Effect of Different Edible Coatings on Postharvest Quality of Mandarin Orange (Citrus reticulata Blanco)

Although Nepal produces a large amount of mandarin it faces huge postharvest losses due to improper postharvest practices. Treating fruits with different edible coatings can minimize postharvest losses. The experiment was carried out in the horticulture lab of Prithu Technical College, Dang, Nepal to evaluate the effects of different edible coating materials on the postharvest quality of mandarin. The experiment was laid in Complete Randomized Design (CRD) with three replications and seven treatments  in each replication. Mandarins were coated with different edible coating materials i.e. paraffin wax (100%, 75% and 50%), mustard oil, Aloe vera, turmeric paste and control (non-coated). After coating with different edible materials, mandarins were kept at ambient room conditions (18±2℃ and 52.41±14.35%). The lowest physiological loss in weight at 7, 14and 21 days was recorded in mandarin coated with 75% paraffin wax which was 3.10%, 4.83% and 10.33%, respectively. The highest titratable acidity (0.68%), juice content (46.33%) and marketable fruit percentage (81.73%) were recorded in 75% paraffin wax. The highest total soluble solid (14.00 ˚Brix) was recorded in control. Based on the result obtained from our research, it is suggested to use 75% paraffin wax for the storage of mandarin at ambient room conditions (18±2℃ and 52.41±14.35% RH) as it gives a high percentage of marketable fruits and juice content and also minimizes the physiological loss in weight

Development, standardization of polyherbal formulation of analgesic ointment of plant Carum copticum, Mentha piperita, Cedrus deodara

Ayurveda is one of the world’s oldest systems of medicine. It originated in India and has evolved there over thousands of years. The term “Ayurveda” combines then Sanskrit words ayur (life) andVeda (science or knowledge). Ayurveda means “the science of life. Medicinal plants and herbal drugs have played a key role in world health. According to world health organization (WHO), about 80% of the world population currently utilizes the herbal drugs. People are using herbal medicines from centuries for safety, efficacy, cultural acceptability, non-toxic, lesser side effects and easily available at affordable prices. In recent times, there has been a move in universal trend from synthetic to herbal medicine due to side effects of synthetic products. Herbal products may contain a single herb or combinations of several different herbs believed to have complementary and /synergistic effects. Some herbal products, including many traditional medicine formulations, also include animal products and minerals. Herbal products are sold as either raw plants or extracts of portions of the plant or in the form formulation i.e. tablet, capsule, syrup, cream and ointment etc. The different parts of plants with analgesic were taken up for the present study and investigated for the phytochemical screening and used for the formulation of analgesic ointment. Present study deals with formulation, Standardization, evaluation of ointment made from alcoholic extract and essential oil of different plants

Assessment of the protein interaction between coagulation factor XII and corn trypsin inhibitor by molecular docking and biochemical validation

Background: Corn trypsin inhibitor (CTI) has selectivity for serine proteases coagulation factor XII (FXII) and trypsin. CTI is in widespread use as a reagent that specifically inhibits the intrinsic pathway of blood coagulation but not the extrinsic pathway. Objectives: To investigate the molecular basis of FXII inhibition by CTI. Methods: We performed molecular docking of CTI, using its known crystal structure, with a model of the activated FXII (FXIIa) protease domain. The interaction model was verified using a panel of recombinant CTI variants tested for their ability to inhibit FXIIa enzymatic activity using a substrate cleavage assay. Results: The docking predicted that (i) the CTI central inhibitory loop P1 Arg34 side chain forms a salt bridge to the FXIIa S1 pocket Asp189 side chain (ii) residue Trp22 from the CTI helix α1 interacts with the FXIIa S3 pocket (iii) Arg43 from CTI helix α2 forms a salt bridge to FXIIa H1 pocket Asp60A. CTI amino acid substitution R34A negated all inhibitory activity whereas variants G32W, L35A, W22A or R42A-R43A reduced activity by a large degree of 108, 41, 158 and 100-fold respectively, with R27A, W37A, W39A, R42A having no effect. Synthetic peptides spanning CTI residues 20-44 had inhibitory activity 3-4000-fold less than full-length CTI. Conclusions: The data confirm the validity of a canonical model of the FXIIa-CTI interaction with helix α1 (Trp22), central inhibitory loop (Arg34) and helix α2 (Arg43) of CTI required for effective binding by contacting the S1, S3 and H1 pockets of FXIIa, respectively

Expression of human ficolin-2 in hepatocytes confers resistance to infection by diverse hepatotropic viruses

