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PloS One 2024Polymeric IgMs are secreted from plasma cells abundantly despite their structural complexity and intricate multimerization steps. To gain insights into IgM's assembly...
Polymeric IgMs are secreted from plasma cells abundantly despite their structural complexity and intricate multimerization steps. To gain insights into IgM's assembly mechanics that underwrite such high-level secretion, we characterized the biosynthetic process of a natural human IgM, SAM-6, using a heterologous HEK293(6E) cell platform that allowed the production of IgMs both in hexameric and pentameric forms in a controlled fashion. By creating a series of mutant subunits that differentially disrupt secretion, folding, and specific inter-chain disulfide bond formation, we assessed their effects on various aspects of IgM biosynthesis in 57 different subunit chain combinations, both in hexameric and pentameric formats. The mutations caused a spectrum of changes in steady-state subcellular subunit distribution, ER-associated inclusion body formation, intracellular subunit detergent solubility, covalent assembly, secreted IgM product quality, and secretion output. Some mutations produced differential effects on product quality depending on whether the mutation was introduced to hexameric IgM or pentameric IgM. Through this systematic combinatorial approach, we consolidate diverse overlapping knowledge on IgM biosynthesis for both hexamers and pentamers, while unexpectedly revealing that the loss of certain inter-chain disulfide bonds, including the one between μHC and λLC, is tolerated in polymeric IgM assembly and secretion. The findings highlight the differential roles of underlying non-covalent protein-protein interactions in hexamers and pentamers when orchestrating the initial subunit interactions and maintaining the polymeric IgM product integrity during ER quality control steps, secretory pathway trafficking, and secretion.
Topics: Humans; Immunoglobulin M; HEK293 Cells; Mutation; Protein Multimerization; Protein Subunits; Endoplasmic Reticulum
PubMed: 38848420
DOI: 10.1371/journal.pone.0291568 -
Applied Microbiology and Biotechnology Jun 2024Lipases are important biocatalysts and ubiquitous in plants, animals, and microorganisms. The high growth rates of microorganisms with low production costs have enabled...
Lipases are important biocatalysts and ubiquitous in plants, animals, and microorganisms. The high growth rates of microorganisms with low production costs have enabled the wide application of microbial lipases in detergent, food, and cosmetic industries. Herein, a novel lipase from Lacticaseibacillus rhamnosus IDCC 3201 (Lac-Rh) was isolated and its activity analyzed under a range of reaction conditions to evaluate its potential industrial application. The isolated Lac-Rh showed a molecular weight of 24 kDa and a maximum activity of 3438.5 ± 1.8 U/mg protein at 60 °C and pH 8. Additionally, Lac-Rh retained activity in alkaline conditions and in 10% v/v concentrations of organic solvents, including glycerol and acetone. Interestingly, after pre-incubation in the presence of multiple commercial detergents, Lac-Rh maintained over 80% of its activity and the stains from cotton were successfully removed under a simulated laundry setting. Overall, the purified lipase from L. rhamnosus IDCC 3201 has potential for use as a detergent in industrial applications. KEY POINTS: • A novel lipase (Lac-Rh) was isolated from Lacticaseibacillus rhamnosus IDCC 3201 • Purified Lac-Rh exhibited its highest activity at a temperature of 60 °C and a pH of 8, respectively • Lac-Rh remains stable in commercial laundry detergent and enhances washing performance.
Topics: Lipase; Lacticaseibacillus rhamnosus; Hydrogen-Ion Concentration; Detergents; Enzyme Stability; Temperature; Molecular Weight; Bacterial Proteins
PubMed: 38842543
DOI: 10.1007/s00253-024-13185-4 -
Frontiers in Microbiology 2024The efficient utilization of straw resources as animal feed has gained considerable attention. The objective of this study was to evaluate whether treatment alters the...
INTRODUCTION
The efficient utilization of straw resources as animal feed has gained considerable attention. The objective of this study was to evaluate whether treatment alters the chemical composition and antioxidant activity of highland barley straw and enhances its functional value as a ruminant feed.
