High Amino Acid Research Articles

Bacillus velezensis strain NA16 shows high poultry feather-degrading efficiency, protease and amino acid production

Isolated Bacillus velezensis strain NA16, which produces proteases, amino acids and the transcription levels of different keratinolytic enzymes and disulfide reductase genes in whole gene sequencing, was evaluated during feather degradation. The result shows under optimum fermentation conditions, chicken feather fermentation showed total amino acid concentration of 7599 mg/L, degradation efficiency of 99.3% at 72 h, and protease activity of 1058 U/mL and keratinase activity of 288 U/mL at 48 h. Goose feather fermentation showed total amino acid concentration of 4918 mg/L (96 h), and degradation efficiency was 98.9% at 120 h. Chicken feather fermentation broth at 72 h showed high levels of 17 amino acids, particularly phenylalanine (1050 ± 1.90 mg/L), valine (960 ± 1.04 mg/L), and glutamic (950 ± 3.00 mg/L). Scanning electron microscopy and Fourier transform infrared analysis revealed the essential role of peptide bond cleavage in structural changes and degradation of feathers. Protein purification and zymographic analyses revealed a key role in feather degradation of the 39-kDa protein encoded by gene1031, identified as an S8 family serine peptidase. Whole genome sequencing of NA16 revealed 26 metalloproteinase genes and 22 serine protease genes. Among the proteins, S8 family serine peptidase (gene1031, gene1428) and S9 family peptidase (gene3132) were shown by transcription analysis to play major roles in chicken feather degradation. These findings revealed the transcription levels of different families of keratinolytic enzymes in the degradation of feather keratin by microorganisms, and suggested potential applications of NA16 in feather waste management and amino acid production.

Anti-diabetic Potentials of Cookies from Wheat, Sweet Potato and African Yam Beans (Sphenostylis stenocarpa) Composite Flour Blends

The consumption of cookies offered reliable advantages to the consumers, which included but not limited to perfect eating ability, prolong shelf live and acceptability among all nations. This study thus, investigated the antioxidant and anti-diabetic properties of cookies obtained from wheat-African yam bean-sweet potato composite flour blends at different ratios. The African yam bean and sweet potato were thoroughly processed, milled, sieved thereafter mixed with the commercial wheat flour. Four blends were prepared by using the trial mixing ratio and 100% wheat flour and later used to bake cookies. The proximate, amino acid compositions, antioxidant, amylase and glucosidase inhibitory activities as well as physical properties of the composite cookies were examined while panelists were assigned to assess the cookie samples. The results of the proximate composition showed that the composite flour technology had significant (p<0.05) positive effects on the cookies. Meanwhile, the amino acid compositions of the cookies revealed that the cookie samples were of high aromatic and hydrophobic amino acids, respectively with optimum physical properties. No panelist showed a total dislike for the taste of any of the samples. The added soybean seeds flours showed no significant (p<0.05) effect on the acceptability and preference of the samples. Hence, it is possible to produce better protein and high nutritional cookies from these flour blends. We therefore concluded that the cookies rich in antioxidants and anti-diabetic potentials could be produced from wheat-African yam bean-sweet potato composite flour.

Non-covalent interactions of roselle anthocyanins with milk proteins and egg white protein

Roselle (Hibiscus sabdariffa L.) anthocyanins (RA) provides limited stability against hydration and pH changes during digestion which limit their bio-accessibility. The formation of protein-anthocyanin complexes offers a promising strategy for maintaining the stability of RA. Animal proteins are potential candidates for these interactions due to their high digestibility and balanced essential amino acid profile. In this study, we investigated the interactions of RA with animal proteins, specifically whey protein (WHEY) and casein (CAS) from milk, as well as ovalbumin (OVA) from egg white. Highly purified RA was obtained using XAD-7HP, SCX, and C18 column chromatography to facilitate the study of protein-anthocyanin interactions. Fluorescence spectra indicate that RA bound to the three proteins, leading to the quenching of their intrinsic fluorescence. UV–visible absorption spectra reveal changes in the polypeptide skeletons of the proteins after interacting with RA. Circular dichroism (CD) spectra show alterations in the secondary structure contents of α-helix, β-sheet, β-turn, and random coil upon interaction with RA. Molecular docking studies elucidate the docking active sites of proteins with RA. These interactions resulted in enhanced stability of RA during in vitro simulated digestion for all three proteins. Whey protein emerges as the most promising candidate for enhancing the stability of RA, thereby contributing to the potential application of RA in various food products. Our research provides a theoretical foundation for the utilization of WHEY-RA complexes in beverages and dairy products.

