ABSTRACT
Haloalkane dehalogenase, LinB, is a member of the α/ß hydrolase family of enzymes. It has a wide range of halogenated substrates, but, has been mostly studied in context of degradation of hexachlorocyclohexane (HCH) isomers, especially ß-HCH (5-12% of total HCH isomers), which is the most recalcitrant and persistent among all the HCH isomers. LinB was identified to directly act on ß-HCH in a one or two step transformation which decreases its toxicity manifold. Thereafter, many studies focused on LinB including its structure determination using X-ray crystallographic studies, structure comparison with other haloalkane dehalogenases, substrate specificity and kinetic studies, protein engineering and site-directed mutagenesis studies in search of better catalytic activity of the enzyme. LinB was mainly identified and characterized in bacteria belonging to sphingomonads. Detailed sequence comparison of LinB from different sphingomonads further revealed the residues critical for its activity and ability to catalyze either one or two step transformation of ß-HCH. Association of LinB with IS6100 elements is also being discussed in detail in sphingomonads. In this review, we summarized vigorous efforts done by different research groups on LinB for developing better bioremediation strategies against HCH contamination. Also, kinetic studies, protein engineering and site directed mutagenesis studies discussed here forms the basis of further exploration of LinB's role as an efficient enzyme in bioremediation projects.
Subject(s)
Hexachlorocyclohexane , Hydrolases , Hydrolases/metabolism , Hydrolases/genetics , Hydrolases/chemistry , Hexachlorocyclohexane/metabolism , Substrate Specificity , Bacterial Proteins/metabolism , Bacterial Proteins/genetics , Bacterial Proteins/chemistry , Kinetics , Biodegradation, Environmental , Crystallography, X-Ray , Mutagenesis, Site-Directed , Sphingomonas/enzymology , Sphingomonas/genetics , Sphingomonas/metabolismABSTRACT
Deinococcus species, noted for their exceptional resistance to DNA-damaging environmental stresses, have piqued scientists' interest for decades. This study dives into the complex mechanisms underpinning radiation resistance in the Deinococcus genus. We have examined the genomes of 82 Deinococcus species and classified radiation-resistance proteins manually into five unique curated categories: DNA repair, oxidative stress defense, Ddr and Ppr proteins, regulatory proteins, and miscellaneous resistance components. This classification reveals important information about the various molecular mechanisms used by these extremophiles which have been less explored so far. We also investigated the presence or lack of these proteins in the context of phylogenetic relationships, core, and pan-genomes, which offered light on the evolutionary dynamics of radiation resistance. This comprehensive study provides a deeper understanding of the genetic underpinnings of radiation resistance in the Deinococcus genus, with potential implications for understanding similar mechanisms in other organisms using an interactomics approach. Finally, this study reveals the complexities of radiation resistance mechanisms, providing a comprehensive understanding of the genetic components that allow Deinococcus species to flourish under harsh environments. The findings add to our understanding of the larger spectrum of stress adaption techniques in bacteria and may have applications in sectors ranging from biotechnology to environmental research.
ABSTRACT
In the era of deep space exploration, extremophile research represents a key area of research w.r.t space survival. This review thus delves into the intriguing realm of 'Space and Astro Microbiology', providing insights into microbial survival, resilience, and behavioral adaptations in space-like environments. This discussion encompasses the modified behavior of extremophilic microorganisms, influencing virulence, stress resistance, and gene expression. It then shifts to recent studies on the International Space Station and simulated microgravity, revealing microbial responses that impact drug susceptibility, antibiotic resistance, and its commercial implications. The review then transitions into Astro microbiology, exploring the possibilities of interplanetary transit, lithopanspermia, and terraforming. Debates on life's origin and recent Martian meteorite discoveries are noted. We also discuss Proactive Inoculation Protocols for selecting adaptable microorganisms as terraforming pioneers. The discussion concludes with a note on microbes' role as bioengineers in bioregenerative life support systems, in recycling organic waste for sustainable space travel; and in promoting optimal plant growth to prepare Martian and lunar basalt. This piece emphasizes the transformative impact of microbes on the future of space exploration.