METHODS
The chemical composition, antioxidant capacity, and metabolomic profile of highland barley straw were determined after 21 days of solid-state fermentation with at 25°C. The differential metabolites between fermented and unfermented highland barley straw were identified by LC-MS and the relationship between the identified metabolites and antioxidant capacity was elucidated.
RESULTS
The results showed that, compared with untreated highland barley straw, the crude protein and ether extract contents were higher (51.55 and 76.43%, respectively) in highland barley straw after 21 days of incubation with , whereas the hemicellulose, cellulose, and acid detergent lignin contents were lower (2.48, 25.08, and 45%, respectively). The total antioxidant capacity was significantly higher in -treated than in untreated highland barley straw. In total, 600 differential metabolites (301 upregulated and 299 downregulated) were identified between -fermented and unfermented highland barley straw. Correlation analysis results showed that Fe scavenging and total phenolic content were strongly correlated with total antioxidant capacity. Meanwhile, the differential flavonoid metabolites between fermented and unfermented highland barley straw were primarily associated with antioxidant activity, with kaempferol 3-xylosylglucoside, isoginkgetin, and rhoifolin being the most representative.
CONCLUSION
Thus, this study demonstrates that could enhance the functional value of highland barley straw, showing the potential of for improving the utilization of agricultural straws in ruminants.
PubMed: 38841071
DOI: 10.3389/fmicb.2024.1365254 -
Journal of Molecular and Cellular... Jun 2024The HSP70 co-chaperone BAG3 targets unfolded proteins to degradation via chaperone assisted selective autophagy (CASA), thereby playing pivotal roles in the proteostasis...
The HSP70 co-chaperone BAG3 targets unfolded proteins to degradation via chaperone assisted selective autophagy (CASA), thereby playing pivotal roles in the proteostasis of adult cardiomyocytes (CMs). However, the complex functions of BAG3 for regulating autophagy in cardiac disease are not completely understood. Here, we demonstrate that conditional inactivation of Bag3 in murine CMs leads to age-dependent dysregulation of autophagy, associated with progressive cardiomyopathy. Surprisingly, Bag3-deficient CMs show increased canonical and non-canonical autophagic flux in the juvenile period when first signs of cardiac dysfunction appear, but reduced autophagy during later stages of the disease. Juvenile Bag3-deficient CMs are characterized by decreased levels of soluble proteins involved in synchronous contraction of the heart, including the gap junction protein Connexin 43 (CX43). Reiterative administration of chloroquine (CQ), an inhibitor of canonical and non-canonical autophagy, but not inactivation of Atg5, restores normal concentrations of soluble cardiac proteins in juvenile Bag3-deficient CMs without an increase of detergent-insoluble proteins, leading to complete recovery of early-stage cardiac dysfunction in Bag3-deficient mice. We conclude that loss of Bag3 in CMs leads to age-dependent differences in autophagy and cardiac dysfunction. Increased non-canonical autophagic flux in the juvenile period removes soluble proteins involved in cardiac contraction, leading to early-stage cardiomyopathy, which is prevented by reiterative CQ treatment.
PubMed: 38838815
DOI: 10.1016/j.yjmcc.2024.06.001 -
PloS One 2024The purpose of the current study was to evaluate the impact of various doses of microencapsulated lemongrass and mangosteen peel (MELM) on gas dynamics, rumen...
Microencapsulation of lemongrass and mangosteen peel as phytogenic compounds to gas kinetics, fermentation, degradability, methane production, and microbial population using in vitro gas technique.