High-energy and high-amino acid diet enhances production performance and antioxidant capacity in yellow-feathered broilers under heat stress

This study investigated the ameliorating effects of high-energy and high-amino acid (HEHA) diets on heat stress (HS) in yellow-feathered broilers. Broilers aged 35 d were randomly assigned to 3 groups: control and HS groups fed the basic normal diet, and the HEHA group fed the HEHA diet (basal diet + 100 kcal/kg AME + 15 % DAAs). The HS and HEHA groups were exposed to cyclic HS (30 ± 1 to 34 ± 1 ℃) for 2 wk, while the control group was maintained at 26 ± 1 ℃. The results indicated that the HEHA diet significantly alleviated HS-induced feed intake and body weight loss. HEHA feeding mitigated the increase in body temperature during HS. Compared with observations in the HS group, the HEHA diet reduced the levels of ALT, Alb, and corticosterone in the serum and downregulated the gene expression of HSP27 and HSP60 in the liver. Moreover, the HEHA group showed higher GSH-px activity in the serum and SOD and GSH-Px activity in the jejunal mucosa than that of the HS group. HEHA supplementation also reduced MDA levels in the liver. In conclusion, the HEHA diet improved the production performance of broilers under HS by increasing their antioxidant capacities. These findings suggest an effective strategy to combat HS in poultry production.

Emulsion stability enhancement against storage and environment stresses using complex plant protein and betanin

Complex plant protein has been proved to be a novel protein with high solubility and comprehensive amino acids. The ability to form and stabilize oil-in-water emulsions of rice protein-pea protein and rice protein-soy protein isolate (RP-PP and RP-SPI) prepared through pH-driven method was investigated. Complex plant protein emulsions, especially RP-SPI emulsion, showed better performance on storage stability and environment stress stability than the single plant protein emulsion. According to laser confocal microscopy and particle size analysis, single protein emulsions aggregated rapidly in a short time, and the particle size increased significantly, while the complex plant protein emulsion maintained good dispersibility within 42 days, and the particle size remained around 570 nm. Furthermore, the bioactive natural pigment betanin was added to explore its effect on the stability of the RP-SPI emulsion. The addition of betanin caused a significant reduction in the surface hydrophobicity of RP-SPI and a promotion in the adsorption of protein on the oil-water interface. Betanin have a tendency to enhance the stability of the emulsion with pleasant red color. This study may provide a promising way to develop functional plant-based food emulsion.

Morphometric analysis of the effects of high molecular weight sodium hyaluronate and amino acids mixture on face rejuvenation.

Skin aging can be observed at various levels in the epidermis, dermis, and dermo-epidermal junction. Reducing the cosmetic effects of skin aging in the facial region is a widespread demand due to common aesthetic concerns. Consequently, many injectable products on the market promise antiaging effects and cosmetic improvements. We aimed to evaluate the cosmetic efficacy of a high molecular weight sodium hyaluronate and amino acids mixture for the facial region using morphometric analysis. This study evaluates the morphometric effectiveness of an injectable mixture (high molecular weight sodium hyaluronate, glycine, L-Proline, L-leucine, L-lysine HCL, L-valine, and L-alanine collagen active ingredient) on the mid-face and jawline in women aged 30-55. We used morphological measurements and digital image data to assess changes and determine effectiveness. Various computational methods were applied simultaneously with statistical tests for validation. The hydration assessment showed a significant increase on both sides of the face. A noticeable decrease was observed in gonion angle, bitragion breadth, bigonion breadth, and marionette wrinkle scale. These results suggest combining mechanical and chemical stimulation from the injection and its components (hyaluronic and amino acids) effectively enhances skin quality. The study indicates that the mechanical stimulation of the injection improves skin quality. Combining hyaluronic and amino acids (collagen, elastin, and pro-synthetic) is a safe and effective alternative for antiaging treatments.

