β-Nicotinamide mononucleotide (NMN) serves as a direct precursor of the essential cofactor NAD⁺, playing a crucial role in regulating mitochondrial function and delaying cellular aging. Synthetic biology, with its environmental and economic advantages, has become an important approach for the production of natural medicines, with the optimized design of metabolic pathways at its core. In this study, we focused on the NMN‑consuming pathway mediated by the nma1 and pof1 genes in Saccharomyces cerevisiae and consequently generated their double‑knockout mutant, DHSX02. Futhermore, a highly efficient NMN phosphoribosyltransferase (CpNampt) derived from Chitinophaga pinensis was mutated via AI-assisted design and introduced into the DHSX02 strain to enhance NMN synthesis. The results showed that the NMN yield in the DHSX02 strain increased by 20.7% compared to the wild-type strain. After multiple rounds of optimization, the strains carrying the mutated CpNampt exhibited significantly enhanced NMN production, with the I229A mutant strain achieving a total yield of 60.64 mg/L in 12h, a 4.4-fold increase compared to the wild-type strain. This work demonstrates an effective synthetic biology strategy, combining metabolic engineering and AI-assisted enzyme design to construct an efficient microbial cell factory for green NMN production.
Background: Phycocyanin (PC), a vibrant blue accessory pigment-protein complex predominantly occurring in cyanobacteria, Rhodophyceae, and some cryptophytes, is a key nutritional component contributing to the commercial value of Spirulina as a food supplement. Beyond its nutritional value, PC has distinctive fluorescent properties and diverse physiological activities, giving it broad potential in food, cosmetics, and biomedicine. Summary: This study discusses the factors restricting PC yield and comprehensively summarizes PC extraction and purification technologies. The advantages and disadvantages of each technology are compared, and its application prospects in laboratory and industrial production are explored. Natural deep eutectic solvents, which are solvent systems formed from naturally occurring cellular metabolites such as sugars, amino acids, and organic acids, offer safety, simple preparation, low cost, and promising prospects for PC purification compared with several conventional technologies. However, the application scope of PC remains limited by its poor stability and sensitivity to temperature, light, and pH. Therefore, this review also discusses the factors affecting PC stability, strategies for improving stability, and applications of PC in food, healthcare products, and related fields. Key Messages: This comprehensive review and critical assessment of production, purification, and stabilization strategies provides valuable insights for advancing PC research and facilitating its broader, more sustainable utilization across diverse industrial sectors.
Introduction: Viroporins are small multifunctional proteins that modify cellular membranes facilitating processes such as viral nucleic acid entry and the release of virions from infected cells. We are interested in studying the evolutionary relationships among these proteins, in particular their organization into families and superfamilies. Methods: We applied a variety of computational strategies to perform comparative genomics analyses of 120 viral genomes, using the phylogenetic profile method. This allowed the identification of 12 families, organized into four functionally related groups. Additionally, we compiled a list of 40 families from the Transporter Classification Database (TCDB) with viroporin-like attributes (i.e., length ≤300 aas, similar topologies, and/or documented viroporin activities). We then used TCDB as a reference to search for evidence of homology among families. Our well-established bioinformatic pipeline for inference of homology included (1) sequence similarity, (2) compatibility of topology and hydropathy profiles, (3) similarity of family-based HMM profiles, (4) shared motifs, and (5) conserved domains. Results: We were able to infer homology among 15 families, four of which (Vpu-C, p10 viroporin/GDU1, FAST, and R-FAST) expanded the established Influenza A/B Virus M2 Protein (M2) superfamily. The other families constituted three novel superfamilies: viroporin-1, consisting of three families (RVP10, NS3, and NSP4); viroporin-2, composed of two functionally linked families (SARS-VP and M-protein); and viroporin-3 composed of 3 functionally related families (viroporin E, IBV-E, and PRRSV). Conclusion: The application of comparative genomics and remote homology identification strategies allowed the classification of homologous and functionally related viroporin-like families into superfamilies. These results will be useful in future functional, mechanistic, and evolutionary studies of viroporins.
INTRODUCTION:The transposase protein, InsAB', is the key cytoplastic determinant that catalyzes IS1 movement within a single DNA molecule or between two DNA molecules. Regulation of InsAB' synthesis, its structural characteristics, and its potent cis effect on IS1 transposition have been subjects of thorough investigation. However, limited efforts have been devoted to measuring the levels of IS1 transcripts and transposases made by native IS1 elements and examining their trans-acting function, particularly concerning their role in transposing otherwise immobile IS1 elements and other target DNA segments within the bacterial genome. METHODS:All constructs were made on the Escherichia coli chromosome using the lambda-Red system. IS1 transposition frequencies were determined using Bgl+ mutation assays, colony PCR and DNA sequencing. IS1 transcripts and transposases were quantitated using lacZ transcriptional and translational reporters. RESULTS:We first confirm that a native IS1 element, IS1E, can transpose at a dramatically elevated rate in the absence of both transcriptional and post-transcriptional regulations. Using lacZ reporters targeting the transposase gene insAB', we reveal that the InsA repressor moderately reduces insAB' transcription by as much as 16.8-fold. However, the combined action of InsA and ribosomal frameshifting leads to a much stronger reduction in InsAB' production, by up to 735-fold. We show that only a small percentage (roughly 1.6%) of IS1 transcripts are successfully translated into InsAB', highlighting the primary role of post-transcriptional regulation over transcriptional repression in governing IS1 transposition. We also demonstrate, for the first time, that high levels of InsAB' exhibit a strong trans effect, being capable of efficiently transposing not only normally non-mobile IS1 elements but also a larger miniIS1 cassette within the genome, with greater amounts of InsAB' leading to higher frequencies of transposition. CONCLUSIONS:By dissecting and quantitating the roles of transcriptional and translational controls in transposase production, our study reveals that post-transcriptional regulation via ribosomal frameshifting is the central mechanism governing IS1 intragenomic transposition. Our findings demonstrate that IS1 transposases function in trans, mediating the movement of DNA segments specifically bordered by its inverted repeat at the left end and right end sequences. These findings advance our comprehension of how IS elements mobilize within bacterial genomes and provide potential strategies to counter IS1-mediated antibiotic resistance in clinical bacterial isolates.
Background: The human microbiome is a dynamic, polymicrobial ecosystem that plays an essential role in nutrition, immune development, barrier integrity, and host physiology, acting as a mutualistic partner under balanced conditions. Summary: The ecological complexity of the human microbiome, its genetic adaptability through horizontal gene transfer, and interactions with other prokaryotes as well as protozoan and metazoan parasites can transform commensals into pathobionts, resulting in weakened host’s barriers, immunity declines with the progression of age, and community composition shifts toward dysbiosis. Factors such as diet, genetics, aging, immune-senescence, impaired autophagy, and environmental exposure, all influence this delicate balance, determining whether the microbiome remains protective or becomes an opportunistic source of inflammation and disease. Key Messages: This review focuses on the study of the intestinal microbiome in humans. Maintaining microbiome homeostasis is promoted through (a) dietary diversity, (b) limited antimicrobial use, (c) use of probiotics, (d) support for gut barrier function, and (e) healthy lifestyle improvements. These actions and considerations are critical to prevent the emergence of pathogenic states and preserving the microbiome’s vital role in host health throughout life.