ABSTRACT
The study focuses on the in silico genomic characterization of Sphingobium indicum B90A, revealing a wealth of genes involved in stress response, carbon monoxide oxidation, ß-carotene biosynthesis, heavy metal resistance, and aromatic compound degradation, suggesting its potential as a bioremediation agent. Furthermore, genomic adaptations among nine Sphingomonad strains were explored, highlighting shared core genes via pangenome analysis, including those related to the shikimate pathway and heavy metal resistance. The majority of genes associated with aromatic compound degradation, heavy metal resistance, and stress response were found within genomic islands across all strains. Sphingobium indicum UT26S exhibited the highest number of genomic islands, while Sphingopyxis alaskensis RB2256 had the maximum fraction of its genome covered by genomic islands. The distribution of lin genes varied among the strains, indicating diverse genetic responses to environmental pressures. Additionally, in silico evidence of horizontal gene transfer (HGT) between plasmids pSRL3 and pISP3 of the Sphingobium and Sphingomonas genera, respectively, has been provided. The manuscript offers novel insights into strain B90A, highlighting its role in horizontal gene transfer and refining evolutionary relationships among Sphingomonad strains. The discovery of stress response genes and the czcABCD operon emphasizes the potential of Sphingomonads in consortia development, supported by genomic island analysis.
Subject(s)
Biodegradation, Environmental , Computer Simulation , Genome, Bacterial , Hexachlorocyclohexane , Phylogeny , Sphingomonadaceae , Sphingomonadaceae/genetics , Sphingomonadaceae/metabolism , Sphingomonadaceae/classification , Hexachlorocyclohexane/metabolism , Genomic Islands , Gene Transfer, HorizontalABSTRACT
Globally, rice is becoming more vulnerable to arsenic (As) pollution, posing a serious threat to public food safety. Previously Debaryomyces hansenii was found to reduce grain As content of rice. To better understand the underlying mechanism, we performed a genome analysis to identify the key genes in D. hansenii responsible for As tolerance and plant growth promotion. Notably, genes related to As resistance (ARR, Ycf1, and Yap) were observed in the genome of D. hansenii. The presence of auxin pathway and glutathione metabolism-related genes may explain the plant growth-promoting potential and As tolerance mechanism of this novel yeast strain. The genome annotation of D. hansenii indicated that it contains a repertoire of genes encoding antioxidants, well corroborated with the in vitro studies of GST, GR, and glutathione content. In addition, the effect of D. hansenii on gene expression profiling of rice plants under As stress was also examined. The Kyoto Encyclopedia of Genes and Genomes (KEGG) database revealed 307 genes, annotated in D. hansenii-treated rice, related to metabolic pathways (184), photosynthesis (12), glutathione (10), tryptophan (4), and biosynthesis of secondary metabolite (117). Higher expression of regulatory elements like AUX/IAA and WRKY transcription factors (TFs), and defense-responsive genes dismutases, catalases, peroxiredoxin, and glutaredoxins during D. hansenii+As exposure was also observed. Combined analysis revealed that D. hansenii genes are contributing to stress mitigation in rice by supporting plant growth and As-tolerance. The study lays the foundation to develop yeast as a beneficial biofertilizer for As-prone areas.
Subject(s)
Arsenic , Debaryomyces , Oryza , Debaryomyces/genetics , Debaryomyces/metabolism , Oryza/metabolism , Arsenic/toxicity , Arsenic/metabolism , Saccharomyces cerevisiae/genetics , Gene Expression Profiling , Glutathione/metabolismABSTRACT
The genus Idiomarina consists of halophilic and/or haloalkaliphilic organisms. We compared the complete genomes of seven strains of the genus Idiomarina to investigate its adaptation to saline environment. A total of 1,313 core genes related to salinity tolerance, stress response, antibiotic resistance genes, virulence factors, and drug targets were found. Comparative genomics revealed various genes involved in halo adaptations of these organisms, including transporters and influx or efflux systems for elements such as Fe, Cu, Zn, Pb, and Cd. In agreement with their isolation sources (such as hydrothermal vents and marine sediments) and environments abundant in heavy metals, various resistance proteins and transporters associated with metal tolerance were also identified. These included copper resistance proteins, zinc uptake transcriptional repressor Zur, MerC domain-containing protein, Cd(II)/Pb(II)-responsive transcriptional regulator, Co/Zn/Cd efflux system protein, and mercuric transporter. Interestingly, we observed that the carbohydrate metabolism pathways were incomplete in all the strains and transporters used for absorption of small sugars were also not found in them. Also, the presence of higher proportion of genes involved in protein metabolism than carbohydrate metabolism indicates that proteinaceous substrates act as the major food substrates for these bacterial strains than carbohydrates. Genomic islands were detected in some species, highlighting the role of horizontal gene transfer for acquisition in novel genes. Genomic rearrangements in terms of partially palindromic regions were detected in all strains. To our knowledge, this is the first comprehensive comparative genomics study among the genus Idiomarina revealing unique genomic features within bacterial species inhabiting different ecological niches. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-023-03887-3.