The purpose of the current study was to evaluate the impact of various doses of microencapsulated lemongrass and mangosteen peel (MELM) on gas dynamics, rumen fermentation, degradability, methane production, and microbial population in in vitro gas experiments. With five levels of microencapsulated-phytonutrient supplementation at 0, 1, 2, 3, and 4% of substrate, 0.5 g of roughage, and a concentrate ratio of 60:40, the trial was set up as a completely randomized design. Under investigation, the amount of final asymptotic gas volume was corresponding responded to completely digested substrate (b) increased cubically as a result of the addition of MELM (P < 0.01) and a cubic rise in cumulative gas output. The amount of MELM form did not change the pH and NH3-N concentration of the rumen after 12 and 24 h of incubation. However, methane production during 24 h of incubation, the levels were cubically decreased with further doses of MELM (P < 0.01) at 12 h of incubation. Increasing the dosage of MELM supplementation at 2% DM resulted in a significant increase in the digestibility of in vitro neutral detergent fiber (IVNDF) and in vitro true digestibility (IVTD) at various incubation times (P < 0.05), but decreased above 3% DM supplementations. Moreover, the concentration of propionic acid (C3) exhibited the variations across the different levels of MELM (P < 0.05), with the maximum concentration obtained at 2% DM. The populations of Fibrobacter succinogenes, Ruminococcus albus, Ruminococcus flavefaciens, and Megasphaera elsdenii revealed a significant increase (P < 0.05), while the quantity of Methanobacteriales decreased linearly with increasing doses of MELM. In conclusion, the inclusion of MELM at a concentration of 2% DM in the substrate which could enhance cumulative gas production, NDF and true digestibility, C3 production, and microbial population, while reducing methane concentration and Methanobacterial abundance.
Topics: Methane; Fermentation; Animals; Rumen; Garcinia mangostana; Digestion; Animal Feed; Kinetics; Gases; Drug Compounding; Phytochemicals; Cattle
PubMed: 38837999
DOI: 10.1371/journal.pone.0304282 -
Journal of Athletic Training Jun 2024The best practice for cleaning wrestling mats is using a residual disinfectant with continued antibacterial action. Recently available wash-in silver additives claim to...
CONTEXT
The best practice for cleaning wrestling mats is using a residual disinfectant with continued antibacterial action. Recently available wash-in silver additives claim to confer a residual effect to fabric.
OBJECTIVE
To test the efficacy of laundering with a wash-in silver additive in reducing exposure of athletes to potentially infectious microbes on apparel.
DESIGN
4-part Controlled Laboratory Study/Parallel Group Comparison Study: (1) To test whether fabrics in athletic clothing would be affected differently, we applied bacteria to control fabrics washed in detergent alone and test counterparts washed in detergent plus wash-in silver additive. Bacteria were applied to fabrics, extracted, plated, incubated, and counted. (2) To see if wash-in silver affected various bacteria differently, we washed cotton t-shirts with detergent alone or with detergent plus wash-in silver. We applied four bacterial species commonly found in the wrestling environment. Bacteria were extracted, plated, incubated, and counted. (3) To see if wash-in silver was effective in reducing bacterial contamination during practice, 32 collegiate wrestlers paired off with one wearing a test silver-treated t-shirt, and their partner wearing a control shirt. Wrestler rotations exposed shirts to 2, 4 or 8 wrestlers. Identical swatches of fabric were cut from the t-shirts. Bacteria were extracted, plated, incubated, and counted. (4) We simulated prolonged/repeated bacterial exposure as occurs during tournaments by applying bacteria directly to silver-treated and untreated singlet material repeatedly over time. Test samples were taken at regular intervals to see if bacterial growth was inhibited by the presence of the silver nanoparticles. Bacteria were extracted, plated, incubated, and counted.
SETTING
Laboratory and practice.
PARTICIPANTS
Collegiate D3 Wrestling Team.
MAIN OUTCOME MEASURE(S)
Wash-in silver would be considered effective if statistically significant reduction in bacterial count was observed at 95% confidence.
RESULTS
Wash-in silver reduced bacterial growth at low levels of contamination but did not significantly reduce bacterial growth at levels seen during contact sport competitions. This was true for all bacterial species and all fabrics tested.
CONCLUSIONS
The environmental and potential health risks in using a wash-in silver nanoparticle laundry additive in the wash cycle for clothing worn by wrestlers outweigh any potential infection control benefits to these athletes. We do not currently recommend adopting wash-in silver treatment as part of the laundering regimen for wrestling programs until further testing of alternate methods of silver impregnation into sports fabrics has been investigated.
