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PubMed2026

Identification of a Novel Thermal Promoter and Its Application in Glutamate Decarboxylase Protein Expression in Bacillus licheniformis.

As a thermotolerant bacterium, Bacillus licheniformis is an attractive chassis for high-temperature biomanufacturing. Here, we identified a novel temperature-responsive promoter, PycgM, which maintained strong transcriptional activity at 37-52 °C. In a promoter-mCherry reporter system, PycgM exhibited 2287.3-fold higher activity than P2 at 52 °C, demonstrating excellent compatibility with a thermotolerant host. Truncation analysis identified a 150-bp core functional region responsible for optimal activity under induction and heat stress. When applied to drive glutamate decarboxylase expression at 50 °C, PycgM enabled γ-aminobutyric acid production of 391.67 g/L with a 98.69% conversion rate, representing a 275% increase over 37 °C fermentation. The whole-cell biocatalyst retained 86% activity after five reuse cycles, and SEM analysis indicated acceptable structural stability despite moderate morphological changes. These results demonstrate that PycgM is a robust, high-temperature genetic element for efficient enzyme and metabolite production in thermotolerant hosts.

Bacillus licheniformis↗

PubMed2025

Approaches to Study Proteins Encoded by Essential Genes.

Although the phenotypes and functions of nonessential proteins can be studied by deletion of their coding sequences (both gene copies in diploid organisms), essential genes cannot be deleted unless loss of the encoded protein can be bypassed. Bypass is often achieved by supplementation with the product of the enzyme. However, supplementation cannot bypass loss of essential genes such as those encoding enzymes of DNA or RNA synthesis. To study proteins encoded by essential genes that cannot be bypassed, the mutations must be conditional in nature. The mutant cells must be able to grow under a permissive condition, but fail to grow under a different condition, the nonpermissive condition. Several methods have been developed to obtain conditional mutations in essential genes. Mutations that result in proteins abnormally sensitive to high temperatures are called temperature-sensitive (Ts) mutants and are a widely used type of conditional mutation. An alternative to Ts mutants is the "degron" system to target proteins for destruction by cellular proteases. Approaches to conditionally control the functions of proteins encoded by essential genes, plus the advantages and disadvantages of these and other approaches, will be considered.

Genes, Essential↗

PubMed2026

Cryo-EM structures of Candida albicans chitin synthase Chs1 reveal a druggable translocation channel.

Invasive candidiasis is a leading cause of hospital-acquired bloodstream infections with high mortality. While the fungal cell wall is an excellent therapeutic target, inhibitor development against the essential chitin synthase (Chs) has been hampered by the absence of structural and mechanistic understanding of class II Chs, which contribute to fungal viability. Here we present cryo-electron microscopy structures of Candida albicans class II Chs (CaChs1) at 2.93-3.38 Å resolution, providing insights into its mechanisms of early elongation, chito-oligomer translocation and inhibition by the CaChs1-specific non-competitive inhibitor diynyl arylamine (DA). Chitin elongation and translocation are coupled to coordinated motion of the glycosyltransferase domain and the dimer interface. Notably, DA binds within the chitin translocation channel where a regulatory lipid resides and inhibits the enzyme by occluding product polymer extrusion. Importantly, DA showed potent synergy with the class I Chs inhibitor nikkomycin Z against C. albicans and Candida auris. These findings establish the chitin translocation channel as a druggable site for rational antifungal design.

Journal Article↗

PubMed2026

Comparative Responses of Invasive and Native Plant Species to Combined Cd and Microplastic Pollution.

