RSC Advances

Copper is an essential metalloelement that plays key fundamental roles in both health and pathology, and is increasingly been implicated in molecular pathogenesis of many cancer types. It has shown promise as a replacement to cisplatin in coordination complexes presently in mainstream chemotherapeutic practices. In this study, two newly synthesized water-soluble ternary copper (II) mixed ligand complexes; complex 1 - (Cu(4-mphen)(tyr)(H2O)]NO3{middle dot}2H2O)(C.1) and complex 2 - (Cu(5-mphen)(tyr)(H2O)]NO3{middle dot}2H2O (C.2) where (4-m= 4-methyl; 5-m = 5-methyl; phen-1, 10 = phenanthroline; tyr = tyrosine)), were investigated on adenocarcinomic human alveolar basal epithelial cell, A549 and non-cancerous human bronchial epithelial cell, BEAS-2B for their antiproliferative effects using the XTT assay (cytotoxicity), Comet assay (genotoxicity) and DCFH-DA assay (intracellular ROS) tests. C.1 was significantly more cytotoxic in A549 than C.2. Data from the Comet and ROS assay tests support each other. C.2 caused more copper-induced DNA damage, possibly through significant induction of ROS-mediated oxidative damage in the cancer cell, but a minimal insignificant ROS rise in normal cells. These results can only be preliminary and further studies are required to better understand the cellular effects and functional interactions of these agents, for an efficient therapeutic design and application.

Green synthesis of nanoparticles (NPs) involves the use of diverse extracts of biological origin as substrates to synthesize nanoparticles and can overcome the hazards associated with chemical methods. Coconut inflorescence sap, which is unfermented phloem sap obtained by tapping of coconut inflorescence, is a rich source of sugars and secondary metabolites. In this study, coconut inflorescence sap was used to synthesize silver nanoparticles (AgNPs). We have initially undertaken metabolomic profiling of coconut inflorescence sap from West Coast Tall cultivar to delineate its individual components. Secondary metabolites constituted the major portion of the inflorescence sap along with sugars, lipids and, peptides. The concentration of silver nitrate, inflorescence sap and incubation temperature for synthesis of AgNPs were optimized. Incubating the reaction mixture at 40{degrees}C was found to enhance AgNP synthesis. The AgNPs synthesized were characterized using UV-Visible spectrophotometry, X-Ray Diffraction (XRD), Fourier Transform Infrared spectroscopy (FTIR), Field Emission Scanning Electron Microscopy (FESEM) and Transmission Electron Microscopy (TEM). Antimicrobial property of AgNP was tested in tissue culture of arecanut (Areca catechu L.) where bacterial contamination (Bacillus pumilus) was a frequent occurrence. Significant reduction in the contamination was observed when plantlets were treated with aqueous solutions of 0.01, 0.02 and 0.03% of AgNPs for one hour. Notably, treatment with AgNPs did not affect growth and development of the arecanut plantlets. Cytotoxicity of AgNPs was quantified in HeLa cells. Viability (%) of HeLa cells declined significantly at 10 ppm concentration of AgNP and complete mortality was observed at 60 ppm. Antimicrobial properties of AgNPs synthesized from inflorescence sap were also evaluated and confirmed in human pathogenic bacteria viz., Salmonella sp., Vibrio parahaemolyticus, and Escherichia coli. The study concludes that unfermented inflorescence sap, with above neutral pH, serves as an excellent reducing agent to synthesize AgNPs from Ag+.\n\nO_FIG O_LINKSMALLFIG WIDTH=177 HEIGHT=200 SRC=\"FIGDIR/small/775940v1_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (64K):\norg.highwire.dtl.DTLVardef@3c3f1forg.highwire.dtl.DTLVardef@1540f6borg.highwire.dtl.DTLVardef@1c069aborg.highwire.dtl.DTLVardef@159772c_HPS_FORMAT_FIGEXP M_FIG Graphical abstract\n\nC_FIG

