Expression patterns of immune checkpoint proteins and Plasmodium falciparum-induced cytokines in chronic hepatitis B virus-infected and uninfected individuals: A cross-sectional study.

Background and Aim: Chronic hepatitis B virus (CHB) infection remains a major public health problem. The American Association for the Study of Liver Diseases (AASLD) 2018 Hepatitis B Guidelines provide that CHB individuals not requiring antiviral therapy yet are monitored to determine the need for antiviral therapy in the future; however, these tests do not include measurement of cytokines and immune cell characterization. This case-control study compared the cytokine and immune checkpoint protein expression profiles between CHB individuals not yet on antiviral treatment and hepatitis B virus (HBV)-negative individuals. Methods: CD4 and CD8 T cells from CHB and HBV-negative individuals were characterized for immune checkpoint proteins programmed cell death-1 (PD1), T cell Immunoglobulin domain and mucin domain-containing protein 3 (TIM-3), and cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) (CD152), and a memory marker CXCR3 (CD183) using flow cytometry. Malaria-induced cytokine expression levels were determined by stimulating their blood cells with Plasmodium falciparum 3D7 strain antigens (CSP, AMA-1, and TRAP) in whole blood assays, and cytokine levels were measured using a 13-plex Luminex kit. Results: HBV-negative and CHB individuals had comparable levels of CD4+ and CD8+ T cells. However, a proportion of the CD4+ and CD8+ populations from both groups, which were CXCR3+, expressed PD-1 and CD152. The ability to produce cytokines in response to malaria antigen stimulation was not significantly different between the groups. Conclusion: These findings support excluding CHB individuals from antiviral therapy at this stage of infection. However, CHB individuals require regular monitoring to determine the need for later antiviral treatment.

Comparative genomics between Trichomonas tenax and Trichomonas vaginalis: CAZymes and candidate virulence factors.

Introduction: The oral trichomonad Trichomonas tenax is increasingly appreciated as a likely contributor to periodontitis, a chronic inflammatory disease induced by dysbiotic microbiota, in humans and domestic animals and is strongly associated with its worst prognosis. Our current understanding of the molecular basis of T. tenax interactions with host cells and the microbiota of the oral cavity are still rather limited. One laboratory strain of T. tenax (Hs-4:NIH/ATCC 30207) can be grown axenically and two draft genome assemblies have been published for that strain, although the structural and functional annotation of these genomes is not available. Methods: GenSAS and Galaxy were used to annotate two publicly available draft genomes for T. tenax, with a focus on protein-coding genes. A custom pipeline was used to annotate the CAZymes for T. tenax and the human sexually transmitted parasite Trichomonas vaginalis, the most well-characterized trichomonad. A combination of bioinformatics analyses was used to screen for homologs of T. vaginalis virulence and colonization factors within the T. tenax annotated proteins. Results: Our annotation of the two T. tenax draft genome sequences and their comparison with T. vaginalis proteins provide evidence for several candidate virulence factors. These include candidate surface proteins, secreted proteins and enzymes mediating potential interactions with host cells and/or members of the oral microbiota. The CAZymes annotation identified a broad range of glycoside hydrolase (GH) families, with the majority of these being shared between the two Trichomonas species. Discussion: The presence of candidate T. tenax virulence genes supports the hypothesis that this species is associated with periodontitis through direct and indirect mechanisms. Notably, several GH proteins could represent potential new virulence factors for both Trichomonas species. These data support a model where T. tenax interactions with host cells and members of the oral microbiota could synergistically contribute to the damaging inflammation characteristic of periodontitis, supporting a causal link between T. tenax and periodontitis.

hnRNPs: roles in neurodevelopment and implication for brain disorders.