Production of Toxins by the Gut Microbiota: The Role of Dietary Protein.

This narrative review will discuss how the intake of specific protein sources (animal and vegetable) providing specific amino acids can modulate the gut microbiota composition and generate toxins. A better understanding of these interactions could lead to more appropriate dietary recommendations to improve gut health and mitigate the risk of complications promoted by the toxic metabolites formed by the gut microbiota. Gut microbiota is vital in maintaining human health by influencing immune function and key metabolic pathways. Under unfavorable conditions, the gut microbiota can produce excess toxins, which contribute to inflammation and the breakdown of the integrity of the intestinal barrier. Genetic and environmental factors influence gut microbiota diversity, with diet playing a crucial role. Emerging evidence indicates that the gut microbiota significantly metabolizes amino acids from dietary proteins, producing various metabolites with beneficial and harmful effects. Amino acids such as choline, betaine, l-carnitine, tyrosine, phenylalanine, and tryptophan can increase the production of uremic toxins when metabolized by intestinal bacteria. The type of food source that provides these amino acids affects the production of toxins. Plant-based diets and dietary fiber are associated with lower toxin formation than animal-based diets due to the high amino acid precursors in animal proteins.

Comprehensive genetic analysis of the first near-complete genome of bovine coronavirus and partial genome of bovine rotavirus in Türkiye through metagenomics.

Obtaining the complete or near-complete genome sequence of pathogens is becoming increasingly crucial for epidemiology, virology, clinical science and practice. This study aimed to detect viruses and conduct genetic characterization of genomes using metagenomics in order to identify the viral agents responsible for a calf's diarrhoea. The findings showed that bovine coronavirus (BCoV) and bovine rotavirus (BRV) are the primary viral agents responsible for the calf's diarrhoea. The current study successfully obtained the first-ever near-complete genome sequence of a bovine coronavirus (BCoV) from Türkiye. The G+C content was 36.31% and the genetic analysis revealed that the Turkish BCoV strain is closely related to respiratory BCoV strains from France and Ireland, with high nucleotide sequence and amino acid identity and similarity. In the present study, analysis of the S protein of the Turkish BCoV strain revealed the presence of 13 amino acid insertions, one of which was found to be shared with the French respiratory BCoV. The study also identified a BRV strain through metagenomic analysis and detected multiple mutations within the structural and non-structural proteins of the BRV strain, suggesting that the BRV Kirikkale strain may serve as an ancestor for reassortants with interspecies transmission, especially involving rotaviruses that infect rabbits and giraffes.

Investigation of genetic diversity in lentil genotypes obtain from different countries using SDS-PAGE methodology

Pulses are an important component of the agricultural production system, providing high-quality food to millions of people worldwide. Among pulses, lentil is an annual and self-pollinated crop known to be its high proteins, amino acids, dietary fibers and low fat in the world. In the current study, the characterization of 44 lentil genotypes was performed using SDS-PAGE (Sodium dodecyl sulfate-polyacrylamide gel electrophoresis) technique. The protein bands were scored according to a binary system as the present (1) or absent (0) that ranged 12-20 bands in lentil genotypes. The Jaccard’s coefficient of genetic dissimilarity (GD) was measured, and the mean GD was 0.216. Maximum genetic distance was found as 0.526 between Syria2 with Iraq3 and these genotypes might be recommended for future lentil breeding. AMOVA explored the presence of higher genetic variety within genotypes (83%) than among genotypes (17% variations). STRUCTURE algorithm separated lentil genotypes into two groups mainly on the basis of their geographic. The UPGMA clustering separated lentil genotypes into two main clusters A and B. The PCoA was also conducted to confirm the results of structure and UPGMA analyses. Findigs from the PCoA and structure analyses were in full agreement with those obtained by UPGMA. The results might be useful for researchers worldwide who are interested in lentil breeding.

Comparative Metabolomics and Transcriptome Studies of Two Forms of Rhododendron chrysanthum Pall. under UV-B Stress.