The co-occurrence of heavy metal contamination and biodegradable microplastic (polylactic acid, PLA) pollution poses increasing risks to terrestrial plant communities and soil functioning, yet species-specific responses to combined stress remain poorly understood. Cd and microplastics frequently co-occur in agricultural soils, where microplastics can alter cadmium mobility, bioavailability, and transport pathways, potentially modifying metal toxicity and plant stress responses compared with single-pollutant exposure. We investigated the responses of the invasive Bidens pilosa and the native Solanum nigrum grown in monoculture and mixed culture under combined cadmium (Cd) and biodegradable microplastic (PLA) stress by integrating plant growth, photosynthetic performance, oxidative physiology, and rhizosphere biochemical processes. Combined Cd-MP exposure markedly reduced plant growth, chlorophyll content (SPAD), photosystem II efficiency (Fv/Fm), nitrogen accumulation, biomass production, and rhizosphere enzyme activities associated with carbon, nitrogen, and phosphorus cycling. However, B. pilosa maintained greater physiological stability under stress, characterized by higher antioxidant enzyme activities (SOD, CAT, POD), lower reactive oxygen species (H2O2, O2˙-) accumulation, and reduced lipid peroxidation (MDA), whereas S. nigrum exhibited stronger oxidative damage and functional impairment. Multivariate analyses further revealed that root antioxidant capacity was closely associated with rhizosphere microbial enzyme activity, suggesting a root-centered regulatory mechanism linking plant stress tolerance to soil functioning. Overall, the invasive species showed greater tolerance to combined contamination and maintained relatively higher rhizosphere functional activity than the native species, indicating that multi-pollutant stress may alter competitive interactions between invasive and native plants in contaminated environments.

Cadmium↗

PubMed2026

Unconventional Biocatalytic Strategies Orchestrate the Synthesis of the Nucleoside Analog Sinefungin.

Sinefungin is a potent nucleoside antimetabolite of S-adenosylmethionine (SAM). Since its discovery in the 1970s, sinefungin has generated significant scientific interest owing to its role as a bioisostere of SAM and its broad range of biological activities. Despite considerable efforts to uncover the enzymes responsible for sinefungin production in the following years, its biosynthesis remained unclear for decades. Here, we characterize the complete sinefungin biosynthetic gene cluster (sin BGC) from Streptomyces incarnatus NRRL 8089. In vitro and in vivo analyses support a recent finding that the defining carbon-carbon (C-C) bond is formed not by a long-hypothesized PLP-dependent process, but by a vitamin B12-dependent radical SAM enzyme. We provide direct mechanistic evidence, via isotope-labeled products, that the adenosyl group of sinefungin originates from adenosylcobalamin and is atypically consumed via a homolytic SH2 substitution reaction. We also characterize two peptide aminoacyl-tRNA ligases (PEARLs) that append alanines onto the nucleoside scaffold using tRNA-activated amino acids. The PEARLs act directly on small molecules rather than macromolecular substrates, with one PEARL capable of iterative elongation. In addition, we perform in-vitro substrate profiling of several sin BGC-encoded enzymes. We reveal that multiple enzymes show specificity toward phosphorylated intermediates, including the earliest-acting PEARL enzyme. These observations provide an explanation for a cryptic phosphorylation-dephosphorylation strategy observed in the pathway, as they prevent the formation of the highly toxic sinefungin inside the cell. Finally, we leverage these enzymes in a reduced multi-enzyme cascade to biosynthesize sinefungin. Together, these findings expand upon our current knowledge of radical-mediated C-C bond formation and PEARL enzyme catalysis, unlocking biocatalytic possibilities to produce amino acid-nucleoside conjugates.

Streptomyces↗

PubMed2026

The anaerobic fungus Caecomyces churrovis produces H2 via a non-bifurcating NADH-dependent enzyme complex.