Heterogeneous nuclear ribonucleoproteins (hnRNPs) constitute a family of multifunctional RNA-binding proteins able to process nuclear pre-mRNAs into mature mRNAs and regulate gene expression in multiple ways. They comprise at least 20 different members in mammals, named from A (HNRNP A1) to U (HNRNP U). Many of these proteins are components of the spliceosome complex and can modulate alternative splicing in a tissue-specific manner. Notably, while genes encoding hnRNPs exhibit ubiquitous expression, increasing evidence associate these proteins to various neurodevelopmental and neurodegenerative disorders, such as intellectual disability, epilepsy, microcephaly, amyotrophic lateral sclerosis, or dementias, highlighting their crucial role in the central nervous system. This review explores the evolution of the hnRNPs family, highlighting the emergence of numerous new members within this family, and sheds light on their implications for brain development.

Causal relationship between mitochondrial-associated proteins and cerebral aneurysms: a Mendelian randomization study.

Background: Cerebral aneurysm is a high-risk cerebrovascular disease with a poor prognosis, potentially linked to multiple factors. This study aims to explore the association between mitochondrial-associated proteins and the risk of cerebral aneurysms using Mendelian randomization (MR) methods. Methods: We used GWAS summary statistics from the IEU Open GWAS project for mitochondrial-associated proteins and from the Finnish database for cerebral aneurysms (uIA, aSAH). The association between mitochondrial-associated exposures and cerebral aneurysms was evaluated using MR-Egger, weighted mode, IVW, simple mode and weighted median methods. Reverse MR assessed reverse causal relationship, while sensitivity analyses examined heterogeneity and pleiotropy in the instrumental variables. Significant causal relationship with cerebral aneurysms were confirmed using FDR correction. Results: Through MR analysis, we identified six mitochondrial proteins associated with an increased risk of aSAH: AIF1 (OR: 1.394, 95% CI: 1.109-1.752, p = 0.0044), CCDC90B (OR: 1.318, 95% CI: 1.132-1.535, p = 0.0004), TIM14 (OR: 1.272, 95% CI: 1.041-1.553, p = 0.0186), NAGS (OR: 1.219, 95% CI: 1.008-1.475, p = 0.041), tRNA PusA (OR: 1.311, 95% CI: 1.096-1.569, p = 0.003), and MRM3 (OR: 1.097, 95% CI: 1.016-1.185, p = 0.0175). Among these, CCDC90B, tRNA PusA, and AIF1 demonstrated a significant causal relationship with an increased risk of aSAH (FDR q < 0.1). Three mitochondrial proteins were associated with an increased risk of uIA: CCDC90B (OR: 1.309, 95% CI: 1.05-1.632, p = 0.0165), tRNA PusA (OR: 1.306, 95% CI: 1.007-1.694, p = 0.0438), and MRM3 (OR: 1.13, 95% CI: 1.012-1.263, p = 0.0303). In the reverse MR study, only one mitochondrial protein, TIM14 (OR: 1.087, 95% CI: 1.004-1.177, p = 0.04), showed a causal relationship with aSAH. Sensitivity analysis did not reveal heterogeneity or pleiotropy. The results suggest that CCDC90B, tRNA PusA, and MRM3 may be common risk factors for cerebral aneurysms (ruptured and unruptured), while AIF1 and NAGS are specifically associated with an increased risk of aSAH, unrelated to uIA. TIM14 may interact with aSAH. Conclusion: Our findings confirm a causal relationship between mitochondrial-associated proteins and cerebral aneurysms, offering new insights for future research into the pathogenesis and treatment of this condition.

Ice recrystallization inhibition activity of pulse protein hydrolysates after immobilized metal affinity separation.

Creating molecules capable of inhibiting ice recrystallization is an active research area aiming to improve the freeze-thaw characteristics of foods and biomedical materials. Peptide mixtures have shown promise in preventing freezing-induced damage, but less is known about the relationship between their amino acid compositions and ice recrystallization inhibition (IRI) activities. In this article, we used Ni2+ immobilized metal affinity chromatography (IMAC) to fractionate pulse protein hydrolysates, created by Alcalase and trypsin, into mixtures lacking and enriched in His, and Cys residues. The aim of this study was to fractionate pulse protein hydrolysates based on their amino acid compositions and evaluate their resulting physicochemical and IRI characteristics. Ni2+ IMAC fractionation induced IRI activity in all of the evaluated soy, chickpea, and pea protein hydrolysates regardless of their amino acid composition. Ni2+ IMAC fractionation produced chemically distinct fractions of peptides, differing by their molecular weights, amino acid composition, and IRI activities. The resulting peptide mixtures' molecular weight, amino acid composition, secondary structure, and sodium ion levels were found to have no correlation with their IRI activities. Thus, we demonstrate for the first time the ability of Ni2+ IMAC fractionation to induce IRI activity in hydrolyzed pulse proteins.