Evading apoptosis is a hallmark of cancer cells, therefore therapeutic strategies have been developed to induce cell death. BCL2 family protein governs the intrinsic pathway of cell death. Targeting the BH4 domain to modulate the anti-apoptosis activities of BCL2 protein has been established however, BDA366 is the only BH4 binding molecule to be reported. Virtually screening ~ 1,000,000 compounds 11 putative BH4 binding small molecules with binding affinity ~ -84kcal/mol to - 64kcal/mol resulted. Using QM-polarized docking, Induced-fit docking, and QM-MM optimization, a putative binding mode for the top 3 compounds is proposed: compound 139068 interactions with GLU13, MET16, LYS17, ASP31, and GLU42; compound 138967 interactions with ASP10, ARG12, GLU13, HIS20, MET16, and GLU42; compound 38831 interactions with ASP10, ARG12, GLU13, LYS17, and GLU42. MD simulations (NMA) data showed the binding of the three compounds to be stable with low eigenvalues. Electronic properties derived via DFT calculations suggest chemical reaction of the compounds be via electrophilic reactions.

Green synthesis of silver nanoparticles has gained much interest over few decades in the field of biomedical research due to their ease of synthesis, cost effectiveness, enhanced bioactivity and biocompatibility compared to the chemical synthesis. Recent studies on silver nanoparticles have shown their potential in various fields like antimicrobial, anticancer, larvicidal, catalytic, and wound healing properties. In the present study, entomopathogenic fungi Metarhizium anisopliae was used to synthesize silver nanoparticles. These silver nanoparticles were synthesized and characterized using UV-Vis spectroscopy, FESEM, FETEM and FTIR. Compared to the chemically synthesized silver nanoparticles, the mycosynthesized silver nanoparticles (MaAgNP) showed high yield. The size of mycosynthesized silver nanoparticles was found to be 5-20 nm and was spherical in shape. FTIR results confirmed the possible functional groups that are responsible for the reduction of silver ions. The mycosynthesized silver nanoparticles showed cytotoxicity on human breast cancer cell line (MCF-7) and the calculated IC50 value for MaAgNP was found to be 16.50 g ml-1 whereas the chemically synthesized silver nanoparticles showed cytotoxicity but with increasing concentration, no further significant reduction in cell viability was observed. The possible reason behind improved cytotoxicity of MaAgNP can be the presence of extracellular secondary metabolites present in the fungal filtrate used to synthesize the nanoparticles. The MaAgNP was also observed to induce cell death through reactive oxygen species (ROS) generation.

Wedelia chinensis is a medicinal herb of Asteraceae family. Preliminary phytochemical screening of internodal extract was done by using its aqueous, methanolic and ethanolic extracts. Preliminary phytochemical screening showed the presence of several pharmaceutically important compounds such as alkaloids, tannins, anthraquinons, saponins, flavonoids, terpenoids, phenols, etc. Aqueous internodal extract was used for the biosynthesis of gold nanoparticles. The characterization of gold nanoparticles were done by using a range of analytical techniques, such as energy-dispersive X-ray spectroscopy (EDS, EDX), X-ray diffraction (XRD), transmission electron microscopy (TEM), dynamic light scattering (DLS), Fourier transformed infrared spectroscopy (FTIR), scanning electron microscopy (SEM). The stability and other characteristics were ascertained using a UV-Vis spectrophotometer. The plasmon resonance occurred at the wavelength of 530 nm, thus showing the formation of gold nanoparticles. X-ray diffraction demonstrated the existence of Au-rich (fcc) phases in gold nanoparticles. FTIR analysis clearly indicated the participation of active constituents of internodal extract in the conversion of gold ions into AuNPs. DLS showed the size distribution (99.71 nm) of the suspended particles. There was variations in the sizes and shapes of the nanoparticles. The zeta potential of synthesized AuNPs was observed as -5.12 mV, thus showing the stability of the synthesized nanoparticles. In TEM study, uniform distribution of nanoparticles was seen with some agglomerates and average size of nanoparticles was observed in between 30-40 nm. The antioxidant potential of internodal extract was slightly higher than the synthesized gold nanoparticles. The green-synthesized AuNPs along with antibiotics demonstrated strong synergistic effect against several bacterial strains such as Escherichia coli and Bacillus subtilis. Gold nanoparticles showed good hemolytic activity against rat erythrocytes (90%). Gold nanoparticles also showed catalytic activity that reduces methylene blue at low concentration (1 mg). Highlights[tpltrtarr] Internodal extracts showed the presence of several active constituents in preliminary phytochemical screening. [tpltrtarr]Internodal extract was used for the synthesis of gold nanoparticles in one step process. [tpltrtarr]Synergistic action of AuNPs and antibiotics were reported against many pathogenic bacterial strains. [tpltrtarr]AuNPs showed its potential as antioxidant. [tpltrtarr]Hemolytic effect of nanoparticles was reported on rat blood sample. [tpltrtarr]Nanoparticles showed dye reducing activity thus enhancing its application in the purification of water bodies. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=159 SRC="FIGDIR/small/660090v1_ufig1.gif" ALT="Figure 1"> View larger version (38K): org.highwire.dtl.DTLVardef@12a27e3org.highwire.dtl.DTLVardef@a0d1b6org.highwire.dtl.DTLVardef@28178aorg.highwire.dtl.DTLVardef@216935_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOFig.C_FLOATNO Graphical abstract of AuNPs C_FIG