UNLABELLED: Hydrogenosomes are mitochondrion-derived organelles that produce ATP and H2 to support energy metabolism in anaerobic eukaryotes. H2 production allows reoxidation of reduced cofactors generated during fermentative metabolism; however, the metabolic mechanisms for H2 production in anaerobic eukaryotes remain incompletely understood. In particular, it remains unclear whether anaerobic fungi (AF) hydrogenosomes use a ferredoxin-dependent pathway or a distinct mechanism to regenerate NAD(P)+ and link electron transfer to H2 formation. Here, by combining genomic search, proteomic analysis, and enzymology, we reveal the molecular mechanism for H2 production in the AF Caecomyces churrovis. Our enzyme assays on the organelle fraction of C. churrovis revealed the activity of H2:NAD+ oxidoreductase but not pyruvate:ferredoxin oxidoreductase, which is usually linked to H2 formation. We identified genes encoding [FeFe] hydrogenase (Hyd) and NADH dehydrogenase subunits E and F (NuoE and NuoF) in C. churrovis and confirmed their expression in the isolated hydrogenosomal fractions by proteomic analysis. Combining the individually purified enzymes, we found Hyd and NuoEF proteins formed H2 directly from NADH independently of ferredoxin, functioning as a non-bifurcating NADH-dependent enzyme rather than an electron-bifurcating enzyme known from anaerobic prokaryotes. We identified homologs of hydrogenosomal NuoE, NuoF, and Hyd in many other AF, indicating this pathway is commonly shared among the AF. This work demonstrates the existence of a non-bifurcating NADH-dependent enzyme complex for H2 production in eukaryotes. Moreover, this complex could potentially be exploited as a target for controlling AF H2 production and altering fungal metabolism. IMPORTANCE: H2 production is a prominent feature of anaerobic energy metabolism, yet our understanding of eukaryotic mechanisms remains limited. Anaerobic fungi (AF) are key decomposers of lignocellulose and contribute to hydrogen flux in anaerobic environments. Although it has been more than 40 years since the H2 production in Neocallimastix was first reported, the molecular mechanism for hydrogenosomal H2 production and redox balance remains unclear. We demonstrate that AF produce H2 from NADH utilizing a non-bifurcating NADH-dependent enzyme complex rather than an electron-bifurcating, ferredoxin-dependent variant. We show that this enzyme complex is conserved across multiple AF lineages and thus demonstrate the occurrence of a non-bifurcating NADH-dependent enzyme in eukaryotes. This discovery expands our understanding of eukaryotic hydrogenosomal metabolism, reveals a previously unknown strategy for redox balancing, and highlights potential targets for manipulating H2 production. These insights have broad implications for microbial energy metabolism, anaerobic ecosystems, and bioengineering of H2-producing systems.

Hydrogen↗

PubMed2026

Key enzyme optimization and multi-node metabolic flux regulation drive l-arginine production in Escherichia coli.

Microbial production of l-arginine is often constrained by tight metabolic regulation and insufficient precursor supply. Here a plasmid-free, non-auxotrophic Escherichia coli strain for high-level production of l-arginine was rationally engineered, based on our previous constructed strain G0 with 12.4 g/L l-arginine production in flask. Glucose metabolism and glutamate/aspartate uptake were initially enhanced, with subsequent semi-rational engineering of key enzymes, ornithine acetyltransferase (OAT) and argininosuccinate synthase (ASS), to promote ATP synthesis. OAT was firstly rational engineered by introducing amide group for the residues near substrate-binding pocket to stabilize oxyanion transition states, with achieving that variant Y386Q showed Km/kcat at 6.58 mM-1 min-1, 2 times higher than that of wild type. Variant Y332L of ASS was novelty fused with argininosuccinate lyase via GGGGS linker for ASS activity measurement, which helped improve l-arginine titer to 17.5 g/L. With further studies by screening of rate-limiting nodes on the genome-scale level based on sRNA strategy, aspartate and glutamate pathways were synergistically enhanced, along with utilizing carbon dioxide recycling for carbamoyl phosphate synthesis to drive ammonia donor supply. The obtained final plasmid-free and non-auxotrophic strain G16 produced 21.1 g/L l-arginine in flask, 76.6% higher than that of original strain G0. In 5 L fermenter, 125.6 g/L l-arginine was produced by fed-batch fermentation, with a yield of 0.53 g/g glucose. This study underscores that the convergence of mechanistic enzyme redesign and systems-level pathway optimization is critical to unlocking high-efficient amino acid production, offering a transferable blueprint for rational strain engineering in industrial biotechnology.

Argininosuccinate synthase↗

PubMed1975

Enzymic reactions of fatty acid hydroperoxides in extracts of potato tuber. II. Conversion of 9- and 13-hydroperoxy-octadecadienoic acids to monohydroxydienoic acid, epoxyhydroxy- and trihydroxymonoenoic acid derivatives.