The expression of immune checkpoint proteins PD-L1 and TIM3 in mouse and human head and neck squamous cell carcinoma.

The aim of this study was to examine the expression of programmed death-ligand 1 (PD-L1) and of T cell immunoglobulin and mucin domain-containing protein (TIM3) in oral epithelial dysplasia and head and neck squamous cell carcinoma (HNSCC). Mouse HNSCC was induced with 4-nitroquinoline-1 oxide (4NQO). Oral epithelial dysplastic lesions, carcinoma in situ and HNSCC lesions were stained with anti-PD-L1 and TIM3 antibodies. The expression of PD-L1 and TIM3 in tumor cells and immune cells was semiquantitatively measured and compared. In parallel, human dysplasia and HNSCC were stained with anti-PD-L1 and anti-TIM3. The expression pattern of PD-L1+ and TIM3+ cells was further compared. In human and mouse samples both PD-L1 and TIM3 were found to be expressed in neoplastic and immune cells in HNSCC, but not in dysplasia. There was no significant difference in PD-L1 and TIM3 expression between metastatic and nonmetastatic HNSCC. We conclude that the 4NQO-induced mouse HNSCC model may be an excellent preclinical model for immune checkpoint therapy.

Proteogenomic Analysis Identifies a Causal Association between Plasma Apolipoprotein B Levels and Liver Cancer Risk.

Liver oncogenesis is accompanied by discernible protein changes in the bloodstream. By employing plasma proteomic profiling, we can delve into the molecular mechanisms of liver cancer and pinpoint potential biomarkers. In this nested case-control study, we applied liquid chromatography-tandem mass spectrometry for proteome profiling in baseline plasma samples. Differential protein expression was determined and was subjected to functional enrichment, network, and Mendelian randomization (MR) analyses. We identified 193 proteins with notable differential levels between the groups. Of these proteins, MR analysis offered a compelling negative association between apolipoprotein B (APOB) and liver cancer. This association was further corroborated in the UK Biobank cohort: genetically predicted APOB levels were associated with a 31% (95% CI 19-42%) decreased risk of liver cancer; and phenotypic analysis indicated an 11% (95% CI 8-14%) decreased liver cancer risk for every 0.1 g/L increase of circulating APOB levels. Multivariable MR analysis suggested that the hepatic fat content might fully mediate the APOB-liver cancer connection. In summary, we identified some plasma proteins, particularly APOB, as potential biomarkers of liver cancer. Our findings underscore the intricate link between lipid metabolism and liver cancer, offering hints for targeted prophylactic strategies and early detection.

Co-sensitizations to Gibberellin Regulated Proteins (GRPs) in Italy: results of a polycentric study.

Summary: Background. Gibberellin Regulated Proteins (GRPs) are small glycoproteins that induce allergy to various types of fruit. This study aimed to evaluate co-sensitization to cypress pollen and other molecules responsible for fruit allergy, such as nsLTP (Pru p 3), PR-10 (Bet v1), and Profilin (Bet v2). Methods. Sixty subjects sensitized to peach GRP (Pru p 7) were consecutively recruited from four Italian centers: 28 males and 32 females (mean age 37.9 years; range 11-79). Specific IgE for Pru p 7, Pru p 3, Bet v 1, Bet v 2, cypress pollen extract (Cup s), and Cup a 1 were determined in all subjects. Results. Sensitization rates to Cup s, Cup a 1, Pru p 3, Bet v 1, and Bet v 2 in the entire studied population were 90.0%, 83.3%, 45.8%, 40.0%, and 30.0%, respectively. In subjects residing in Northern Italy, the respective sensitization rates were 96.4%, 80.0%, 50.0%, 73.3%, and 40.0%, while in those residing in Southern Italy, they were 83.3%, 86.7%, 40.0%, 6.7%, and 20.0%. The only significant difference was observed for PR-10 (p less than 0.0001) Co-sensitization to PR-10 was found to be associated with a reduced risk of anaphylaxis (OR: 0.125). Allergic reactions were most commonly triggered by peach (26/40), followed by orange (12/40), with other foods being less frequently implicated. Conclusions. This study confirms a high association between sensitization to Pru p 7 and cypress pollen and highlights a high percentage of co-sensitization to nsLTP, PR-10, and profilin. PR-10 emerged as a protective factor against anaphylaxis.