Rhododendron chrysanthum Pall. (R. chrysanthum), a plant with UV-B resistance mechanisms that can adapt to alpine environments, has gained attention as an important plant resource with the ability to cope with UV-B stress. In this experiment, R. chrysanthums derived from the same origin were migrated to different culture environments (artificial climate chamber and intelligent artificial incubator) to obtain two forms of R. chrysanthum. After UV-B irradiation, 404 metabolites and 93,034 unigenes were detected. Twenty-six of these different metabolites were classified as UV-B-responsive metabolites. Glyceric acid is used as a potential UV-B stress biomarker. The domesticated Rhododendron chrysanthum Pall. had high amino acid and SOD contents. The study shows that the domesticated Rhododendron chrysanthum Pall. has significant UV-B resistance. The transcriptomics results show that the trends of DEGs after UV-B radiation were similar for both forms of R. chrysanthum: cellular process and metabolic process accounted for a higher proportion in biological processes, cellular anatomical entity accounted for the highest proportion in the cellular component, and catalytic activity and binding accounted for the highest proportion in the molecular function category. Through comparative study, the forms of metabolites resistant to UV-B stress in plants can be reflected, and UV-B radiation absorption complexes can be screened for application in future specific practices. Moreover, by comparing the differences in response to UV-B stress between the two forms of R. chrysanthum, references can be provided for cultivating domesticated plants with UV-B stress resistance characteristics. Research on the complex mechanism of plant adaptation to UV-B will be aided by these results.

Extraction and purification of seaweed protein from Ulva sp. - Challenges to overcome

Ulva sp. has gained attention as a promising and sustainable protein source, due to its high essential amino acid content. This study investigated traditional extraction methods using a double screw press followed by acid or heat precipitation. Furthermore, the screw pressing was combined by a novel purification method that combines mild heat treatment with microfiltration for removal of chlorophyll, to produce white Ulva protein. The double screw press followed by acid precipitation or heat denaturation showed low total protein yields, with 14% of the biomass protein extracted, of which half was soluble. The highest total protein yield (5.5%) was achieved with acid precipitation at pH 2, while heat denaturation at 90 °C resulted in a total protein yield of 4.1%. Acid precipitation proved to be effective with ∼ 81% precipitation of solubilized protein compared to ∼ 43% for heat denaturation. While microfiltration effectively retained chlorophyll in the retentate, the recovery of protein in the permeate remained challenging. SDS-PAGE analysis of proteins in different microfiltration-fractions showed that larger protein complexes (>250 kDa) were unable to transmit through the membranes, resulting in a maximum recovery of 24.9% of the feed-protein in the permeate. Further refinement of Ulva fractionation protocols is imperative for improvement.

Characterization of Heat-labile Uracil-DNA Glycosylase from Oncorhynchus mykiss and its Application for Carry-over Contamination Control in RT-qPCR.

Heat-labile uracil-DNA glycosylase (HL-UDG) is commonly employed to eliminate carry-over contamination in DNA amplifications. However, the prevailing HL-UDG is markedly inactivated at 50°C, rendering it unsuitable for specific one-step RT-qPCR protocols utilizing reverse transcriptase at an optimal temperature of 42°C. This study aimed to explore novel HL-UDG with lower inactivation temperature and for recombinant expression. The gene encoding an HL-UDG was cloned from the cold-water fish rainbow trout (Oncorhynchus mykiss) and expressed in Escherichia coli with high yield. The thermostability of this enzyme and other enzymatic characteristics were thoroughly examined. The novel HL-UDG was then applied for controlling carry-over contamination in one-step RT-qPCR. This recombinantly expressed truncated HL-UDG of rainbow trout (OmUDG) exhibited high amino acids similarity (84.1% identity) to recombinant Atlantic cod UDG (rcUDG) and was easily denatured at 40°C. The optimal pH of OmUDG was 8.0, and the optimal concentrations of both Na+ and K+ were 10 mM. Since its inactivation temperature was lower than that of rcUDG, the OmUDG could be used to eliminate carry-over contamination in one-step RT-qPCR with moderate reverse transcription temperature. We successfully identified and recombinantly expressed a novel HL-UDG with an inactivation temperature of 40°C. It is suitable for eliminating carry-over contamination in one-step RT-qPCR.