1. Crude extracts and partially purified enzyme preparations from potato tubers catalyse, at pH 5-7, the conversion of linoleic acid hydroperoxides to a range of oxygenated fatty acid derivatives. 2. 9-D- and 13-L-hydroperoxide isomers are converted at similar rates to equivalent (isomeric) products. 3. The major products from the 13-hydroperoxide isomer were identified as the corresponding monohydroxydienoic acid derivative, threo-11-hydroxy-trans12,13-epoxy-octadec-cis9-enoic acid and 9,12,13-trihydroxy-octadec-trans10-enoic acid. The corresponding products from the 9-hydroperoxide were the monohydroxydienoic acid, 9,10-epoxy-11-hydroxy-octadec-12-enoic acid and 9,10,13-trihydroxy-octadec-11-enoic acid. 4. No separation of activities forming the different products was achieved by partial purification of enzyme extracts. 5. Product formation was unaffected by EDTA, CN-, sulphydryl reagents or glutathione but was reduced by boiling the extracts. 6. This system is compared with the 9-hydroperoxide-specific enzymic formation of divinyl ether derivatives by potato extracts.

Fatty Acids, Unsaturated↗

PubMed2024

A metabolomics pipeline highlights microbial metabolism in bloodstream infections.

The growth of antimicrobial resistance (AMR) highlights an urgent need to identify bacterial pathogenic functions that may be targets for clinical intervention. Although severe infections profoundly alter host metabolism, prior studies have largely ignored microbial metabolism in this context. Here, we describe an iterative, comparative metabolomics pipeline to uncover microbial metabolic features in the complex setting of a host and apply it to investigate gram-negative bloodstream infection (BSI) in patients. We find elevated levels of bacterially derived acetylated polyamines during BSI and discover the enzyme responsible for their production (SpeG). Blocking SpeG activity reduces bacterial proliferation and slows pathogenesis. Reduction of SpeG activity also enhances bacterial membrane permeability and increases intracellular antibiotic accumulation, allowing us to overcome AMR in culture and in vivo. This study highlights how tools to study pathogen metabolism in the natural context of infection can reveal and prioritize therapeutic strategies for addressing challenging infections.

Metabolomics↗

PubMed2026

Bioprospecting microbial genomes to expand the biocatalytic toolbox of rubber oxygenases.

A set of rubber oxygenases was discovered through phylogenetic analysis and AI-based structural modeling of complexes of the putative enzymes with a substrate mimicking cis-1,4-polyisoprene. Sixteen candidate proteins were selected from thermophilic microorganisms, all sequence-related to the Latex clearing protein from Streptomyces sp. K30 (LcpK30). Sequence truncation and solubility tags were then evaluated to enhance protein expression, with the SUMO tag proving to be the most effective. Including LcpK30, nine heme-containing oxygenases were successfully expressed in E. coli NEB 10-beta cells, purified (35-157 mg L-1 yield) and characterized. Steady-state kinetics revealed significant rubber latex-degrading properties for six of them, with the truncated SUMO-fused LcpK30 (SUMO-LcpK30T) showing activity in agreement with literature. Notably, the catalytic efficiencies of all the expressed homologs lay within one order of magnitude and the oxygenase from Thermomonospora echinospora was found to be particularly promising in terms of activity, especially at high latex concentrations (more than 1% w/v). The analysis of reaction mixtures by both HPLC and HPLC-MS confirmed the oxidation of cis-1,4-polyisoprene to form the expected isoprenoid oligomers (n = 2-12), whose distribution was consistent with the usual endo-type cleavage pattern in all but one case. This bioprospecting effort afforded a platform of new rubber-degrading enzymes with diverse efficiencies and product profiles, capable of adapting to targeted applications.

Oxygenases↗

PubMed2026

Flux rewiring enables native D-glucosamine production in Escherichia coli.