ABSTRACT
The taxonomic status of 43 Psychrobacter species was examined based upon the genome sequences of their type strains. Three groups of type strains were found to be conspecific, Psychrobacter salsus Shivaji et al. (Syst Appl Microbiol 27:628-635, 2004. 10.1078/0723202042369956) and Psychrobacter submarinus Romanenko et al. (Int J Syst Evol Microbiol 52:1291-1297, 2002. 10.1099/00207713-52-4-1291); Psychrobacter oceani Matsuyama et al. (Int J Syst Evol Microbiol 65:1450-1455, 2015. 10.1099/ijs.0.000118) and Psychrobacter pacificensis Maruyama et al. (Int J Syst Evol Microbiol 50:835-846, 2000. 10.1099/00207713-50-2-835); and Psychrobacter proteolyticus Denner et al. (Syst Appl Microbiol 24:44-53, 2001. 10.1078/0723-2020-00006), Psychrobacter marincola Romanenko et al. (Int J Syst Evol Microbiol 52:1291-1297, 2002. 10.1099/00207713-52-4-1291) and Psychrobacter adeliensis Shivaji et al. (Syst Appl Microbiol 27:628-635, 2004. 10.1078/0723202042369956). For all three groups, the average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values are > 97.69% and > 80.2%, respectively. This conclusion is supported by similarities in morphology, growth properties, and fatty acid compositions. Based on this evidence, we propose the reclassification of Psychrobacter salsus Shivaji et al. (Syst Appl Microbiol 27:628-635, 2004. 10.1078/0723202042369956) as a later heterotypic synonym of Psychrobacter submarinus Romanenko et al. (Int J Syst Evol Microbiol 52:1291-1297, 2002. 10.1099/00207713-52-4-1291); Psychrobacter oceani Matsuyama et al. (Int J Syst Evol Microbiol 65:1450-1455, 2015. 10.1099/ijs.0.000118) as a later heterotypic synonym of Psychrobacter pacificensis Maruyama et al. (Int J Syst Evol Microbiol 50:835-846, 2000. 10.1099/00207713-50-2-835), and Psychrobacter marincola Romanenko et al. (Int J Syst Evol Microbiol 52:1291-1297, 2002. 10.1099/00207713-52-4-1291) and Psychrobacter adeliensis Shivaji et al. (Syst Appl Microbiol 27:628-635, 2004. 10.1078/0723202042369956) as later heterotypic synonyms of Psychrobacter proteolyticus Denner et al. (Syst Appl Microbiol 24:44-53, 2001. 10.1078/0723-2020-00006).
Subject(s)
Psychrobacter , Psychrobacter/genetics , Phylogeny , DNA, Bacterial/geneticsABSTRACT
Monkeypox is a viral zoonosis with symptoms that are reminiscent of those experienced in previous smallpox cases. The GSAID database (Global Initiative on Sharing Avian Influenza Data) was used to assess 630 genomes of MPXV. The phylogenetic study revealed six primary clades, as well as a smaller percentage in radiating clades. Individual clades that make up various nationalities may have formed as a result of a particular SNP hotspot type that mutated in a specific population. The most significant mutation based on a mutational hotspot analysis was found at G3729A and G5143A. The gene ORF138, which encodes the Ankyrin repeat (ANK) protein, was found to have the most mutations. This protein mediates molecular recognition via protein-protein interactions. It was shown that 243 host proteins interacted with 10 monkeypox proteins identified as the hub proteins E3, SPI2, C5, K7, E8, G6, N2, B14, CRMB, and A41 through 262 direct connections. The interaction with chemokine system-related proteins provides further evidence that the monkeypox virus suppresses human proteins to facilitate its survival against innate immunity. Several FDA-approved molecules were evaluated as possible inhibitors of F13, a significant envelope protein on the membrane of extracellular versions of the virus. A total of 2500 putative ligands were individually docked with the F13 protein. The interaction between the F13 protein and these molecules may help prevent the monkeypox virus from spreading. After being confirmed by experiments, these putative inhibitors could have an impact on the activity of these proteins and be used in monkeypox treatments.
