Bioscience, Biotechnology, and Biochemistry最新文献

The oleaginous yeast Lipomyces starkeyi has a high capacity for starch assimilation, but the genes involved and specific mechanisms in starch degradation remain unclear. This study aimed to identify the critical carbohydrate-active enzyme (CAZyme) genes contributing to starch degradation in L. starkeyi. Comparative transcriptome analysis of cells cultured in glucose and soluble starch medium revealed that 55 CAZymes (including transcript IDs 3772, 1803, and 7314) were highly expressed in soluble starch medium. Protein domain structure and disruption mutant analyses revealed that 3772 encodes the sole secreted α-amylase (LsAmy1p), whereas 1803 and 7314 encode secreted α-glucosidase (LsAgd1p and LsAgd2p, respectively). Triple-gene disruption exhibited severely impaired growth in soluble starch, dextrin, and raw starch media, highlighting their critical role in degrading polysaccharides composed of glucose linked by α-1,4-glucosidic bonds. This study provided insights into the complex starch degradation mechanism in L. starkeyi.

Recently, noninvasive spot hemoglobin measurement (SpHb) using Pulse CO-Oximeter Rad-67™ Spot-check (Rad-67) has been validated, although anemia diagnosis typically relies on blood hemoglobin concentration measurement. In this large-scale survey of Japanese children aged 1-5 years, we evaluated SpHb distribution to understand the prevalence of suspected anemia, and further examined the relationship between SpHb and background factors. Children were recruited from large retail stores in Japan between November 2022 and August 2023. SpHb was measured by nutritionists or registered dietitians using Rad-67. Four thousand one hundred thirty-three participants were included and stratified by age and sex. The prevalence of children below the World Health Organization threshold value for anemia was found to be 5.2% in total (ranging between 2.6% and 7.8% in subgroups). Mean SpHb values increased with age, and were higher in boys. Age and sex were independently related to SpHb. Overall, this study shows that approximately 3%-8% of young children in Japan are suspected to be anemic.

Plant growth is finely tuned by environmental changes, with abscisic acid (ABA) playing a key role in balancing stress tolerance and growth regulation. The target genes of MYB50, which regulate root growth, include genes that respond to ABA; however, the precise role of MYB50 in ABA signaling remains unclear. Therefore, this study aimed to elucidate the function of MYB50 under ABA signaling. Our experiments demonstrated that ABA treatment reduced MYB50 expression and promoted the degradation of MYB50 protein. This degradation alleviates the inhibitory effects of MYB50 on root growth. Furthermore, ABA differentially regulates MYB50 compared with ABI5, another key transcription factor involved in root growth under ABA signaling, suggesting that ABA uses distinct regulatory pathways for root growth. Our study suggests that ABA controls root growth by modulating MYB50 at both the transcriptional and post-translational levels, thus ensuring balanced root development in response to ABA.

Maillard reaction products (MRPs), including melanoidins and volatile odor compounds, are associated with distinct flavors and colors during food processing and cooking. Although MRPs have health benefits, such as antioxidant activity, they are also associated with pathophysiological effects. Several in vivo models, especially rodents, are used to demonstrate physiological effects. Caenorhabditis elegans (C. elegans), an easy-to-rear free-living nematode with a short lifespan, has been used as a promising in vivo organism for the evaluation of functional properties in food components, including antiaging, antioxidant, and antiobesity properties. Furthermore, the high olfactory discrimination of this organism allows for the basic elucidation of behavior and regulation of aging. In this minireview, I discuss the various attributes of C. elegans that make it a promising in vivo model for studying the biological effects of MRPs.

Cloning of small DNA segments has been established using Escherichia coli plasmids. The cloned DNA can be transferred to various cells using transformation. In contrast, cloning of large DNA segments of more than several hundred kilobase pairs has been limited to the Bacillus subtilis genome cloning system. The advantage of giant DNA cloned by B. subtilis is that all kinds of gene editing can be implemented by the high and strict natural transformation ability of the host. However, the following transfer step of giant synthesized and edited genomes to different cell systems requires a special system by avoiding exposure in liquid. The use of a conjugational plasmid pLS20 that was developed for 20 years improves the B. subtilis genome vector establishment process from scratch. The use of the unique B. subtilis genome vector system from synthesis to transmitting genomes is now being manipulated and summarized for the first time.