Methicillin-resistant Staphylococcus aureus (MRSA) is a critical global problem in case of infection due to its multidrug resistance and adaptive nature. Polyisoprenyl-teichoic acid--peptidoglycan teichoic acid transferase (PTA), an essential enzyme for biosynthesis of teichoic acid is present in almost all of the gram-positive species, especially S. aureus. A biocompatible gel-patch formulated using acetone extract of microalgae Haematococcus pluvialis was developed and evaluated for its favourable physicochemical properties such as moisture retention and thickness. Antioxidant activity of the patch and extract was assessed via DPPH assay and results showed a dose-dependent radical scavenging potential and an IC of 24.23 {micro}g/mL, while antibacterial assay against S. aureus showed decent inhibition zones for the extract (5 - 7 mm) and the gel-patch (1 - 3.5 mm). In addition to invitro assays, in-silico analysis was carried out for predicting the effective usage of H. pluvialis against S. aureus strains. Conserved domain analysis identified the presence of a Cps2A domain of the LytR-CpsA-Psr (LCP) family in PTA. Physicochemical profiling revealed a hydrophilic, stable protein nature having high aliphatic index (85.07), low GRAVY score (-0.679), and a structured catalytic core with flanked disordered terminal regions. GC-MS analysis showed the presence of 15 compounds in the algal extract and ADMET analysis identified compounds with suitable drug-likeness, skin permeability and low systemic toxicity. The diversified residue-specific interactions for control drugs and the algal ligands in the conserved Cps2A domain favour a dual binding and synergistic therapeutic approach to target MRSA using the H. pluvialis-based gel-patch.

Nanocarbons, such as fullerenes, carbon nanotubes, and graphene have attracted a great deal of attention as next-generation materials because of their unprecedented structures and unique physicochemical properties; however, almost all nanocarbons reported previously were used as mixtures. Thus, there are still many unsolved issues about their biological functions at the molecular level. Our synthetic campaign in the last decade has synthesized structurally uniform and atomically precise nanocarbons, leading to the preparation of a library consisting of eighty structurally diverse nanocarbon molecules. This resource motivated us to explore the as yet uncovered biological functions of these nanocarbon molecules in organisms. Recently, nanotubes were used to deliver genes to plants; however, the effects of the molecules on plants are not well known. To monitor the effects of nanocarbon molecules on plants, we analyzed the transcriptome of Arabidopsis thaliana seedlings treated with [9]cycloparaphenylene (CPP), decaborylated warped nanographene (WNG), and dimethoxyhexabenzotetracene (HBT). Clustering analysis indicated few effects of nanocarbon molecules on the transcriptome, perhaps suggesting a low toxicity of nanocarbon molecules on plants. We found that AT1G05880 (ARIADNE 12) gene, categorized into response to hypoxia genes, was up-regulated by nanocarbon molecules, suggesting that this gene is usable as maker for treatment of nanocarbon molecules.