Investigation on adaptations of broiler chickens to high dietary free amino acid levels in nitrogen utilisation and plasma amino acid concentrations

ABSTRACT 1. A reduction in crude protein (CP) in feed for broiler chickens necessitates elevated free amino acid (AA) levels to meet the requirement of each AA. This study investigated adaptations following a change to diets with increasing free AA concentrations and possible reasons for the limitation caused by the inclusion of more free AA. 2. Male Ross 308 broiler hatchlings received a starter diet (164 g CP/kg containing 80 g/kg soy protein isolate (SPI)) until d 7. From d 7–22, birds received a diet almost identical to the starter diet or two other diets, where 50% or 100% of digestible AA in SPI were substituted with a free AA mixture. Birds were allocated to metabolism units located in the same barn to determine performance (n = 7 units) and blood traits (n = 14 birds). Total excreta collection was performed on d 7–8, 8–9, 9–10, 11–12, 14–15 and 21–22. Blood samples were collected on d 7, 8, 9, 11, 14 and 21. 3. Average daily weight gain (ADG) and average daily feed intake (ADFI) was unaffected at 50% AA substitution but decreased at 100% AA substitution on d 7–22 (p ≤ 0.001). The 100% substitution led to a decline in ADG and ADFI consistently on all days (p ≤ 0.037) except on d 11–12. A 50% AA substitution resulted in lower ADFI on d 7–8 and 14–15 (p ≤ 0.032). Nitrogen utilisation efficiency (NUE) was on a level of ~ 0.74 and was only affected by treatment up to d 11–12 (p ≤ 0.008). Concentrations of 10, 9, 8, 10 and 4 plasma free AA were affected on d 8, 9, 11, 14 and 21, respectively (p ≤ 0.037). 4. Following a change to diets containing high levels of free AA, NUE and free AA concentrations in the circulation became more balanced within 3 to 7 d. The results suggested that peptide-bound and free AA did not cause different NUE, particularly 3 and 7 d after the diet change.

Composites flour formulation made from yellow pumpkin, purple sweet potato, corn, and wolf-herring flour for replacement of wheat flour on low- and high- moisture foods part I: Cookies and muffin

IntroductionThe study investigated the potential of yellow pumpkin, purple sweet potato, corn, and wolf-herring flour as local-based ingredients in generating cookies and muffin rich in minerals and amino acids. MethodsFive different composites were prepared and applied for cookies and muffins preparation replacing wheat flour. Sensory acceptability was conducted, and selected formulas were further evaluated for their chemical composition including proximate, dietary fiber, minerals (Mg, P, Fe, Zn and Ca) and amino acids. Results and discussionA formula consisting of 45 g yellow pumpkin-, 45 g purple sweet potato-, 5 g corn- and 5 g wolf-herring flour was selected as the highest acceptability. Compared to muffins, cookies had a higher level of fat (21.6%), protein (8.6%), ash (2.9%), dietary fiber (14.4%), calorie (462.6 cal/100 g), Mg (47.9 mg/100 g), P (1725.9 mg/kg), Fe (2.0 mg/100 g), Zn (0.9 mg/100 g) and Ca (136.8 mg/100 g). Cookies and muffins contained 15 L-amino acids in which 7 were amino acids essentials. The higher amino acids were L-aspartic acid and L-glutamat acids ranged between 6.9 – 8.1 and 8.9 – 9.5 g/kg, respectively. The lowest amino acids obtained was L-Histidine at 1.9 g/kg for cookies and muffin. ConclusionIn conclusion, cookies, and muffin rich in mineral and amino acids can be formulated based on low-cost locally available food ingredients.

Identification and Genome Characterization of a Novel Virus within the Genus Totivirus from Chinese Bayberry (Myrica rubra).