Subject(s)
Monkeypox virus , Mpox (monkeypox) , Animals , Humans , Phylogeny , Genomics , MutationABSTRACT
Bacterial transmission is considered one of the potential risks for communicable diseases, requiring promising antibiotics. Traditional drugs possess a limited spectrum of effectiveness, and their frequent administration reduces effectiveness and develops resistivity. In such a situation, we are left with the option of developing novel antibiotics with higher efficiency. In this regard, nanoparticles (NPs) may play a pivotal role in managing such medical situations due to their distinct physiochemical characteristics and impressive biocompatibility. Metallic NPs are found to possess extraordinary antibacterial effects that are useful in vitro as well as in vivo as self-modified therapeutic agents. Due to their wide range of antibacterial efficacy, they have potential therapeutic applications via diverse antibacterial routes. NPs not only restrict the development of bacterial resistance, but they also broaden the scope of antibacterial action without binding the bacterial cell directly to a particular receptor with promising effectiveness against both Gram-positive and Gram-negative microbes. This review aimed at exploring the most relevant types of metal NPs employed as antimicrobial agents, particularly those based on Mn, Fe, Co, Cu, and Zn metals, and their antimicrobial mechanisms. Further, the challenges and future prospects of NPs in biological applications are also discussed.
ABSTRACT
The study aimed to explicate the role of microbial co-inoculants for the mitigation of arsenic (As) toxicity in rice. Arsenate (AsV) reducer yeast Debaryomyces hansenii NBRI-Sh2.11 (Sh2.11) with bacterial strains of different biotransformation potential was attempted to develop microbial co-inoculants. An experiment to test their efficacy (yeast and bacterial strains) on plant growth and As uptake was conducted under a stressed condition of 20 mg kg-1 of arsenite (AsIII). A combination of Sh2.11 with an As(III)-oxidizer, Citrobacter sp. NBRI-B5.12 (B5.12), resulted in â¼90% decrease in grain As content as compared to Sh2.11 alone (â¼40%). Reduced As accumulation in rice roots under co-treated condition was validated with SEM-EDS analysis. Enhanced As expulsion in the selected combination under in vitro conditions was found to be correlated with higher As content in the soil during their interaction with plants. Selected co-inoculant mediated enhanced nutrient uptake in association with better production of indole acetic acid (IAA) and gibberellic acid (GA) in shoot, support microbial co-inoculant mediated better biomass under stressful condition. Boosted defense response in association with enhanced glutathione-S-transferase (GST) and glutathione reductase (GR), activities under in vitro and in vivo conditions were observed. These results indicated that the As(III) oxidizer-B5.12 accelerated the As detoxification property of the As(V) reducer-Sh2.11. Henceforth, the results confer that the coupled reduction-oxidation process of the co-inoculant reduces the accumulation of As in rice grain. These co-inoculants can be further developed for field trials to achieve higher biomass with alleviated As toxicity in rice.
