Genetically encoded biosensors for the circular plastics bioeconomy.

Current plastic production and consumption routes are unsustainable due to impact upon climate change and pollution, and therefore reform across the entire value chain is required. Biotechnology offers solutions for production from renewable feedstocks, and to aid end of life recycling/upcycling of plastics. Biology sequence/design space is complex requiring high-throughput analytical methods to facilitate the iterative optimisation, design-build, test-learn (DBTL), cycle of Synthetic Biology.

Metabolomic heterogeneity of ageing with ethnic diversity: a step closer to healthy ageing.

Introduction: Outside of case-control settings, ethnicity specific changes in the human metabolome are understudied especially in community dwelling, ageing men. Characterising serum for age and ethnicity specific features can enable tailored therapeutics research and improve our understanding of the interplay between age, ethnicity, and metabolism in global populations.

Objective: A metabolomics approach was adopted to profile serum metabolomes in middle-aged and elderly men of different ethnicities from the Northwest of England, UK.

Complex waste stream valorization through combined enzymatic hydrolysis and catabolic assimilation by Pseudomonas putida.

Biogenic waste-derived feedstocks for production of fuels, chemicals, and materials offer great potential supporting the transition to net-zero and greater circularity. However, such feedstocks are heterogeneous and subject to geographical and seasonal variability. Here, we show that, through careful strain selection and metabolic engineering, Pseudomonas putida can be employed to permit efficient co-utilization of highly heterogeneous substrate compositions derived from hydrolyzed mixed municipal-like waste fractions (food, plastic, organic, paper, cardboard, and textiles) for growth and synthesis of exemplar bioproducts.

Tuning the performance of a TphR-based terephthalate biosensor with a design of experiments approach.

Transcription factor-based biosensors are genetic tools that aim to predictability link the presence of a specific input stimuli to a tailored gene expression output. The performance characteristics of a biosensor fundamentally determines its potential applications. However, current methods to engineer and optimise tailored biosensor responses are highly nonintuitive, and struggle to investigate multidimensional sequence/design space efficiently.

Synthesis and screening of a library of Lewis deoxyfluoro-analogues reveals differential recognition by glycan-binding partners.

Glycan-mediated interactions play a crucial role in biology and medicine, influencing signalling, immune responses, and disease pathogenesis. However, the use of glycans in biosensing and diagnostics is limited by cross-reactivity, as certain glycan motifs can be recognised by multiple biologically distinct protein receptors. To address this specificity challenge, we report the enzymatic synthesis of a 150-member library of site-specifically fluorinated Lewis analogues ('glycofluoroforms') using naturally occurring enzymes and fluorinated monosaccharides.

Effect of Peptide-Polymer Host-Guest Electrostatic Interactions on Self-Assembling Peptide Hydrogels Structural and Mechanical Properties and Polymer Diffusivity.

Peptide-based supramolecular hydrogels are an attractive class of soft materials for biomedical applications when biocompatibility is a key requirement as they exploit the physical self-assembly of short self-assembling peptides avoiding the need for chemical cross-linking. Based on the knowledge developed through our previous work, we designed two novel peptides, (FKFE) and (FEFK), that form transparent hydrogels at pH 7. We characterized the phase behavior of these peptides and showed the clear link that exists between the charge carried by the peptides and the physical state of the samples.

Engineering Biocatalysts for the C-H Activation of Fatty Acids by Ancestral Sequence Reconstruction.

Selective, one-step C-H activation of fatty acids from biomass is an attractive concept in sustainable chemistry. Biocatalysis has shown promise for generating high-value hydroxy acids, but to date enzyme discovery has relied on laborious screening and produced limited hits, which predominantly oxidise the subterminal positions of fatty acids. Herein we show that ancestral sequence reconstruction (ASR) is an effective tool to explore the sequence-activity landscape of a family of multidomain, self-sufficient P450 monooxygenases.