Tdh3 and Rom2 are functional modulators of a conserved condensate-resident RNA-binding protein, Scd6 in Saccharomyces cerevisiae.

RGG-motif proteins play a crucial role in determining mRNA fate. Suppressor of clathrin deficiency 6 (Scd6) is a conserved RGG-motif containing RNP condensate-resident, translation repressor, and decapping activator protein in Saccharomyces cerevisiae. Identifying protein factors that can modulate Scd6 function is critical to understanding the regulation of mRNA fate by Scd6. In this study, using an approach that combined mRNA tethering assay with flow cytometry, we screened 50 genes for their role in modulating the translation repression activity of Scd6. We identified eight conserved modulators with human homologs. Of these, we further characterised in detail guanine nucleotide exchange factor (GEF) Rom2 (Rho1 multicopy suppressor) and glycolytic enzyme Tdh3 (Triose phosphate dehydrogenase 3), which, respectively, impede and promote translation repression activity of Scd6. Our study reveals that Rom2 negatively regulates the arginine methylation of Scd6 and antagonises its localisation to P-bodies. Tdh3, on the other hand, promotes Scd6 interaction with Hmt1, thereby promoting the arginine methylation of Scd6 and enhanced eIF4G1 interaction, which is known to promote its repression activity. Identifying these novel modulators provides exciting new insights into the role of a metabolic enzyme of the glycolytic pathway and guanine nucleotide exchange factor implicated in the cell wall integrity pathway in regulating Scd6 function and, thereby, cytoplasmic mRNA fate.

Polymerization of hydroxyethyl methacrylate (HEMA) under rotation to form core-annular hydrogels.

HYPOTHESIS: We propose to polymerize high water content hydroxyethyl methacrylate (HEMA) formulations in a rotating cylinder to explore the effect of the rotation on microstructure and critical parameters such as diffusivity of model proteins in porous poly-HEMA gels. EXPERIMENTS: Cylindrical molds were partially filled with water-HEMA-initiator-crosslinker mixtures and exposed to UV light while undergoing rotation to polymerize into a cylindrical tube. The process was repeated multiple times to manufacture a core annular rod with multiple concentric rings, in which at least one ring was porous. The porous gels were imaged by scanning electron microscopy to explore the microstructure. The transport of model proteins bovine serum albumin and human γ-globulin was measured and modeled, in radial and axial directions, to obtain the effective diffusivity and partition coefficient. Also, the true diffusivity of proteins was calculated by accounting for the effects of porosity and tortuosity. FINDINGS: The porous gels exhibited diffusion-controlled release of both model proteins. The hydrogels prepared with 55% water in the monomer mixture were porous with non-isotropic structure likely due to axially oriented pores with minimal radial connectivity. The gels with higher water content were isotropic with interconnected pores in both directions. The pore volume increased with water content, but the partition coefficient was relatively constant and less than one likely due to presence of isolated unconnected pores.

Visualizing DNA/RNA, Proteins, and Small Molecule Metabolites within Live Cells.