Subject(s)
Agricultural Inoculants , Arsenic , Arsenites , Oryza , Soil Pollutants , Arsenates/toxicity , Arsenates/metabolism , Arsenic/toxicity , Arsenic/metabolism , Saccharomyces cerevisiae , Oryza/metabolism , Arsenites/toxicity , Arsenites/metabolism , Bacteria/metabolism , Oxidation-Reduction , Agricultural Inoculants/metabolism , Plant Roots/metabolism , Soil Pollutants/analysisABSTRACT
Arsenic (As) has become natural health hazard for millions of people across the world due to its distribution in the food chain. Naturally, it is present in different oxidative states of inorganic [As(V) and As(III)] and organic (DMA, MMA and TMA) forms. Among different mitigation approaches, microbe mediated mitigation of As toxicity is an effective and eco-friendly approach. The present study involves the characterization of bacterial strains containing arsenite methyltransferase (Pseudomonas oleovorans, B4.10); arsenate reductase (Sphingobacterium puteale, B4.22) and arsenite oxidase (Citrobacter sp., B5.12) activity with plant growth promoting (PGP) traits. Efficient reduction of grain As content by 61 % was observed due to inoculation of methyltransferase containing B4.10 as compared to B4.22 (47 %) and B5.12 (49 %). Reduced bioaccumulation of As in root (0.339) and shoot (0.166) in presence of B4.10 was found to be inversely related with translocation factor for Mn (3.28), Fe (0.073), and Se (1.82). Bioaccumulation of these micro elements was found to be associated with the modulated expression of different mineral transporters (OsIRT2, OsFRO2, OsTOM1, OsSultr4;1, and OsZIP2) in rice shoot. Improved dehydrogenase (407 %), and ß-glucosidase (97 %) activity in presence of P. oleovorans (B4.10) as compared to arsenate reductase (198 and 50 %), and arsenite oxidase (134 and 69 %) containing bacteria was also observed. Our finding confers the potential of methyltransferase positive P. oleovorans (B4.10) for As stress amelioration. Reduced grain As uptake was found to be mediated by improved plant growth and nutrient uptake associated with enhanced soil microbial activity.
Subject(s)
Arsenic , Arsenicals , Arsenites , Oryza , Pseudomonas oleovorans , Humans , Arsenic/toxicity , Arsenic/metabolism , Arsenate Reductases/metabolism , Pseudomonas oleovorans/metabolism , Plant Roots/metabolism , Edible Grain/metabolism , Arsenicals/metabolism , Methyltransferases , Arsenites/metabolismABSTRACT
A rigorous exploration of microbial diversity has revealed its presence on Earth, deep oceans, and vast space. The presence of microbial life in diverse environmental conditions, ranging from moderate to extreme temperature, pH, salinity, oxygen, radiations, and altitudes, has provided the necessary impetus to search for them by extending the limits of their habitats. Microbiology started as a distinct science in the mid-nineteenth century and has provided inputs for the betterment of mankind during the last 150 years. As beneficial microbes are assets and pathogens are detrimental, studying both have its own merits. Scientists are nowadays working on illustrating the microbial dynamics in Earth's subsurface, deep sea, and polar regions. In addition to studying the role of microbes in the environment, the microbe-host interactions in humans, animals and plants are also unearthing newer insights that can help us to improve the health of the host by modulating the microbiota. Microbes have the potential to remediate persistent organic pollutants. Antimicrobial resistance which is a serious concern can also be tackled only after monitoring the spread of resistant microbes using disciplines of genomics and metagenomics The cognizance of microbiology has reached the top of the world. Space Missions are now looking for signs of life on the planets (specifically Mars), the Moon and beyond them. Among the most potent pieces of evidence to support the existence of life is to look for microbial, plant, and animal fossils. There is also an urgent need to deliberate and communicate these findings to layman and policymakers that would help them to take an adequate decision for better health and the environment around us. Here, we present a glimpse of recent advancements by scientists from around the world, exploring and exploiting microbial diversity.
ABSTRACT
Polyphosphate (polyP) accumulation is an important trait of microorganisms. Implication of polyP accumulating bacteria (PAB) in enhanced biological phosphate removal, heavy metal sequestration, and dissolution of dental enamel is well studied. Phosphorous (P) accumulated within microbial biomass also regulates labile P in soil; however, abundance and diversity of the PAB in soil is still unexplored. Present study investigated the genetic and functional diversity of PAB in rhizosphere soil. Here, we report the abundance of Pseudomonas spp. as high PAB in soil, suggesting their contribution to global P cycling. Additional subset analysis of functional genes i.e., polyphosphate kinase (ppk) and exopolyphosphatase (ppx) in all PAB, indicates their significance in bacterial growth and metabolism. Distribution of functional genes in phylogenetic tree represent a more biologically realistic discrimination for the two genes. Distribution of ppx gene disclosed its phylogenetic conservation at species level, however, clustering of ppk gene of similar species in different clades illustrated its environmental condition mediated modifications. Selected PAB showed tolerance to abiotic stress and strong correlation with plant growth promotary (PGP) traits viz. phosphate solubilization, auxin and siderophore production. Interaction of PAB with A. thaliana enhanced the growth and phosphate status of the plant under salinity stress, suggestive of their importance in P cycling and stress alleviation. IMPORTANCE Study discovered the abundance of Pseudomonas genera as a high phosphate accumulator in soil. The presence of functional genes (polyphosphate kinase [ppk] and exopolyphosphatase [ppx]) in all PAB depicts their importance in polyphosphate metabolism in bacteria. Genetic and functional diversity reveals conservation of the ppx gene at species level. Furthermore, we found a positive correlation between PAB and plant growth promotary traits, stress tolerance, and salinity stress alleviation in A. thaliana.