Genetic and process engineering for polyhydroxyalkanoate production from pre- and post-consumer food waste.

Biopolymers produced as microbial carbon storage systems, such as polyhydroxyalkanoates (PHAs), offer potential to be used in place of petrochemically derived plastics. Low-value organic feedstocks, such as food waste, have been explored as a potential substrate for the microbial production of PHAs. In this review, we discuss the biosynthesis, composition and producers of PHAs, with a particular focus on the genetic and process engineering efforts to utilise non-native substrates, derived from food waste from across the entire supply chain, for microbial growth and PHA production.

Engineered co-culture for consolidated production of phenylpropanoids directly from aromatic-rich biomass.

Consolidated bioprocesses for the in situ hydrolysis and conversion of biomass feedstocks into value-added products offers great potential for both process and cost reduction. However, to date few consolidated bioprocesses have been developed that target aromatic rich feedstock fractions. Reported here is the development of synthetic co-cultivation for the consolidated hydrolysis and valorisation of corncob hydroxycinnamic acids.

Sustainable biosynthetic pathways to value-added bioproducts from hydroxycinnamic acids.

The biorefinery concept, in which biomass is utilized for the production of fuels and chemicals, emerges as an eco-friendly, cost-effective, and renewable alternative to petrochemical-based production. The hydroxycinnamic acid fraction of lignocellulosic biomass represents an untapped source of aromatic molecules that can be converted to numerous high-value products with industrial applications, including in the flavor and fragrance sector and pharmaceuticals. This review describes several biochemical pathways useful in the development of a biorefinery concept based on the biocatalytic conversion of the hydroxycinnamic acids ferulic, caffeic, and p-coumaric acid into high-value molecules.

Biochemical characterization of a glycoside hydrolase family 43 β-D-galactofuranosidase from the fungus Aspergillus niger.

β-D-Galactofuranose (Galf) and its polysaccharides are found in bacteria, fungi and protozoa but do not occur in mammalian tissues, and thus represent a specific target for anti-pathogenic drugs. Understanding the enzymatic degradation of these polysaccharides is therefore of great interest, but the identity of fungal enzymes with exclusively galactofuranosidase activity has so far remained elusive. Here we describe the identification and characterization of a galactofuranosidase from the industrially important fungus Aspergillus niger.

The feruloyl esterase from Thermobacillus xylanilyticus shows broad specificity for processing pre-biotic feruloylated xylooligosaccharides at high temperatures.

Ferulic acid has antioxidant properties of interest to the food industry and can be released from natural plant fibres using feruloyl esterases. Esterases active at high temperatures are highly desirable but currently underrepresented. Here we report the biochemical characterization of the feruloyl esterase from Thermobacillus xylanilyticus.

Three-Component Stereoselective Enzymatic Synthesis of Amino-Diols and Amino-Polyols.

Amino-polyols represent attractive chemical building blocks but can be challenging to synthesize because of the high density of asymmetric functionalities and the need for extensive protecting-group strategies. Here we present a three-component strategy for the stereoselective enzymatic synthesis of amino-diols and amino-polyols using a diverse set of prochiral aldehydes, hydroxy ketones, and amines as starting materials. We were able to combine biocatalytic aldol reactions, using variants of d-fructose-6-phosphate aldolase (FSA), with reductive aminations catalyzed by IRED-259, identified from a metagenomic library.

Directed Evolution of Transcription Factor-Based Biosensors for Altered Effector Specificity.

Transcription factor-based biosensors are important tools in Synthetic Biology for the sensing of industrially valuable molecules and clinically important metabolites, therefore presenting applications in the bioremediation, industrial biotechnology, and biomedical fields. The directed evolution of allosteric transcription factors (aTFs) with the aim of altering effector specificity has the potential for the development of new biosensors to detect natural and nonnatural molecules, expanding the scope of available aTF-based biosensors. In this chapter, we delineate a general method for the directed evolution of aTFs.