Indirubin can be applied as an anti-cancer drug for inhibition of brain tumors. However, its performance is reduced due to hydrophobicity. In this study, we synthesized cationic human serum albumin (CHSA) nanoparticle by a new hybrid approach for improvement the surface chemistry of albumin and investigate the amount of indirubin loaded CHSA nanoparticle. In this study, the generated mechanical force from a high-pressure homogenizer (HPH) was used to make nanoparticles with a certain size with narrow polydispersity. The results indicated that the size of indirubin loaded CHSA nanoparticles were 130 nm and their zeta potential were +9. Besides, the encapsulation efficiency and drug loading capacity were found to be 85% and 5.8 %, respectively. To the best to our knowledge, this is the first time that indirubin has been used in albumin nanoparticles. In this study, indirubin loaded CHSA nanoparticles was shown can be a potential candidate for drug delivery in the treatment of glioblastoma. Moreover, the cationized form allows the chemical agent to be transmitted to the brain.

Metallo-{beta}-lactamases (MBLs) producing bacteria especially the ones with New Delhi metallo-beta-lactamase-1 (NDM-1) and its variants can potentially hydrolyse all the major {beta}-lactam antibiotics, ultimately escalating anti-microbial resistance world-wide. There is a dearth of approved inhibitors to combat NDM and other MBLs producing bacteria. Hence we focussed to find novel inhibitor(s) in-silico which can potentially suppress the activity of NDM/ MBLs. 2400 compounds were virtually screened to identify a promising carboxylic acid-containing compound (CID-53986787) analogous to NDM antagonist Captopril. Our lead compound can bind adjacent to the active site zinc ions (Zn1 and Zn2) in all highly resistant NDM variants. CID-53986787 possesses ~5-8% higher binding affinity than Captopril, exhibiting molecular interactions with crucial residues that can destabilize the hydrolytic activity of NDM. CID-53986787 was virtually evaluated to ascertain its safe pharmacological/ toxicity profile. Molecular dynamics simulation studies elucidated its stable interaction with the target protein (NDM-1).

In this study, the crystalline structure of a sample of trans-cinnamic acid, isolated from an aqueous-alcohol extract of Scutellaria baicalensis in vitro root culture was analyzed using diffraction analysis. The diffraction analysis results confirmed that the investigated trans-cinnamic acid sample corresponds to the declared compound: trans-cinnamic acid. Additionally, the structure of this sample was examined using NMR spectroscopy. The obtained 1H, 13C{1H}, 1H1H-COSY, 1H13C-HSQC, 1H13C-HMBC NMR spectra of the studied sample correspond to the structure of the target compound, trans-cinnamic acid. No impurity signals were detected in the analyzed trans-cinnamic acid sample.

Using mycorrhizal fungi with their helper bacteria (MHB) to alleviate phosphate deficiency improve plant growth and rejuvenate the soil is the newer safer and most promising environment-friendly approach to limiting synthetic agrochemicals. Here 65 MHBs were isolated from the mycorrhizosphere of the Banana plant and 12 were screened based on their biofilm formation protease and N-acyl homoserine lactones production. A newly reported MHB Enterobacter sp. was identified through the 16S ribotyping method and posed plant growth-promoting properties i.e solubilized phosphate produced indole acetic acid ammonia and hydrogen sulfide. The biocompatibility experiment showed a 4-fold increased mycorrhizal proliferation in MHB added plate compared to the control. Further field experiments suggested the banana plants 10-4-fold increment in plant height (216.54 cm) and stem diameter (10.66 cm). At the same time10-fold significant improvements were noted in leaf carbohydrate (639.9 mg g-1) protein (432.76 mg g-1) chlorophyll (268.66 mg g-1) phenol (1.9 mg g-1) and proline (uMole g-1) content of experimental plant compared to control. The study also showed a 5-fold increase in phosphate content (12.84 mg g-1) than the control (2.4 mg g-1). The isolates increased the mycorrhizal colonization and spore number by 79.33 % and 14.31 g-1 in the rhizospheric soil. Total organic carbon and nitrogen (0.68 %) total phosphorus (55 kg ha-1) and potassium (499.66 kg ha-1) content of soil were positively affected by MHB and AMF. Further-more principal component analysis (PCA) and Pearsons correlation analysis of all the obtained results clearly showed the positive insights of inoculated MHB and AMF on the growth of the banana plant and soil restoration.