Keratinase from Nocardiopsis sp. TOA-1 (NAPase) holds significant potential for industrial and medical applications. Here, we developed a heterologous secretory expression system for NAPase in Bacillus subtilis. The recombinant enzyme exhibited catalytic properties comparable to the native enzyme, demonstrating its suitability for further protein engineering. This work provides a foundation for enhancing NAPase activity and stability, expediting its biotechnological applications.

We successfully constructed a heterologous expression system for L-glutamate oxidase from the marine actinomycete Streptomyces lydicamycinicus NBRC 110027 (Sl-LGOX) in Escherichia coli BL21(DE3) as a host. This is the first example of L-glutamate oxidase from a marine microorganism. A chemically synthesized gene optimized for codon usage in E. coli was used as the inserted fragment, which was effective for enzyme expression. We expressed Sl-LGOX in the soluble fraction of E. coli BL21(DE3)/pET21b-Sl-lgox. We also succeeded in purifying the recombinant Sl-LGOX (rSl-LGOX) to homogeneity from the cell-free extract of this clone via an Ni-NTA column. rSl-LGOX showed high specificity for L-Glu and was active and stable over a wide range of temperatures and pH values. In particular, it showed high specific activity and stability at an acidic pH. A variety of applications can take advantage of the unique enzymatic properties of rSl-LGOX.

The nematode Caenorhabditis elegans is an excellent model organism for elucidating higher life phenomena. C. elegans and humans are common in many aspects. During our research on development and life span regulation, we identified RAB-18, a small guanosine triphosphatase (GTPase) involved in the membrane trafficking of NCR-1, a cholesterol transporter mainly expressed in the intestine. We expressed the human NPC1L1, an intestinal cholesterol transporter, in mutant C. elegans lacking NCR-1. NPC1L1-expressing animals revealed almost the same larval diapause in the presence of a diapause-inducing pheromone and lipid droplets containing cholesterol as in wild-type C. elegans. This result indicates that C. elegans NCR-1 and human NPC1L1 are exchangeable and that C. elegans RAB-18 transports human NPC1L1 to the apical membrane in the C. elegans intestine. This transgenic C. elegans could be adapted to evaluate functional foods and ingredients regarding cholesterol absorption.

Angiogenin (Ang), an endoribonuclease belonging to the RNase A superfamily, cleaves the anticodon-loops of tRNAs to produce tRNA-half molecules. Although previous studies have demonstrated the involvement of Ang in the pathobiology of neurodegenerative disorders, the characterization of Ang-generated tRNA halves in neuronal cells remains limited. This is partly due to the technical limitations of standard RNA-seq methods, which cannot capture Ang-generated RNAs containing a 2',3'-cyclic phosphate (cP). In this report, we established an Ang-treatment model using SH-SY5Y, a human neuroblastoma cell line, and demonstrated Ang-dependent accumulation of tRNA halves. By performing cP-RNA-seq, which selectively captures cP-containing RNAs, we identified Ang-generated tRNA halves and the specific cleavage positions within tRNA anticodon-loops responsible for their generation. Our results provide insights into the anticodon-loop cleavage and the selective production of a specific subset of tRNA halves by Ang.

Circadian rhythms are biological systems that provide approximately 24-h cycles for the behavior and physiological functions of organisms. As diverse modern lifestyles often cause disturbances in circadian rhythms, new approaches to their regulation are required. Therefore, new compounds that affect circadian rhythms have been explored in edible mushrooms. The extract from the culture filtrate of Cyclocybe cf. erebia showed activity that advanced the circadian rhythm in a bioassay with mouse fibroblasts expressing the LUCIFERASE protein under the control of the Period2 promoter. Bioassay-guided fractionation of the extract resulted in the isolation of the compound. Spectroscopic analyses identified the compound as a phthalide derivative, and the compound was named cyclocybelide. Treatment of mouse fibroblasts with the compound shifted the circadian rhythm forward, irrespective of the timing of treatment. In addition, some phthalide derivatives with hydroxy and methoxy groups showed similar effects on circadian rhythms.