Subject(s)
Arabidopsis/growth & development , Polyphosphates/metabolism , Pseudomonas/genetics , Pseudomonas/metabolism , Soil Microbiology , Acid Anhydride Hydrolases/genetics , Acid Anhydride Hydrolases/metabolism , Arabidopsis/microbiology , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Genetic Variation , Indoleacetic Acids/metabolism , Phosphorus/metabolism , Phosphotransferases (Phosphate Group Acceptor)/genetics , Phosphotransferases (Phosphate Group Acceptor)/metabolism , Phylogeny , Pseudomonas/classification , Pseudomonas/enzymology , Rhizosphere , Siderophores/biosynthesis , Soil/chemistryABSTRACT
Silicon (Si) has gained considerable attention for its utility in improved plant health under biotic and abiotic stresses through alteration of physiological and metabolic processes. Its interaction with arsenic (As) has been the compelling area of research amidst heavy metal toxicity. However, microbe mediated Si solubilization and their role for reduced As uptake is still an unexplored domain. Foremost role of Bacillus amyloliquefaciens (NBRISN13) in impediment of arsenite (AsIII) translocation signifies our work. Reduced grain As content (52-72%) during SN13 inoculation under feldspar supplementation (Si+SN+As) highlight the novel outcome of our study. Upregulation of Lsi1, Lsi2 and Lsi3genes in Si+SN+As treated rice plants associated with restricted As translocation, frames new propositions for future research on microbemediated reduced As uptake through increased Si transport. In addition to low As accumulation, alleviation of oxidative stress markers by modulation of defense enzyme activities and differential accumulation of plant hormones was found to be associated with improved growth and yield. Thus, our findings confer the potential role of microbe mediated Si solubilization in mitigation of As stress to restore plant growth and yield.
Subject(s)
Arsenic , Bacillus amyloliquefaciens , Oryza , Arsenic/toxicity , Plant Roots , Plants , Silicon/toxicityABSTRACT
What microbiology beholds after a decade and a half in the future requires a vision based on the facts and ongoing trends in research and technological advancements. While the latter, assisted by microbial dark matter, presents a greater potential of creating an upsurge in in-situ and ex-situ rapid microbial detection techniques, this anticipated change will also set forth a revolution in microbial cultivation and diversity analyses. The availability of a microbial genetic toolbox at the expanse will help complement the current understanding of the microbiome and assist in real-time monitoring of the dynamics for detecting the health status of the host with utmost precision. Alongside, in light of the emerging infectious diseases, antimicrobial resistance (AMR) and social demands for safer and better health care alternatives, microbiology laboratories are prospected to drift in terms of the volume and nature of research and outcomes. With today's microbiological lens, one can predict with certainty that in the years to come, microbes will play a significant role in therapeutic treatment and the designing of novel diagnostic techniques. Another area where the scope of microbial application seems to be promising is the use of novel probiotics as a method to offer health benefits whilst promoting metabolic outputs specific for microbiome replenishment. Nonetheless, the evolution of extraterrestrial microbes or the adaptation of earth microbes as extraterrestrial residents are also yet another prominent microbial event one may witness in the upcoming years. But like the two sides of the coin, there is also an urgent need to dampen the bloom of urbanization, overpopulation and global trade and adopting sustainable approaches to control the recurrence of epidemics and pandemics.