Biological synthesis reflects as an eco-friendly, nontoxic and easy method of nanoparticle preparation. Present investigation deals with biosynthesis of silver nanoparticles using Drimia indica leaf extract. Initially the synthesized silver nanoparticles were characterized and confirmed by UV-Vis spectroscopy, X-ray diffraction (XRD) and SEM. The synthesized nanoparticles when used for determination of antibacterial activity, by Microtitre Broth Dilution method exhibited remarkable activity against Pseudomonas aeruginosa, Klebsiella pneumoniae and Escherichia coli.

The current study was carried out to compare the antileishmanial potential of the characterized chemically and plant mediated (Trigonella foenum-graecum) FeO-NPs (FeO-NPs) against L.tropica KWH23. The promastigotes and amastigotes mortality, ROS generation, and biocompatibility assessment were estimated in time-dose dependent manner by PDT. LED exposed bio-inspired FeO-NPs after 72 hours, express significant suppressive effects by exhibiting 50% inhibitory concentration (IC50) 0.001572{+/-}0.02g/ml and 0.011408{+/-}0.02g/ml for promastigotes and amastigotes, respectively. Leishmanial DNA damage and membrane integrity caused by FeO-NPs were owe to high production of reactive oxygen species (ROS). By applying different ROS scavengers (mannitol and sodium azide) hydroxyl radical and singlet oxygen were found as main moieties for cell death by giving quantum yield 0.15 and 0.28 through chemically and green synthesized FeO-NPs, respectively. The light exposed green synthesized nanoparticles were found biocompatible on human red blood cells (RBCs), by exhibiting LD50 value 2659g/mL. From these findings, it can be concluded that plant mediated FeO-NPs can be used as promising antiprotozoal agent.

Metabolites belonging to the propionate metabolism pathway can regulate immune cell responses in the context of autoimmunity and chronic inflammation. Methyl malonic acid (MMA), a metabolite in this pathway is known to cause dysregulation of T cell oxidative phosphorylation (OXPHOS) and downregulating pro inflammatory T cell effector functions. However, the effects of MMA on T cell signaling and T cell activation is not clearly known. Furthermore, since MMA is a small molecule, using it in the context of therapy remains a problem. It gets metabolized in a short time and millimolar concentrations are required to get effective results. This work describes a novel polymer, 1,6 MMA, synthesized using 1,6 Hexane-diol and MMA, which helps in slow, steady and continuous release of the small molecule. Doses in micromolar ranges generate long lasting and robust immunosuppression of activated T cells via an IL2 dependent mechanism in both human and mice T cells without causing non-specific toxicity. This causes a dysregulated expression of pSTAT5 which eventually enhances BLIMP1 mediated T cell apoptosis. Finally, 1,6 MMA mediated T cell suppression is caused due to increase in mitochondrial ROS production. Extrapolation of our findings in-vivo showed the polymer inhibited autoreactive T cell responses in mice with collagen induced arthritis (CIA). Overall, 1,6 MMA, a novel metabolite polymer, has major therapeutic potential in combating inflammatory disorders.