Live cell imaging is essential for obtaining spatial and temporal insights into dynamic molecular events within heterogeneous individual cells, in situ intracellular networks, and in vivo organisms. Molecular tracking in live cells is also a critical and general requirement for studying dynamic physiological processes in cell biology, cancer, developmental biology, and neuroscience. Alongside this context, this review provides a comprehensive overview of recent research progress in live-cell imaging of RNAs, DNAs, proteins, and small-molecule metabolites, as well as their applications in molecular diagnosis, immunodiagnosis, and biochemical diagnosis. A series of advanced live-cell imaging techniques have been introduced and summarized, including high-precision live-cell imaging, high-resolution imaging, low-abundance imaging, multidimensional imaging, multipath imaging, rapid imaging, and computationally driven live-cell imaging methods, all of which offer valuable insights for disease prevention, diagnosis, and treatment. This review article also addresses the current challenges, potential solutions, and future development prospects in this field.

Analysis of host factor networks during hepatitis B virus infection in primary human hepatocytes.

BACKGROUND: Chronic hepatitis B virus (HBV) infection affects around 250 million people worldwide, causing approximately 887,000 deaths annually, primarily owing to cirrhosis and hepatocellular carcinoma (HCC). The current approved treatments for chronic HBV infection, such as interferon and nucleos(t)ide analogs, have certain limitations as they cannot completely eradicate covalently closed circular DNA (cccDNA). Considering that HBV replication relies on host transcription factors, focusing on host factors in the HBV genome may provide insights into new therapeutic targets against HBV. Therefore, understanding the mechanisms underlying viral persistence and hepatocyte pathogenesis, along with the associated host factors, is crucial. In this study, we investigated novel therapeutic targets for HBV infection by identifying gene and pathway networks involved in HBV replication in primary human hepatocytes (PHHs). Importantly, our study utilized cultured primary hepatocytes, allowing transcriptomic profiling in a biologically relevant context and enabling the investigation of early HBV-mediated effects. METHODS: PHHs were infected with HBV virion particles derived from HepAD38 cells at 80 HBV genome equivalents per cell (Geq/cell). For transcriptomic sequencing, PHHs were harvested 1, 2-, 3-, 5-, and 7 days post-infection (dpi). After preparing the libraries, clustering and sequencing were conducted to generate RNA-sequencing data. This data was processed using Bioinformatics tools and software to analyze DEGs and obtain statistically significant results. Furthermore, qRT-PCR was performed to validate the RNA-sequencing results, ensuring consistent findings. RESULTS: We observed significant alterations in the expression patterns of 149 genes from days 1 to 7 following HBV infection (R2 > 0.7, q < 0.05). Functional analysis of these genes identified RNA-binding proteins involved in mRNA metabolism and the regulation of alternative splicing during HBV infection. Results from qRT-PCR experiments and the analysis of two validation datasets suggest that RBM14 and RPL28 may serve as potential biomarkers for HBV-associated HCC. CONCLUSIONS: Transcriptome analysis of gene expression changes during HBV infection in PHHs provided valuable insights into chronic HBV infection. Additionally, understanding the functional involvement of host factor networks in the molecular mechanisms of HBV replication and transcription may facilitate the development of novel strategies for HBV treatment.

Protein Network Alterations in G-CSF Treated Severe Congenital Neutropenia Patients and Beneficial Effects of Oral Health Intervention.

PURPOSE: Severe congenital neutropenia (SCN) is a raredisorder characterized by diminished neutrophil levels. Despite granulocytecolony-stimulating factor (G-CSF) treatment, SCN patients remain still prone tosevere infections, including periodontal disease-a significant oral healthrisk. This study investigates the host proteome and metaproteome in saliva andgingival crevicular fluid (GCF) of G-CSF-treated patients. EXPERIMENTAL DESIGN: We used label-free quantitative proteomics on saliva and GCF samples from SCN patients before (n = 10, mean age: 10.7 ± 6.6 years) and after a 6-month oral hygiene intervention (n = 9,mean age: 11.6 ± 5.27 years), and from 12 healthy controls. RESULTS: We quantified 894 proteins in saliva (648 human,246 bacterial) and 756 proteins in GCF (493 human, 263 bacterial). Predominant bacterial genera included Streptococcus, Veillonella, Selenomonas, Corynebacterium, Porphyromonas, and Prevotella. SCN patients showed reduced antimicrobial peptides (AMPs) and elevated complement proteins compared tohealthy controls. Oral hygiene intervention improved oral epithelial conditionsand reduced both AMPs and complement proteins. CONCLUSIONS AND CLINICAL RELEVANCE: SCN patients have aunique proteomic profile with reduced AMPs and increased complement proteins, contributing to infection susceptibility. Oral hygiene intervention not onlyimproved oral health in SCN patients but also offers potential overall therapeuticbenefits.