Self-Assembling Peptide Hydrogels as Functional Tools to Tackle Intervertebral Disc Degeneration.

Low back pain (LBP), caused by intervertebral disc (IVD) degeneration, is a major contributor to global disability. In its healthy state, the IVD is a tough and well-hydrated tissue, able to act as a shock absorber along the spine. During degeneration, the IVD is hit by a cell-driven cascade of events, which progressively lead to extracellular matrix (ECM) degradation, chronic inflammation, and pain.

Microarray screening reveals two non-conventional SUMO-binding modules linked to DNA repair by non-homologous end-joining.

SUMOylation is critical for numerous cellular signalling pathways, including the maintenance of genome integrity via the repair of DNA double-strand breaks (DSBs). If misrepaired, DSBs can lead to cancer, neurodegeneration, immunodeficiency and premature ageing. Using systematic human proteome microarray screening combined with widely applicable carbene footprinting, genetic code expansion and high-resolution structural profiling, we define two non-conventional and topology-selective SUMO2-binding regions on XRCC4, a DNA repair protein important for DSB repair by non-homologous end-joining (NHEJ).

Acidic and basic self-assembling peptide and peptide-graphene oxide hydrogels: characterisation and effect on encapsulated nucleus pulposus cells.

Extracellular pH can have a profound effect on cell metabolism, gene and protein expression. Nucleus pulposus (NP) cells, for example, under acidic conditions accelerate the production of degradative enzymes and pro-inflammatory cytokines, leading ultimately to intervertebral disc degeneration, a major cause of back pain. Self-assembling peptide hydrogels constitute a well-established class of biomaterials that could be exploited as pH-tunable platform to investigate cell behaviour under normal and non-physiological pH.

Structural basis of terephthalate recognition by solute binding protein TphC.

Biological degradation of Polyethylene terephthalate (PET) plastic and assimilation of the corresponding monomers ethylene glycol and terephthalate (TPA) into central metabolism offers an attractive route for bio-based molecular recycling and bioremediation applications. A key step is the cellular uptake of the non-permeable TPA into bacterial cells which has been shown to be dependent upon the presence of the key tphC gene. However, little is known from a biochemical and structural perspective about the encoded solute binding protein, TphC.

Genetic and process engineering strategies for enhanced recombinant N-glycoprotein production in bacteria.

Background: The production of N-linked glycoproteins in genetically amenable bacterial hosts offers great potential for reduced cost, faster/simpler bioprocesses, greater customisation, and utility for distributed manufacturing of glycoconjugate vaccines and glycoprotein therapeutics. Efforts to optimize production hosts have included heterologous expression of glycosylation enzymes, metabolic engineering, use of alternative secretion pathways, and attenuation of gene expression. However, a major bottleneck to enhance glycosylation efficiency, which limits the utility of the other improvements, is the impact of target protein sequon accessibility during glycosylation.

DL_FFLUX: A Parallel, Quantum Chemical Topology Force Field.

DL_FFLUX is a force field based on quantum chemical topology that can perform molecular dynamics for flexible molecules endowed with polarizable atomic multipole moments (up to hexadecapole). Using the machine learning method kriging (aka Gaussian process regression), DL_FFLUX has access to atomic properties (energy, charge, dipole moment, etc.) with quantum mechanical accuracy.

Electrostatics Explains the Reverse Lewis Acidity of BH and Boron Trihalides: Infrared Intensities and a Relative Energy Gradient (REG) Analysis of IQA Energies.

The reaction path for the formation of BX-NH (X = H, F, Cl, Br) complexes was divided into two processes: (i) rehybridization of the acid while adopting a pyramidal geometry, and (ii) the complex formation from the pyramidal geometries of the acid and base. The interacting quantum atom (IQA) method was used to investigate the Lewis acidity trend of these compounds. This topological analysis suggests that the boron-halogen bond exhibits a considerable degree of ionicity.