In every region of the world, antibiotic resistance is increasing to dangerously high levels. Our ability to cure widespread infectious diseases is being threatened by the emergence of new resistance mechanisms. Klebsiella pneumoniae is one of the most prevalent nosocomial Gram-negative bacteria in the world. NDM-1 is a brand-new class of metallo-{beta}-lactamase (MBL) that makes the bacteria almost total resistant to all {beta}-lactam antibiotics, including penicillins, cephalosporins and carbapenems. Dangerous infections could develop if NDM-1 switches to a bacterium that is already resistant to antibiotics. It could be untreatable and spread quickly among humans. Using the available insilico tools, in the present research work, an attempt has been made to develop inhibitors for NDM-1 {beta} Lactamase of K. pneumoniae. In the present study, 8 standard ligands were identified and docked against the NDM-1 protein using PyRx. Among these standards, Sulfonamide was selected as the best compound and virtual screening of a large number of sulfonamide moieties was conducted with the identified active site in NDM-1 protein using PyRx. On analyzing the obtained results, about 60 molecules were selected as best hits for Docking studies and ADMET studies. On analyzing the ADMET properties and binding energies of the top hits, (3Z)-N-hydroxypenta-1,3-diene-2-sulfonamide (Pubchem CID 118156306) and N-hydroxyfuran-2-sulfonamide (Pubchem CID 46175386). Molecular dynamics simulations were carried out on the complxes formed by these 2 compounds with target protein and the results showed the complexes were stable which validated the earlier findings. Based on the findings of the study, it was concluded that (3Z)-N-hydroxypenta-1,3-diene-2-sulfonamide (Pubchem CID 118156306) and N-hydroxyfuran-2-sulfonamide (Pubchem CID 46175386) had the potential to be used as lead candidates against infections caused by K. pneumoniae producing NDM-1 strains.

Tyramine-Fe complex is a bioinorganic complex that is formed in brotowali. The complex is thought to play a role in the anti-inflammatory activity of brotowali. The mechanism of anti-inflammatory drugs such as aspirin and ibuprofen is to inhibit the reaction of prostaglandin formation from COX2 and arachidonic acid. In this study, a docking simulation was performed between COX2 with single tyramine, tyramine-Fe complex ibuprofen, and aspirin. The results showed that the COX2-single tyramine bond overlapped with COX2-aspirin, while COX2-tyramine-Fe complex bond overlapped with COX2-ibuprofen. The energy required to form COX2-tyramine-Fe complex bond was smaller than that of single tyramine, aspirin, and ibuprofen. The number of bonds of COX2-tyramine-Fe complex bond is higher than that of single tyramine. These results proved that COX2-tyramine-Fe complex bond had anti-inflammatory activity and greater activity than single tyramine.

Trimethyleneamine N-oxide (TMAO); a gut microbiota derived metabolite has been involved in human health and diseases. It is enhanced by insulin resistivity and linked with various metabolic syndromes in human being such as renal, neuro-degenerative, and cardiovascular diseases. The primary mechanism through which TMAOs promotes disease is not clear yet. TMAO with MW= 75.11 g/mol is a small biomolecule hence, it becomes crucial to develop the conjugate of TMAO with BSA for aptamer synthesis. The binding interactions among TMAO and BSA were investigated using spectroscopic methods like UV-Vis, photoluminescence, Fourier transform infrared and circular dichroism. Hydrophilicity/Hydrophobicity of the conjugate was monitored by using contact angle ([O]) measurement. Sodium dodecyl sulphate polyacryl amide gel electrophoresis (SDS-PAGE) confirmed the different ratio of conjugate formation with the help of band size. This interaction study reveals that TMAO bind with BSA on two sites and with high affinity on one site. Docking studies also showed TMAO is involved in non-covalent interaction with bovine serum albumin forming stable docking complex with binding score of -3.6 kcal/mol obtained from the docking simulation. TMAO is involved in interaction with BSA via amino acid residues forming the stable docking complex through hydrogen bond and electrostatic interaction. This kind of interaction study may be helpful in making strategies to break the conjugation between serum albumin and uremic toxin and pave the way for the treatment for CKD and other diseases wherein TMAO is implicated. Also, conjugation of TMAO and BSA studied here may also serve as premise to develop aptamers for the detection of TMAO in the body fluids.

Traditional drug design focus on specific target (s) expressed by cancer cells. However, cancer cells outsmart the interventions by activating survival pathways and/or downregulating cell death pathways. As the research in molecular biology of cancer grows exponentially, new methods of drug designs are needed to target multiple pathways/targets which are involved in survival of cancer cells. Vitamin E analogues including a-tocopheryl succinate (-TOS) is a well-known anti-tumoregenic agent which is well studied both in vitro and in vivo tumor models. However, lack of targeting cancer cells and unexpected toxicity along with the poor water solubility of -TOS compelled a rational drug design using both targeting and cleavable technologies incorporated in the new drug design. A plethora of Vitamin E derivatives (AMP-001, 002 and 003) were synthesized, characterized and studied for the improved efficacy and lowered toxicity in various cancer cells in vitro. Preliminary studies revealed AAAPT leading candidates reduced the invasive potential of brain tumor stem cells, synergized with different drugs and different treatments. AAAPT leading drug AMP-001 enhanced the therapeutic index of front-line drug Doxorubicin in triple negative breast cancer (TNBC) tumor rat model preserving the ventricular function when used as a neoadjuvant to Doxorubicin. These results may pave the way for reducing the cardiotoxicity of chemotherapy in clinical settings.