PubMed: 38835322
DOI: 10.4085/1062-6050-0084.24 -
Scientific Reports Jun 2024In the field of biotechnology, the utilization of agro-industrial waste for generating high-value products, such as microbial biomass and enzymes, holds significant...
In the field of biotechnology, the utilization of agro-industrial waste for generating high-value products, such as microbial biomass and enzymes, holds significant importance. This study aimed to produce recombinant α-amylase from Anoxybacillus karvacharensis strain K1, utilizing whey as an useful growth medium. The purified hexahistidine-tagged α-amylase exhibited remarkable homogeneity, boasting a specific activity of 1069.2 U mg. The enzyme displayed its peak activity at 55 °C and pH 6.5, retaining approximately 70% of its activity even after 3 h of incubation at 55 °C. Its molecular weight, as determined via SDS-PAGE, was approximately 69 kDa. The α-amylase demonstrated high activity against wheat starch (1648.8 ± 16.8 U mg) while exhibiting comparatively lower activity towards cyclodextrins and amylose (≤ 200.2 ± 16.2 U mg). It exhibited exceptional tolerance to salt, withstanding concentrations of up to 2.5 M. Interestingly, metal ions and detergents such as sodium dodecyl sulfate (SDS), Triton 100, Triton 40, and Tween 80, 5,5'-dithio-bis-[2-nitrobenzoic acid (DNTB), β-mercaptoethanol (ME), and dithiothreitol (DTT) had no significant inhibitory effect on the enzyme's activity, and the presence of CaCl (2 mM) even led to a slight activation of the recombinant enzyme (1.4 times). The Michaelis constant (K) and maximum reaction rate (V), were determined using soluble starch as a substrate, yielding values of 1.2 ± 0.19 mg mL and 1580.3 ± 183.7 μmol mg protein min, respectively. Notably, the most favorable conditions for biomass and recombinant α-amylase production were achieved through the treatment of acid whey with β-glucosidase for 24 h.
Topics: alpha-Amylases; Whey; Anoxybacillus; Detergents; Hydrogen-Ion Concentration; Enzyme Stability; Recombinant Proteins; Starch; Temperature
PubMed: 38830978
DOI: 10.1038/s41598-024-63606-7 -
Microbial Biotechnology Jun 2024Microbial communities from extreme environments are largely understudied, but are essential as producers of metabolites, including enzymes, for industrial processes. As...
Metagenomic exploration of cold-active enzymes for detergent applications: Characterization of a novel, cold-active and alkali-stable GH8 endoglucanase from ikaite columns in SW Greenland.
Microbial communities from extreme environments are largely understudied, but are essential as producers of metabolites, including enzymes, for industrial processes. As cultivation of most microorganisms remains a challenge, culture-independent approaches for enzyme discovery in the form of metagenomics to analyse the genetic potential of a community are rapidly becoming the way forward. This study focused on analysing a metagenome from the cold and alkaline ikaite columns in Greenland, identifying 282 open reading frames (ORFs) that encoded putative carbohydrate-modifying enzymes with potential applications in, for example detergents and other processes where activity at low temperature and high pH is desired. Seventeen selected ORFs, representing eight enzyme families were synthesized and expressed in two host organisms, Escherichia coli and Aliivibrio wodanis. Aliivibrio wodanis demonstrated expression of a more diverse range of enzyme classes compared to E. coli, emphasizing the importance of alternative expression systems for enzymes from extremophilic microorganisms. To demonstrate the validity of the screening strategy, we chose a recombinantly expressed cellulolytic enzyme from the metagenome for further characterization. The enzyme, Cel240, exhibited close to 40% of its relative activity at low temperatures (4°C) and demonstrated endoglucanase characteristics, with a preference for cellulose substrates. Despite low sequence similarity with known enzymes, computational analysis and structural modelling confirmed its cellulase-family affiliation. Cel240 displayed activity at low temperatures and good stability at 25°C, activity at alkaline pH and increased activity in the presence of CaCl, making it a promising candidate for detergent and washing industry applications.