The liver-expressed pattern recognition receptors (PRRs) mannose binding lectin (MBL), ficolin-2 and ficolin-3 contribute to the innate immune response by activating complement. Binding of soluble ficolin-2 to viral pathogens can directly neutralize virus entry. We observed that the human hepatoma cell line HuH7.5, which is routinely used for the study of hepatotropic viruses, is deficient in expression of MBL, ficolin-2 and ficolin-3. We generated a cell line that expressed and secreted ficolin-2. This cell line (HuH7.5 [FCN2]) was more resistant to infection with hepatitis C virus (HCV), ebolavirus (EBOV) and vesicular stomatitis virus (VSV), but surprisingly was more susceptible to infection with rabies virus (RABV). Cell-to-cell spread of HCV was also inhibited in ficolin-2 expressing cells. This illustrates that ficolin-2 expression in hepatocytes contributes to innate resistance to virus infection, but some viruses might utilise ficolin-2 to facilitate entry

Plasma kallikrein structure reveals apple domain disc rotated conformation compared to factor XI

BackgroundPlasma prekallikrein (PK) and factor XI (FXI) are apple domain‐containing serine proteases that when activated to PKa and FXIa cleave substrates kininogen, factor XII, and factor IX, respectively, directing plasma coagulation, bradykinin release, inflammation, and thrombosis pathways.ObjectiveTo investigate the three‐dimensional structure of full‐length PKa and perform a comparison with FXI.MethodsA series of recombinant full‐length PKa and FXI constructs and variants were developed and the crystal structures determined.Results and conclusionsA 1.3 Å structure of full‐length PKa reveals the protease domain positioned above a disc‐shaped assemblage of four apple domains in an active conformation. A comparison with the homologous FXI structure reveals the intramolecular disulfide and structural differences in the apple 4 domain that prevents dimer formation in PK as opposed to FXI. Two latchlike loops (LL1 and LL2) extend from the PKa protease domain to form interactions with the apple 1 and apple 3 domains, respectively. A major unexpected difference in the PKa structure compared to FXI is the 180° disc rotation of the apple domains relative to the protease domain. This results in a switched configuration of the latch loops such that LL2 interacts and buries portions of the apple 3 domain in the FXI zymogen whereas in PKa LL2 interacts with the apple 1 domain. Hydrogen‐deuterium exchange mass spectrometry on plasma purified human PK and PKa determined that regions of the apple 3 domain have increased surface exposure in PKa compared to the zymogen PK, suggesting conformational change upon activation

Structures of factor XI and prekallikrein bound to domain 6 of high–molecular weight kininogen reveal alternate domain 6 conformations and exosites

High molecular weight kininogen (HK) circulates in plasma as a complex with zymogen prekallikrein (PK). HK is both substrate and co-factor for activated plasma kallikrein (PKa) and the principal exosite interactions occur between PK N-terminal apple domains and the C-terminal D6 domain of HK. To determine the structure of the complex formed between PK apple domains and a HKD6 fragment and compare this to the FXI-HK complex. We produced recombinant FXI and PK heavy chains (HC) spanning all four apple domains. We co-crystallised PKHC (and subsequently FXIHC) with a 31 amino acid synthetic peptide spanning HK residues Ser565-Lys595 and determined the crystal structure. We also analysed the full length FXI-HK complex in solution using hydrogen deuterium exchange mass spectrometry (HDX-MS). The 2.3Å PKHC-HK peptide crystal structure revealed that the HKD6 sequence WIPDIQ (Trp569-Gln574) binds to the apple 1 domain and HK FNPISDFPDT (Phe582-Thr591) binds to the apple 2 domain with a flexible intervening sequence resulting in a bent double conformation. A second 3.2Å FXIHC-HK peptide crystal structure revealed a similar interaction with the apple 2 domain but an alternate, straightened conformation of the HK peptide where residues LSFN (Leu579-Asn583) interacts with a unique pocket formed between the apple 2 and 3 domains. HDX-MS of full length FXI-HK complex in solution confirmed interactions with both apple 2 and apple 3. The alternate conformations and exosite binding of the HKD6 peptide likely reflects the diverging relationship of HK to the functions of PK and FXI. [Abstract copyright: Crown Copyright © 2023. Published by Elsevier Inc. All rights reserved.

