through chemical hydrolysis and engineered Pseudomonas putida
Aliphatic polyamides, or nylons, are widely used in the textile and automotive industry due to their high durability and tensile strength, but recycling rates are below 5%. Chemical recycling of polyamides is possible but typically yields mixtures of monomers and oligomers which hinders downstream purification. Here, Pseudomonas putida KT2440 was engineered to metabolize C6-polyamide monomers such as 6-aminohexanoic acid, ε-caprolactam and 1,6-hexamethylenediamine, guided by adaptive laboratory evolution. Heterologous expression of nylonases also enabled P. putida to metabolize linear and cyclic nylon oligomers derived from chemical polyamide hydrolysis. RNA sequencing and reverse engineering revealed the metabolic pathways for these non-natural substrates. To demonstrate microbial upcycling, the phaCAB operon from Cupriavidus necator was heterologously expressed to enable production of polyhydroxybutyrate (PHB) from PA6 hydrolysates. This study presents a microbial host for the biological conversion, in combination with chemical hydrolysis, of polyamide monomers and mixed polyamids hydrolysates to a value-added product.
Geochemistry of lithospheric aqueous fluids modified by nanoconfinement
Water is a principal component of Earth’s fluids, and its interaction with rocks governs lithospheric geochemical and geodynamic processes. Water–rock interactions are crucial in societally relevant resource management, including subsurface extraction and storage of energy, the deep carbon cycle and generating critical metal deposits. The prevailing view is that fluids navigate through the lithosphere without being influenced by the distinct properties that arise from matter confined at the nanoscale. Here we use electron microscopy and neutron scattering data to show that a diverse range of lithospheric rocks, including sandstones, peridotites and serpentinites, consistently show nanoporosity, predominantly with pore sizes < 100 nanometres. Using molecular dynamics simulations, we demonstrate that water’s dielectric permittivity—a fundamental property that governs its geochemical behaviour—diverges in nanoconfinement from its bulk counterpart under conditions ranging from ambient to extremes of 700 °C and 5 GPa. Our geochemical simulations suggest that changes in water permittivity due to confinement will decrease mineral solubility, a process that is not currently considered in models of fluid–rock interactions. Given that permittivity is also intimately linked to ion speciation, pore-size-dependent properties should be expected to exert a primary influence on rock reactivity and the geochemical evolution of fluids during fluid–rock interactions.
ON–OFF nanopores for optical control of transmembrane ionic communication
Nanoscale photoswitchable proteins could facilitate precise spatiotemporal control of transmembrane communication and support studies in synthetic biology, neuroscience and bioelectronics. Here, through covalent modification of the α-haemolysin protein pore with arylazopyrazole photoswitches, we produced ‘photopores’ that transition between iontronic resistor and diode modes in response to irradiation at orthogonal wavelengths. In the diode mode, a low-leak OFF-state nanopore exhibits a reversible increase in unitary conductance of more than 20-fold upon irradiation at 365 nm. A rectification ratio of >5 was achieved with photopores in the diode state by either direct or alternating voltage input. Unlike conventional electronic phototransistors with intensity-dependent photoelectric responses, the photopores regulated current output solely based on the wavelength(s) of monochromatic or dual-wavelength irradiation. Dual-wavelength irradiation at various relative intensities allowed graded adjustment of the photopore conductance. By using these properties, photonic signals encoding text or graphic messages were converted into ionic signals, highlighting the potential applications of photopores as components of smart devices in synthetic biology.
Dact1 induces Dishevelled oligomerization to facilitate binding partner switch and signalosome formation during convergent extension
Convergent extension (CE) is a universal morphogenetic engine that promotes polarized tissue extension. In vertebrates, CE is regulated by non-canonical Wnt ligands signaling through “core” proteins of the planar cell polarity (PCP) pathway, including the cytoplasmic protein Dishevelled (Dvl), receptor Frizzled (Fz) and tetraspan protein Van gogh-like (Vangl). PCP was discovered in Drosophila to coordinate polarity in the plane of static epithelium, but does not regulate CE in flies. Existing evidence suggests that adopting PCP for CE might be a vertebrate-specific adaptation with incorporation of new regulators. Herein we use Xenopus to investigate Dact1, a chordate-specific protein. Dact1 induces Dvl to form oligomers that dissociate from Vangl, but stay attached with Fz as signalosome-like clusters and co-aggregate with Fz into protein patches upon non-canonical Wnt induction. Functionally, Dact1 antagonizes Vangl, and synergizes with wild-type Dvl but not its oligomerization-defective mutants. We propose that, by promoting Dvl oligomerization, Dact1 couples Dvl binding partner switch with signalosome-like cluster formation to initiate non-canonical Wnt signaling during vertebrate CE.
Vastly different energy landscapes of the membrane insertions of monomeric gasdermin D and A3
Gasdermin D and gasdermin A3 belong to the same family of pore-forming proteins and executors of pyroptosis, a form of programmed cell death. To unveil the process of their pore formation, we examine the energy landscapes upon insertion of the gasdermin D and A3 monomers into a lipid bilayer by extensive atomistic molecular dynamics simulations. We reveal a lower free energy barrier of membrane insertion for gasdermin D than for gasdermin A3 and a preference of gasdermin D for the membrane-inserted and of gasdermin A3 for the membrane-adsorbed state, suggesting that gasdermin D first inserts and then oligomerizes while gasdermin A3 oligomerizes and then inserts. Gasdermin D stabilizes itself in the membrane by forming more salt bridges and pulling phosphatidylethanolamine lipids and more water into the membrane. Gasdermin-lipid interactions support the pore formation. Our findings suggest that both the gasdermin species and the lipid composition modulate gasdermin pore formation.
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