D-Glucosamine is an industrially important amino sugar used in pharmaceuticals, nutraceuticals, and functional materials, yet its production remains dominated by chemical extraction from chitinous biomass, raising sustainability and allergen concerns. Escherichia coli natively synthesizes D-glucosamine directly from D-glucose through endogenous metabolism, revealing an underutilized amino sugar biosynthetic capability. Building on this native pathway, D-glucosamine production was enhanced through targeted genetic modifications and systematic optimization of nitrogen metabolism and cultivation conditions, reaching 9.2 g L-1 under shake-flask conditions. This work extends a phosphorylation-dephosphorylation strategy previously developed for neutral rare sugars to amino sugar biosynthesis, demonstrating the broader applicability of this metabolic design principle. Phosphatase identity emerged as a key control point for product formation: YbiV was the most effective phosphatase for selective D-glucosamine production, whereas alternative phosphatases redirected flux toward D-sedoheptulose. This enzyme-dependent flux partitioning further enabled tunable co-production of D-glucosamine and D-sedoheptulose. Native amino sugar biosynthesis in E. coli provides a controllable framework for producing chemically distinct sugars through endogenous metabolism and establishes a generalizable strategy for engineering amino sugar and other nitrogen-containing metabolite biosynthesis.

Escherichia coli↗

PubMed2025

Defining the networks that connect RNase III and RNase J-mediated regulation of primary and specialized metabolism in Streptomyces venezuelae.

UNLABELLED: RNA metabolism involves coordinating RNA synthesis with RNA processing and degradation. Ribonucleases play fundamental roles within the cell, contributing to the cleavage, modification, and degradation of RNA molecules, with these actions ensuring appropriate gene regulation and cellular homeostasis. Here, we employed RNA sequencing to explore the impact of RNase III and RNase J on the transcriptome of Streptomyces venezuelae. Differential expression analysis comparing wild-type and RNase mutant strains at distinct developmental stages revealed significant changes in transcript abundance, particularly in pathways related to multicellular development, nutrient acquisition, and specialized metabolism. Both RNase mutants exhibited dysregulation of the BldD regulon, including altered expression of many cyclic-di-GMP-associated enzymes. We also observed precocious chloramphenicol production in these RNase mutants and found that in the RNase III mutant, this was associated with PhoP-mediated regulation. We further found that RNase III directly targeted members of the PhoP regulon, suggesting a link between RNA metabolism and a regulator that bridges primary and specialized metabolism. We connected RNase J function with translation through the observation that RNase J directly targets multiple ribosomal protein transcripts for degradation. These findings establish distinct but complementary roles for RNase III and RNase J in coordinating the gene expression dynamics critical for S. venezuelae development and specialized metabolism. IMPORTANCE: RNA processing and metabolism are mediated by ribonucleases and are fundamental processes in all cells. In the morphologically complex and metabolically sophisticated Streptomyces bacteria, RNase III and RNase J influence both development and metabolism through poorly understood mechanisms. Here, we show that both ribonucleases are required for the proper expression of the BldD developmental pathway and contribute to the control of chloramphenicol production, with an interesting connection to phosphate regulation for RNase III. Additionally, we show that both RNases have the potential to impact translation through distinct mechanisms and can function cooperatively in degrading specific transcripts. This study advances our understanding of RNases in Streptomyces biology by providing insight into distinct contributions made by these enzymes and the intriguing interplay between them.

Streptomyces↗

PubMed2025

Municipal sewage as a pathway for multidrug-resistant KPC-producing Klebsiella pneumoniae from hospital effluent to urban stream: challenges for wastewater management.

Carbapenemase-producing Klebsiella pneumoniae is among the mainly reasons for death from bacterial infection associated with antibiotic resistance. Its widespread dissemination, especially due to KPC enzyme, is one of the main challenges in One Health perspective. Here, we studied 42 KPC-producing K. pneumoniae isolates from hospital wastewater, municipal wastewater from wastewater treatment plant (WWTP), and urban stream which receives treated municipal effluent. The isolates presented broad resistance to β-lactams antibiotics, as well as to fluoroquinolones, and show antibiotic resistance profile very similar, even those from out-of-hospital settings. Along to blaKPC gene, blaCTX-M-1 (33,3 %, n = 14), blaCTX-M-8 (19 %, n = 8), qnrB (52,3 %, n = 22), qnrS (2,38 %, n = 1), and rmtB (19 %, n = 8) were detected. There was a predominance of gene that confers tolerance to silver and copper metals, as well as to virulence factor related to enterobactin and colibactin production. Macrorestriction genomic analysis by XbaI enzyme demonstrated several pulsotype, but some ones are related. Isolates from hospital wastewater were detected after 4 months at the same sampling point, as well as similar to those detected in WWTP and urban stream demonstrating the effluents role as spreaders of antibiotic resistance. This study provides data on the characterization of KPC-producing K. pneumoniae, which contributes to the epidemiological characterization of human pathogens transmitted by aquatic matrices. In view of the universal sanitation and control of antimicrobial resistance in the One Health perspective, greater investment in effluent treatment is necessary to avoid contamination and environmental dissemination of antibiotic-resistant bacteria.