Characterisation of geranylgeranyl diphosphate synthase from the sandfly Lutzomyia longipalpis

Leishmaniasis is a debilitating and often fatal neglected tropical disease. Males from sub-populations of the Leishmania-harbouring sandfly, Lutzomyia longipalpis, produce the diterpene sex and aggregation pheromone, sobralene, for which geranylgeranyl diphosphate (GGPP) is the likely isoprenoid precursor. We have identified a GGPP synthase (lzGGPPS) from L. longipalpis, which was recombinantly expressed in bacteria and purified for functional and kinetic analysis. In vitro enzymatic assays using LC-MS showed that lzGGPPS is an active enzyme, capable of converting substrates dimethylallyl diphosphate (DMAPP), (E)-geranyl diphosphate (GPP), (E,E)-farnesyl diphosphate (FPP) with co-substrate isopentenyl diphosphate (IPP) into (E,E,E)-GGPP, while (Z,E)-FPP was also accepted with low efficacy. Comparison of metal cofactors for lzGGPPS highlighted Mg2+ as most efficient, giving increased GGPP output when compared against other divalent metal ions tested. In line with previously characterised GGPPS enzymes, GGPP acted as an inhibitor of lzGGPPS activity. The molecular weight in solution of lzGGPPS was determined to be ∼221 kDa by analytical SEC, suggesting a hexameric assembly, as seen in the human enzyme, and representing the first assessment of GGPPS quaternary structure in insects

"Like sugar in milk": reconstructing the genetic history of the Parsi population.

BACKGROUND: The Parsis are one of the smallest religious communities in the world. To understand the population structure and demographic history of this group in detail, we analyzed Indian and Pakistani Parsi populations using high-resolution genetic variation data on autosomal and uniparental loci (Y-chromosomal and mitochondrial DNA). Additionally, we also assayed mitochondrial DNA polymorphisms among ancient Parsi DNA samples excavated from Sanjan, in present day Gujarat, the place of their original settlement in India. RESULTS: Among present-day populations, the Parsis are genetically closest to Iranian and the Caucasus populations rather than their South Asian neighbors. They also share the highest number of haplotypes with present-day Iranians and we estimate that the admixture of the Parsis with Indian populations occurred ~1,200 years ago. Enriched homozygosity in the Parsi reflects their recent isolation and inbreeding. We also observed 48% South-Asian-specific mitochondrial lineages among the ancient samples, which might have resulted from the assimilation of local females during the initial settlement. Finally, we show that Parsis are genetically closer to Neolithic Iranians than to modern Iranians, who have witnessed a more recent wave of admixture from the Near East. CONCLUSIONS: Our results are consistent with the historically-recorded migration of the Parsi populations to South Asia in the 7th century and in agreement with their assimilation into the Indian sub-continent's population and cultural milieu "like sugar in milk". Moreover, in a wider context our results support a major demographic transition in West Asia due to the Islamic conquest

Crystal structures of the recombinant β-factor XIIa protease with bound Thr-Arg and Pro-Arg substrate mimetics

© 2019 International Union of Crystallography. Coagulation factor XII (FXII) is a key initiator of the contact pathway, which contributes to inflammatory pathways. FXII circulates as a zymogen, which when auto-activated forms factor XIIa (FXIIa). Here, the production of the recombinant FXIIa protease domain (βFXIIaHis) with yields of ~1–2 mg per litre of insect-cell culture is reported. A second construct utilized an N-terminal maltose-binding protein (MBP) fusion (MBP-βFXIIaHis). Crystal structures were determined of MBP-βFXIIaHis in complex with the inhibitor d-Phe-ProArg chloromethyl ketone (PPACK) and of βFXIIaHis in isolation. The βFXIIaHis structure revealed that the S2 and S1 pockets were occupied by Thr and Arg residues, respectively, from an adjacent molecule in the crystal. The Thr-Arg sequence mimics the P2–P1 FXIIa cleavage-site residues present in the natural substrates prekallikrein and FXII, and Pro-Arg (from PPACK) mimics the factor XI cleavage site. A comparison of the βFXIIaHis structure with the available crystal structure of the zymogen-like FXII protease revealed large conformational changes centred around the S1 pocket and an alternate conformation for the 99-loop, Tyr99 and the S2 pocket. Further comparison with activated protease structures of factors IXa and Xa, which also have the Tyr99 residue, reveals that a more open form of the S2 pocket only occurs in the presence of a substrate mimetic. The FXIIa inhibitors EcTI and infestin-4 have Pro-Arg and Phe-Arg P2–P1 sequences, respectively, and the interactions that these inhibitors make with βFXIIa are also described. These structural studies of βFXIIa provide insight into substrate and inhibitor recognition and establish a scaffold for the structure-guided drug design of novel antithrombotic and antiinflammatory agents