Proteomics analysis of immune response-related proteins in Guillain-Barré Syndrome (GBS) and Chronic Inflammatory Demyelinating Polyneuropathy (CIDP).

The objective is to characterize differentially expressed proteins (DEPs) in Guillain-Barré Syndrome (GBS) and Chronic Inflammatory Demyelinating Polyneuropathy (CIDP) through high-throughput analysis. Sera from 11 healthy controls (HCs), 21 GBS and 19 CIDP patients were subjected to Olink Proteomics Analysis. In the comparison between CIDP and GBS groups, up-regulation of ITM2A and down-regulation of NTF4 were observed. Comparing GBS with HCs revealed 18 up-regulated and 4 down-regulated proteins. Comparing CIDP with the HCs identified 15 up-regulated and 4 down-regulated proteins. Additionally, the correlation between clinical characteristics and DEPs were uncovered. In conclusion, the DEPs have significant potential to advance our understanding of the pathogenesis in these debilitating neurological disorders.

Exploring faba beans (Vicia faba L.): bioactive compounds, cardiovascular health, and processing insights.

Faba beans (Vicia faba L.), integral to the legume family, are a significant component of the global pulse market because of their nutritional richness and positive health implications. While existing reviews have extensively covered the nutritional composition and anti-nutritional factors of faba beans, and their utilization in food product development, the insights into the optimization of processing methods and upcycling the wastewater during faba bean processing remain insufficient. Therefore, this review focuses on consolidating information about their bioactive compounds, elucidating associated health benefits and unveiling the possible application of processing water derived from faba beans. Key issues discussed include the impact of bioactive compounds in faba beans on cardiovascular health and carcinogenic condition, the challenges in processing that affect bioactive content, and the potential nutritional and functional applications of processing water in food production.

Macrophages in the infarcted heart acquire a fibrogenic phenotype, expressing matricellular proteins, but do not undergo fibroblast conversion.

Although some studies have suggested that macrophages may secrete structural collagens, and convert to fibroblast-like cells, macrophage to fibroblast transdifferentiation in infarcted and remodeling hearts remains controversial. Our study uses linage tracing approaches and single cell transcriptomics to examine whether macrophages undergo fibroblast conversion, and to characterize the extracellular matrix expression profile of myeloid cells in myocardial infarction. To examine whether infarct macrophages undergo fibroblast conversion, we identified macrophage-derived progeny using the inducible CX3CR1CreER mice crossed with the PDGFRαEGFP reporter line for reliable fibroblast identification. The abundant fibroblasts that infiltrated the infarcted myocardium after 7 and 28 days of coronary occlusion were not derived from CX3CR1+ macrophages. Infarct macrophages retained myeloid cell characteristics and did not undergo conversion to myofibroblasts, endothelial or vascular mural cells. Single cell RNA-seq of CSF1R+ myeloid cells harvested from control and infarcted hearts showed no significant expression of fibroblast identity genes by myeloid cell clusters. Moreover, infarct macrophages did not express significant levels of genes encoding structural collagens. However, infarct macrophage and monocyte clusters were the predominant source of the fibrogenic growth factors Tgfb1 and Pdgfb, and of the matricellular proteins Spp1/Osteopontin, Thbs1/Thrombospondin-1, Emilin2, and Fn1/fibronectin, while expressing significant amounts of several other matrix genes, including Vcan/versican, Ecm1 and Sparc. ScRNA-seq data suggested similar patterns of matrix gene expression in human myocardial infarction. In conclusion, infarct macrophages do not undergo fibroblast or myofibroblast conversion and do not exhibit upregulation of structural collagens but may contribute to fibrotic remodeling by producing several fibrogenic matricellular proteins.