Klebsiella pneumoniae↗

PubMed2026

Degradation of a graphene-reinforced polyamide by fungi: When culture conditions matter.

The large-scale production, marketing and disposal of polymer-based graphene products can lead to the dispersal of graphene-enriched plastic particles into terrestrial ecosystems, where they might accumulate if not degraded by organisms. The objective of this work is to test the degradability and compatibility of one polyamide-6 polymer reinforced with reduced graphene-oxide (PA6-rGO) and its base constituents (polyamide-6, PA6; reduced graphene oxide, rGO) using mono- and co-cultures of two lignin-degrading fungi (Bjerkandera adusta and Morchella esculenta) grown under different nutrient conditions. Fungal (co-)cultures were exposed to pure rGO or abraded powders of PA6 and PA6-rGO in two different liquid media, and monitored over time for biomass growth, H2O2 production, and activity of two lignolytic enzymes (i.e., Laccase, Lac, and Lignin peroxidase, LiP). The changes in polyamide structure were evaluated by proton nuclear magnetic resonance and mass spectrometry, and changes in rGO were evaluated by Raman spectroscopy. The materials had no effect on fungal growth. PA6 increased Lac secretion only in low nutrient medium, while PA6-rGO slightly suppressed LiP activity. Only M. esculenta promoted polyamides oxidation when cultured in a low nutrient medium, as evidenced by a change in mass distribution values (m/z: 400-420) and the appearance of a new resonance peak (at 5.37 ppm). Lignolytic exudates in co-cultures low in nutrients caused a greater change in rGO, as shown by the increase in the ID/IG ratio. The degradation of rGO, PA6 and PA6-rGO depended on culture conditions.

Graphite↗

PubMed2026

Biochemical analysis of the TPS-b subfamily reveals a cineole-centered monoterpene biosynthetic module in Medicago truncatula.

Terpenoids constitute one of the largest and most structurally diverse classes of plant specialized metabolites, with diversity generated by terpene synthases (TPSs) and downstream tailoring enzymes. In Medicago truncatula, the TPS-b subfamily comprises five putative synthases, two of which are embedded within a previously uncharacterized genomic locus containing a cytochrome P450 (CYP) and a BAHD-type acyltransferase. Here, we present a comprehensive biochemical analysis of the M. truncatula TPS-b subfamily and define a cineole-centered monoterpene biosynthetic module. Heterologous expression and in vitro assays with multiple prenyl diphosphate substrates revealed three catalytically active TPS-b enzymes with distinct substrate preferences and product profiles. MtTPS4 functions as a dedicated (E)-β-ocimene synthase, whereas MtTPS15 exhibits substrate-dependent bifunctionality, producing (E)-β-ocimene from geranyl diphosphate and α-farnesene from farnesyl diphosphate. MtTPS36 generates 1,8-cineole as the predominant product alongside α-terpineol from geranyl and neryl diphosphate. Genome analysis revealed that MtTPS36 is colocalized with a cytochrome P450 belonging to the CYP736 family. Biochemical characterization of this CYP identified a previously undescribed plant cineole hydroxylase that catalyzes oxidation of 1,8-cineole to yield 2α-hydroxy-1,8-cineole (also known as 2-exo-hydroxy-1,8-cineole), establishing a TPS-CYP biosynthetic module. These results define the gene-to-metabolite relationships within the TPS-b subfamily in M. truncatula and expand the known enzymatic biosynthetic capacity underlying oxygenated monoterpene biosynthesis in plants.

1,8-Cineole↗

PubMed2024

Accessing and exploring the unusual chemistry by radical SAM-RiPP enzymes.