Chinese bayberry (Myrica rubra) is an economically significant fruit tree native to eastern Asia and widely planted in south-central China. However, studies about the viruses infecting M. rubra remain largely lacking. In the present study, we employed the metatranscriptomic method to identify viruses in M. rubra leaves exhibiting yellowing and irregular margin symptoms collected in Fuzhou, a city located in China's Fujian province in the year 2022. As a consequence, a novel member of the genus Totivirus was identified and tentatively named "Myrica rubra associated totivirus 1" (MRaTV1). The genome sequencing of MRaTV1 was determined by overlapping reverse transcription polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE). The two deduced proteins encoded by MRaTV1 have the highest amino acid (aa) sequence identity to the coat protein (CP) and RNA-dependent RNA polymerase (RdRP) of Panax notoginseng virus A (PNVA), a member of the genus Totivirus within the family Totiviridae, at 49.7% and 61.7%, respectively. According to the results of the phylogenetic tree and the species demarcation criteria of the International Committee on Taxonomy of Viruses (ICTV) for the genus Totivirus, MRaTV1 is considered a new member of the genus Totivirus.

Metabolomics Profiles of Solid State and Submerge Fermentation of Corn Silk using Mixed Microbes

Corn silk is a waste product when harvesting corn, but it contains many bioactive compounds. This research characterized and assessed the antioxidant bioactivity of compounds isolated from corn silk. Two different biological fermentation processes were compared: Solid-state fermentation (SSF) and submerged fermentation (SmF). Fresh corn silk (Zea mays L.) from both sweet corn and feed corn were fermented separately with various microorganisms (Bacillus subtilis, Lactobacillus sp. and Saccharomyces cerevisiae). Biological compounds were identified using LC-MS/MS. The extract was then tested for total phenolic content (TPC) and total flavonoid content (TFC). Next, the extract was tested for xanthine oxidase inhibition and antioxidant inhibition using the ABTS method. Potential contamination with the heavy metals As, Pb, and Hg was checked. The results show that fermented corn silk extract contains numerous bioactive compounds and amino acids. Among the different extract process variations, the highest TPC and TFC were 6.1 ± 0.7 µg GAE/g dry wt. and 68.3 ± 1.1 µg QE/g dry wt., respectively, and the highest xanthine oxidase inhibition was 76.5 ± 6.4 %. The above highest results were obtained using SmF of sweet corn for 60 days. In contrast, the highest antioxidant activity of 66.9 % was found using SSF of sweet corn for 20 days. Neither sweet corn nor feed corn were found to contain heavy metal ions. In summary, fermented corn silk extract samples contain numerous bioactive compounds, including both flavonoid and phenolic compounds, and the extract demonstrates antioxidant activity. Corn silk extracts from fermentation thus have potential for utilization in health products. HIGHLIGHTS Feed corn silk and sweet corn silk underwent solid-state fermentation (SSF) or submerged fermentation (SmF) for extraction using heterofermentative microbes of Bacillus subtilis, Lactobacillus and Saccharomyces cerevisiae to produce the high bioactive compounds, amino acids, and organic acids Metabolomic analysis of the fermentation extracts was carried out by LC-ESI-QTOF-MS/MS (Liquid Chromatography-Mass Spectrometry) using the heatmaps generated metabolites profiles The main amino acids found in the extractions were phenylalanine, leucine, and their derivatives SmF of sweet corn silk yielded the highest xanthine oxidase inhibitory activity (76 %) SSF of feed corn silk and sweet corn silk displayed the high antioxidant activity GRAPHICAL ABSTRACT

Microalgae as fishmeal alternatives in aquaculture: current status, existing problems, and possible solutions.

Fishmeal is an indispensable ingredient for most aquatic animals. However, the finite supply and escalating price of fishmeal seriously limit its use in aquaculture. Thus the development of new, sustainable protein ingredients has been a research focus. Microalgae are potential fishmeal alternatives owing to their high protein content and balanced amino acid profile. Studies suggest that suitable replacement of fishmeal with microalgae is beneficial for fish growth performance, but excessive replacement would induce poor growth and feed utilization. Therefore, this paper aims to review research on the maximum substitutional level of fishmeal by microalgae and propose the main issues and possible solutions for fishmeal replacement by microalgae. The maximum replacement level is affected by microalgal species, fish feeding habits, quality of fishmeal and microalgal meals, and supplemental levels of fishmeal in the control group. Microalgae could generally replace 100%, 95%, 95%, 64.1%, 25.6%, and 18.6% fishmeal protein in diets of carp, shrimp, catfish, tilapia, marine fish, and salmon and trout, respectively. The main issues with fishmeal replacement using microalgae include low production and high production cost, poor digestibility, and anti-nutritional factors. Possible solutions to these problems are recommended in this paper. Overall, microalgae are promising fishmeal alternatives in aquaculture.