PEL-PVP: Application of plant vacuolar protein discriminator based on PEFT ESM-2 and bilayer LSTM in an unbalanced dataset.

Plant vacuoles, play a crucial role in maintaining cellular stability, adapting to environmental changes, and responding to external pressures. The accurate identification of vacuolar proteins (PVPs) is crucial for understanding the biosynthetic mechanisms of intracellular vacuoles and the adaptive mechanisms of plants. In order to more accurately identify vacuole proteins, this study developed a new predictive model PEL-PVP based on ESM-2. Through this study, the feasibility and effectiveness of using advanced pre-training models and fine-tuning techniques for bioinformatics tasks were demonstrated, providing new methods and ideas for plant vacuolar protein research. In addition, previous datasets for vacuolar proteins were balanced, but imbalance is more closely related to the actual situation. Therefore, this study constructed an imbalanced dataset UB-PVP from the UniProt database，helping the model better adapt to the complexity and uncertainty in real environments, thereby improving the model's generalization ability and practicality. The experimental results show that compared with existing recognition techniques, achieving significant improvements in multiple indicators, with 6.08 %, 13.51 %, 11.9 %, and 5 % improvements in ACC, SP, MCC, and AUC, respectively. The accuracy reaches 94.59 %, significantly higher than the previous best model GraphIdn. This provides an efficient and precise tool for the study of plant vacuole proteins.

Hansenula polymorpha methanol metabolism genes enhance recombinant protein production in Komagataella phaffi.

Recombinant protein production in Komagataella phaffi (K. phaffi), a widely utilized host organism, can be optimized by enhancing the metabolic flux in the central carbon metabolism pathways. The methanol utilization pathway (MUT) during methanol-based growth plays a crucial role in providing precursors and energy for cell growth and development. This study investigated the impact of boosting the methanol dissimilation pathway, a branch of MUT that plays a vital role in detoxifying formaldehyde and providing energy in the form of NADH, in K. phaffi. This was achieved by integrating two orthologous genes from Hansenula polymorpha into the K. phaffi genome: formaldehyde dehydrogenase (HpFLD) and formate dehydrogenase (HpFMDH). The HpFLD and HpFMDH genes were isolated from the Hansenula polymorpha genome and inserted under the regulation of the pAOX1 promoter in the genome of recombinant K. phaffi that already contained a single copy of model protein genes (eGFP or EGII). The expression levels of these model proteins were assessed through protein activity assays and gene expression analysis. The findings revealed that while both orthologous genes positively influenced model protein production, HpFMDH exhibited a more pronounced upregulation in expression compared to HpFLD. Co-expression of both orthologous genes demonstrated synergistic effects, resulting in approximately a twofold increase in the levels of the model proteins detected. This study provides valuable insights into enhancing the production capacity of recombinant proteins in K. phaffi.

Protein blend extrusion: Crafting meat analogues with varied textural structures and characteristics.

With an increasing emphasis on health and environmental consciousness, there is a growing inclination toward plant protein-based meat substitutes as viable alternatives to animal meat. In the pursuit of creating diverse and functional plant protein-based substitutes, innovative plant proteins have been introduced in conjunction with soy protein isolate (SPI), encompassing pea protein isolate (PPI), rice bran protein (RBP), fava bean protein isolate (FPI), and spirulina protein isolate (SPPI). Notably, SPI-WG extrudates and SPI-PPI extrudates exhibited superior fiber structures (fiber degrees were 1.72 and 1.88, respectively), with coarse fibers in SPI-WG extrudates and fine, dense fibers in SPI-PPI extrudates. The addition of RBP, FPI and SPPI had minimal effect on fiber structure. Fresh SPI-FPI displayed the slowest rate of water loss, losing about 7.11% of their total weight in 5 h. Different plant proteins can be selected for the preparation of plant protein-based meat substitutes according to practical needs.