Plant diseases pose threat to global food security. The excessive use of synthetic agro-chemical engender pesticide resistance. The exploration of alternative sustainable diseases management practices are crucial to overcome the devastative plant diseases. In this study, a facile innocuous approach was adopted for biogenic synthesis of Fe2O3-CsNPs via Crocus sativus corm aqueous extract and was evaluated for their antifungal efficacy against the Verticillium wilt pathogen Verticillium dahliae. The physico-chemical characterization of biosynthesized nanoparticles were performed through UV-visible Spectroscopy, Fourier Transform Infrared Spectroscopy, Energy Dispersive X-ray Spectroscopy, X-Ray Diffraction, and Scanning Electron Microscopy. The fungus mycelium growth was significantly inhibited in the media containing 3mg/mL Fe2O3-CsNPs. Degenerated, concentrated and shriveled hyphae were revealed in Scanning Electron Microscopy. The overall results demonstrated that the biogenic Fe2O3-CsNPs have the efficacy to control devastative phytopathogens.

BackgroundTerpenoids form a large pool of highly diverse organic compounds possessing several economically important properties, including nutritional, aromatic, and pharmacological properties. The DXP pathways end enzyme, nuclear distribution protein (NudF), interacting with isopentenyl pyrophosphate (IPP) and dimethylallyl pyrophosphate (DMAPP), is critical for the synthesis of isoprenol/prenol/downstream compounds. The enzyme is yet to be thoroughly investigated to increase the overall yield of terpenoids in the Bacillus subtilis, which is widely used in industry and is generally regarded as safe (GRAS) bacterium. The study aims to analyze the evolutionary conservation across the active site, and map the key residues for mutagenesis studies. The study would allow us customize the metabolic load towards the synthesis of prenol or isoprenol or any of the downstream molecules. ResultsThe 37-sequence dataset, extracted from 103 Bacillus subtilis entries, show a high phylogenetic divergence, and only six one-motif sequences ASB92783.1, ASB69297.1, ASB56714.1, AOR97677.1, AOL97023.1, and OAZ71765.1 show monophyly relationship, unlike a complete polyphyly relationship between the other 31 three-motif sequences. Further, only 47 of 179 residues of the representative sequence CUB50584.1 are observed to be significantly conserved. Docking analysis shows a preferential bias of ADP-ribose pyrophosphatase towards IPP, and a nearly 3-fold energetic difference is observed between IPP and DMAPP. Computational saturation mutagenesis of the seven hotspot residues identifies two key positions LYS78 and PHE116, encoded within loop1 and loop7, majorly interact with the ligands DMAPP and IPP, and their mutants K78I/K78L and PHE116D/PHE116E are found to stabilize the overall conformation. The loops are hereby shown to play a regulatory role in guiding the promiscuity of NudF towards a specific ligand. ConclusionThe study map the phylogenetic relationship between the 37 representative B.subtiis NudF sequences, and through sequence conservation, structural contact map, topological flexibility, and saturation mutagenesis of the active site residues, the essential residues regulating the interaction of NudF with IPP/DMAPP are deciphered. The study robustly screens its mutational landscape and localizes the two crucial residues LYS78 and PHE116 for directing the mutagenesis studies. The preliminary docking and simulation results also suggest a preferential bias of ADP-ribose pyrophosphatase towards IPP over DMAPP. The findings would pave the way for the development of novel enzyme variants with highly improved catalytic ability for the large-scale bioproduction of specific terpenoids with significant neutraceutical or commercial value.