An LQT2-related mutation in the voltage-sensing domain is involved in switching the gating polarity of hERG

BackgroundCyclic Nucleotide-Binding Domain (CNBD)-family channels display distinct voltage-sensing properties despite sharing sequence and structural similarity. For example, the human Ether-a-go-go Related Gene (hERG) channel and the Hyperpolarization-activated Cyclic Nucleotide-gated (HCN) channel share high amino acid sequence similarity and identical domain structures. hERG conducts outward current and is activated by positive membrane potentials (depolarization), whereas HCN conducts inward current and is activated by negative membrane potentials (hyperpolarization). The structural basis for the “opposite” voltage-sensing properties of hERG and HCN remains unknown.ResultsWe found the voltage-sensing domain (VSD) involves in modulating the gating polarity of hERG. We identified that a long-QT syndrome type 2-related mutation within the VSD, K525N, mediated an inwardly rectifying non-deactivating current, perturbing the channel closure, but sparing the open state and inactivated state. K525N rescued the current of a non-functional mutation in the pore helix region (F627Y) of hERG. K525N&F627Y switched hERG into a hyperpolarization-activated channel. The reactivated inward current induced by hyperpolarization mediated by K525N&F627Y can be inhibited by E-4031 and dofetilide quite well. Moreover, we report an extracellular interaction between the S1 helix and the S5-P region is crucial for modulating the gating polarity. The alanine substitution of several residues in this region (F431A, C566A, I607A, and Y611A) impaired the inward current of K525N&F627Y.ConclusionsOur data provide evidence that a potential cooperation mechanism in the extracellular vestibule of the VSD and the PD would determine the gating polarity in hERG.

Evolutionary analysis of seasonal influenza A viruses in Pakistan 2020-2023.

Influenza A viruses cause global health concerns due to their high amino acid substitution rates. They are linked to yearly seasonal epidemics and occasional pandemics. This study focused on sequencing influenza A virus strains in Pakistan. We analyzed the genetic characteristics of influenza A(H1N1)pdm09 and A(H3N2) viruses circulating in Pakistan from January 2020 to January 2023. Whole genome sequences from influenza A (n = 126) virus isolates were amplified and sequenced by the Oxford Nanopore (MinION) platform. The HA genes of influenza A(H1N1)pdm09 underwent amino acid substitutions at positions K54Q, A186T, Q189E, E224A, R259K, and K308R in sequenced samples. The HA genes of influenza A(H3N2) had amino acid substitutions at G53D, E83K, D104G, I140M, S205F, A212T, and K276R in the sequenced samples. Furthermore, the HA gene sequences of influenza A(H1N1)pdm09 in this study belonged to subclade 6B.1A.5a.2a. Similarly, the HA gene sequences of influenza A(H3N2) were classified under six subclades (3C.3a.1 and 3C.2a1b.2a [2, 2a.1, 2b, 2c, and 2a.3b]). Notably, amino acid substitutions in other gene segments of influenza A(H1N1)pdm09 and A(H3N2) were also found. These findings indicate influenza A(H1N1)pdm09 and A(H3N2) viruses co-circulated during the 2020-2023 influenza season in Pakistan. Continued surveillance is crucial for real-time monitoring of possible high-virulence variation and their relevance to existing vaccine strains.

Deep learning-based characterization and redesign of major potato tuber storage protein

Potato is one of the most important crops worldwide, to feed a fast-growing population. In addition to providing energy, fiber, vitamins, and minerals, potato storage proteins are considered as one of the most valuable sources of non-animal proteins due to their high essential amino acid (EAA) index. However, low tuber protein content and limited knowledge about potato storage proteins restrict their widespread utilization in the food industry. Here, we report a proof-of-concept study, using deep learning-based protein design tools, to characterize the biological and chemical characteristics of patatins, the major potato storage proteins. This knowledge was then employed to design multiple cysteines on the patatin surface to build polymers linked by disulfide bonds, which significantly improved viscidity and nutrient of potato flour dough. Our study shows that deep learning-based protein design strategies are efficient to characterize and to create novel proteins for future food sources.