Subject(s)
MicrobiotaABSTRACT
In Saccharomyces cerevisiae, the selective autophagic degradation of mitochondria, termed mitophagy, is critically regulated by the adapter protein Atg32. Despite our knowledge about the molecular mechanisms by which Atg32 controls mitophagy, its physiological roles in yeast survival and fitness remains less clear. Here, we demonstrate a requirement for Atg32 in promoting spermidine production during respiratory growth and heat-induced mitochondrial stress. During respiratory growth, mitophagy-deficient yeast exhibit profound heat-stress induced defects in growth and viability due to impaired biosynthesis of spermidine and its biosynthetic precursor S-adenosyl methionine. Moreover, spermidine production is crucial for the induction of cytoprotective nitric oxide (NO) during heat stress. Hence, the re-addition of spermidine to Atg32 mutant yeast is sufficient to both enhance NO production and restore respiratory growth during heat stress. Our findings uncover a previously unrecognized physiological role for yeast mitophagy in spermidine metabolism and illuminate new interconnections between mitophagy, polyamine biosynthesis and NO signaling.
Subject(s)
Mitophagy , Saccharomyces cerevisiae Proteins , Autophagy/genetics , Autophagy-Related Proteins/genetics , Heat-Shock Response/genetics , Nitric Oxide , Receptors, Cytoplasmic and Nuclear , Saccharomyces cerevisiae Proteins/genetics , SpermidineABSTRACT
LinB is an important haloalkane dehalogenase involved in the degradation pathway of different isomers of hexachlorocyclohexane (HCH), mainly in catalyzing degradation of the notorious ß-HCH. The HCH isomers are known to have neurotoxic, carcinogenic and estrogenic effects. Enzymatic bioremediation for decontamination of ß- as well as other HCH isomers can prove to be a potential remediation strategy. For any bioremediation technology that is to be developed, apart from having high turnover number, the candidate enzyme must also be available in sufficient amounts. In this direction, the LinB variants reported in database were tested in laboratory studies. The variant LinBSSO4-3 however could not be obtained in soluble fraction by using standard procedures. The protein LinBSSO4-3 was cloned in pDEST17 vector and codon optimized for better expression in Escherichia coli BL21AI using a strong T7 promoter. However, the over-expression of this protein in ectopic host E. coli, led to aggregation of the protein in form of inclusion bodies, which are insoluble aggregates of misfolded or partially folded proteins. SEM analysis of the inclusion bodies showed them as aggregated spherical particles. The inclusion bodies were isolated using high speed sonication and homogenization. This was followed by solubilization in the strong denaturing agent urea. Refolding into its native state was done by using pulsatile refolding. This was done by slowly decreasing the denaturant concentration in the presence of sucrose. The turnover number of the refolded protein was then determined for different isomers of HCH. The protein was found to have a turnover number of â¼43 molecules min-1 on ß-HCH and â¼13 molecules min-1 on δ-HCH. Additionally, a mutation I253 M in the active site of the enzyme was found to drastically decrease the enzyme activity on ß-HCH. Taking into consideration the wide range of substrates of haloalkane dehalogenases, such a protocol for inclusion body refolding will contribute to the field of bioremediation technology development for organochlorines, specifically HCH. Such a protocol for refolding of haloalkane dehalogenases from inclusion bodies has not been developed or reported before.
Subject(s)
Escherichia coli , Hexachlorocyclohexane , Escherichia coli/genetics , Hydrolases/genetics , Inclusion BodiesABSTRACT
COVID-19 patients presenting for emergency laparotomy require evaluation of surgical illness and viral disease. As these patients are likely to have a wide spectrum of deranged physiology and organ dysfunction, optimization should start preoperatively and continue through intraoperative and postoperative recovery periods along with appropriate antimicrobial cover. The goal should be not to delay damage control surgery in favor of evaluation and optimization. When a COVID-19 positive or suspected patient is to be operated for laparotomy, the situation often demands general anesthesia with invasive monitoring and analgesia complemented by regional anesthesia to minimize postoperative opioid requirements to facilitate early recovery. This particular article addresses the issues related to emergency laparotomy management in relation to COVID-19 patient. Healthcare workers should diligently use effective PPE and practice disinfection to prevent spread. Video-communication is an effective means of evaluation. Information expected from investigations should be weighed against risk of exposure to healthcare workers/laypersons. Simulation and memory aids should be used to familiarize team members with roles and techniques of management while in PPE. Step-wise detailed planning for patient transfer, anesthesia induction, maintenance and emergence, aid in enhancing HCW safety without compromising patient care.