Topics: Cold Temperature; Enzyme Stability; Greenland; Metagenomics; Detergents; Escherichia coli; Cellulase; Metagenome; Hydrogen-Ion Concentration; Recombinant Proteins; Gene Expression; Open Reading Frames
PubMed: 38829370
DOI: 10.1111/1751-7915.14466 -
Journal of Oleo Science 2024Bicellar mixtures containing diacetylene molecules, such as diynoic acids, can be used as parent materials for functional membranes. A bicellar mixture consisting of a...
Bicellar mixtures containing diacetylene molecules, such as diynoic acids, can be used as parent materials for functional membranes. A bicellar mixture consisting of a diynoic acid-10,12-tricosadiynoic acid (TCDA)-, a phospholipid-1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC)-, and a detergent-3-[(3-cholamidopropyl) dimethylammonio]-2-hydroxypropanesulfonate (CHAPSO)-was evaluated for its morphology and packing of TCDA molecules in its bicellar mixture. A TCDA/DMPC vesicle was prepared at different molar ratios, TCDA/DMPC = 2/8, 5/5, and 8/2; a TCDA/DMPC/CHAPSO bicellar mixture was prepared by mixing a CHAPSO solution with a TCDA/DMPC vesicle solution as a detergent at different composition ratios, x = [TCDA/DMPC]/([TCDA/DMPC]+[CHAPSO]), of 1.0, 0.70, 0.50, and 0.30. A DMPC molecule formed a bilayer membrane structure and was used to suppress its precipitation. The packing density of the TCDA/DMPC/CHAPSO bicellar mixtures was increased by mixing a CHAPSO molecule in x = 1.0 to 0.70 or 0.50. A TEM image of a TCDA/DMPC/CHAPSO bicellar mixture showed many discoidal assemblies at x = 0.5 of TCDA/DMPC = 5/5. Polymerization of the TCDA molecules in the bicellar mixture by UV light suggested an ordered arrangement of TCDA. Polymerization at x = 0.70 and 0.50 correlated with improved packing density.
Topics: Dimyristoylphosphatidylcholine; Detergents; Lipid Bilayers; Phase Separation
PubMed: 38825541
DOI: 10.5650/jos.ess24006 -
The Science of the Total Environment Sep 2024The emergence of SARS-CoV-2 has heightened the need to evaluate the detection of enveloped viruses in the environment, particularly in wastewater, within the context of... (Review)
Review
The emergence of SARS-CoV-2 has heightened the need to evaluate the detection of enveloped viruses in the environment, particularly in wastewater, within the context of wastewater-based epidemiology. The studies published over the past 80 years focused primarily on non-enveloped viruses due to their ability to survive longer in environmental matrices such as wastewater or sludge compared to enveloped viruses. However, different enveloped viruses survive in the environment for different lengths of time. Therefore, it is crucial to be prepared to assess the potential infectious risk that may arise from future emerging enveloped viruses. This will require appropriate tools, notably suitable viral concentration methods that do not compromise virus infectivity. This review has a dual purpose: first, to gather all the available literature on the survival of infectious enveloped viruses, specifically at different pH and temperature conditions, and in contact with detergents; second, to select suitable concentration methods for evaluating the infectivity of these viruses in wastewater and sludge. The methodology used in this data collection review followed the systematic approach outlined in the PRISMA (Preferred Reporting Items for Systematic Review and Meta-Analysis) guidelines. Concentration methods cited in the data gathered are more tailored towards detecting the enveloped viruses' genome. There is a lack of suitable methods for detecting infectious enveloped viruses in wastewater and sludge. Ultrafiltration, ultracentrifugation, and polyethylene glycol precipitation methods, under specific/defined conditions, appear to be relevant approaches. Further studies are necessary to validate reliable concentration methods for detecting infectious enveloped viruses. The choice of culture system is also crucial for detection sensitivity. The data also show that the survival of infectious enveloped viruses, though lower than that of non-enveloped ones, may enable environmental transmission. Experimental data on a wide range of enveloped viruses is required due to the variability in virus persistence in the environment.
Topics: Sewage; Wastewater; SARS-CoV-2; Viruses; COVID-19
PubMed: 38825204
DOI: 10.1016/j.scitotenv.2024.173648