Radical SAM enzymes involved in the biosynthesis of ribosomally synthesized and post-translationally modified peptides catalyze unusual transformations that lead to unique peptide scaffolds and building blocks. Several natural products from these pathways show encouraging antimicrobial activities and represent next-generation therapeutics for infectious diseases. These systems are uniquely configured to benefit from genome-mining approaches because minimal substrate and cognate modifying enzyme expression can reveal unique, chemically complex transformations that outperform late-stage chemical reactions. This report highlights the main strategies used to reveal these enzymatic transformations, which have relied mainly on genome mining using enzyme-first approaches. We describe the general biosynthetic components for rSAM enzymes and highlight emerging approaches that may broaden the discovery and study of rSAM-RiPP enzymes. The large number of uncharacterized rSAM proteins, coupled with their unpredictable transformations, will continue to be an essential and exciting resource for enzyme discovery.

S-Adenosylmethionine↗

PubMed2026

Hepatic ketogenic insufficiency blunts exercise-induced energy expenditure and alters mitochondrial proteins in skeletal muscle.

Ketone body (KB) utilization increases during fasting and exercise due to enhanced hepatic fatty acid oxidation and KB production via the rate-limiting mitochondrial enzyme hydroxymethylglutaryl-CoA synthase (HMGCS2). Since KB metabolism intersects with multiple metabolic pathways and skeletal muscle KB catabolism rises during exercise, we tested the hypothesis that liver-specific HMGCS2 knockouts (KO) would have reduced energy expenditure (EE) and changes in the mitochondrial proteome of skeletal muscle with chronic exercise through voluntary wheel running (VWR), time-restricted feeding (TRF), or both combined to boost hepatic KB production and utilization. Control (CON) and HMGCS2 knockout (KO) mice (n = 6-8 per group) underwent sedentary ad libitum feeding (SED + AL), SED + TRF, VWR + AL, and VWR + TRF for 16 wk, with whole body EE measured using indirect calorimetry. In CON mice, VWR increased total EE by 19.5% and nonresting EE by 50% under AL conditions, and total EE by 16% and nonresting EE by 47.9% under TRF conditions. However, the EE increases seen with VWR did not occur in KO mice. Proteomic analysis revealed that the loss of liver HMGCS2 significantly impacted proteins involved in metabolic processes within skeletal muscle, including reduced oxidative phosphorylation (OXPHOS) protein expression in SED KO mice compared with sedentary CON. Notably, VWR restored OXPHOS protein expression in the muscle of the liver HMGCS2 KO but did not alter it in the CON. Furthermore, muscle from liver HMGCS2 KO mice had elevated expression of glycolytic pathways in sedentary and VWR conditions. These results indicate that hepatic ketogenic deficiency (HMGCS2 KO) diminishes exercise-induced increases in EE and uniquely impacts baseline and exercise-related adaptations in the metabolic and mitochondrial proteome of skeletal muscle.

Hydroxymethylglutaryl-CoA Synthase↗

PubMed2026

Yeast Strain Development and Process Intensification in High-Gravity Fermentation.

High- and very-high-gravity (HG/VHG) fermentation increases substrate loading and product titers, thereby improving fermenter utilisation and potentially reducing water use and downstream processing requirements. Initially developed for brewing and fuel ethanol production, these approaches are now applied more broadly in food, beverage, and bioproduct manufacturing. This MiniReview summarises operational definitions and industrial drivers of HG/VHG fermentation and examines the associated constraints in rheology, mass and heat transfer, osmotic and ethanol stress, nutrient availability, and oxidative damage. Yeast improvement strategies are reviewed, including adaptive laboratory evolution, mutagenesis, genome shuffling, multiplex genome editing, non-conventional yeasts, and multi-omics-guided selection. Process developments such as no-cook simultaneous liquefaction, saccharification and fermentation (SLSF), enzyme formulation, nutrient management, and in situ product recovery are considered together with applications in alcoholic beverages, organic acids, microbial lipids, and other value-added products. The review also discusses coproduct valorisation and the need to integrate strain development with process design. Current evidence supports HG/VHG fermentation as a useful process-intensification platform, although performance and sustainability depend strongly on feedstock, operating conditions, product requirements, and the basis used to report fermentation outcomes.

circular bioeconomy↗