Differential action of proteases from Trimeresurus malabaricus, Naja naja and Daboia russellii venoms on hemostasis

The action of venom proteases and their role in hemostasis has been compared in the venoms of Trimeresurus malabaricus, Daboia russellii and Naja naja from the Southern region of Western Ghats, India. These venoms exhibit varying amounts of proteolytic activity and also influence hemostasis differently. Casein hydrolyzing activity of T. malabaricus venoms was 16 and 24 fold higher than those of N. naja and D. russellii venoms, respectively. With the synthetic substrate TAME, the highest activity was observed in T. malabaricus venom. N. naja venom did not hydrolyze TAME even at higher concentrations. These variations in proteolytic activity also influenced the coagulation process. T. malabaricus and D. russellii venoms are strongly procoagulant and reduce the re-calcification time from 148 to 14 and 12 s, respectively. Similarly, both T. malabaricus and D. russellii venoms reduce the prothrombin time from 12.5 to 6.0 s. On the other hand, N. naja venom is anticoagulant and prolongs re-calcification time to 600 s and prothrombin time to 42 s. In spite of varied effects on hemostasis, all the venoms hydrolyze fibrinogen. T. malabaricus venom hydrolyses both Aalpha and Bbeta subunits. While D. russellii and N. naja venoms hydrolyse only Aalpha. None of these venoms hydrolyze the gamma subunit of fibrinogen. Inhibition studies with specific protease inhibitors revealed that both N. naja and T. malabaricus venoms contain only metalloproteases. D. russellii venom contained both serine and metalloproteases. Only, T. malabaricus venom exhibited thrombin-like activity and induces fibrin clot formation with purified fibrinogen within 58 s. Even though D. russellii venom exhibits procoagulant activity, it did not show thrombin-like activity and may act on other coagulation factors.     

Subdomain 3 of Plasmodium falciparum VAR2CSA DBL3x Is Identified as a Minimal Chondroitin Sulfate A-binding Region

Molecular interactions between the VAR2CSA protein, expressed on the surface of Plasmodium falciparum-infected erythrocytes, and placental chondroitin sulfate A (CSA) are primarily responsible for pregnancy-associated malaria (PAM). Interrupting these interactions may prevent or ameliorate the severity of PAM. Several of the Duffy binding-like (DBL) domains of VAR2CSA, including the DBL3x domain, have been shown to bind CSA in vitro, but a more detailed understanding of how DBL domains bind CSA is needed. In this study, we demonstrate that subdomain 3 (S3), one of the three subdomains of VAR2CSA DBL3x by itself, is the major contributor toward CSA binding. NMR spectroscopy and flow cytometry analyses show that S3 and the intact DBL3x domain bind CSA similarly. Mutations within the S3 portion of DBL3x markedly affect CSA binding. Both recombinant molecules, S3 and DBL3x, are recognized by antibodies in the plasma of previously pregnant women living in malaria-endemic regions of Mali, but much less so by plasma from men of the same regions. As the S3 sequence is highly conserved in all known VAR2CSA proteins expressed by different parasite isolates obtained from various malaria endemic areas of the world, the identification of S3 as an independent CSA-binding region provides a compelling molecular basis for designing interventions against PAM.     

Deficiency of mannose-binding lectin greatly increases susceptibility to postburn infection with Pseudomonas aeruginosa

Burn injury disrupts the mechanical and biological barrier that the skin presents against infection by symbionts like the Pseudomonas aeruginosa, a Gram-negative bacteria. A combination of local factors, antimicrobial peptides, and resident effector cells form the initial response to mechanical injury of the skin. This activity is followed by an inflammatory response that includes influx of phagocytes and serum factors, such as complement and mannose-binding lectin (MBL), which is a broad-spectrum pattern recognition molecule that plays a key role in innate immunity. A growing consensus from studies in humans and mice suggests that lack of MBL together with other comorbid factors predisposes the host to infection. In this study we examined whether MBL deficiency increases the risk of P. aeruginosa infection in a burned host. We found that both wild-type and MBL null mice were resistant to a 5% total body surface area burn alone or s.c. infection with P. aeruginosa alone. However, when mice were burned then inoculated s.c. with P. aeruginosa at the burn site, all MBL null mice died by 42 h from septicemia, whereas only one-third of wild-type mice succumbed (p = 0.0005). This result indicates that MBL plays a key role in containing and preventing a systemic spread of P. aeruginosa infection following burn injury and suggests that MBL deficiency in humans maybe a premorbid variable in the predisposition to infection in burn victims.     

Dual-function protein in plant defence: seed lectin from Dolichos biflorus (horse gram) exhibits lipoxygenase activity

Plant-pathogen interactions play a vital role in developing resistance to pests. Dolichos biflorus (horse gram), a leguminous pulse crop of the subtropics, exhibits amazing defence against attack by pests/pathogens. Investigations to locate the possible source of the indomitable pest resistance of D. biflorus, which is the richest source of LOX (lipoxygenase) activity, have led to a molecule that exhibits LOX-like functions. The LOX-like activity associated with the molecule, identified by its structure and stability to be a tetrameric lectin, was found to be unusual. The evidence for the lectin protein with LOX activity has come from (i) MALDI-TOF (matrix-assisted laser-desorption ionization-time-of-flight) MS, (ii) N-terminal sequencing, (iii) partial sequencing of the tryptic fragments of the protein, (iv) amino acid composition, and (v) the presence of an Mn2+ ion. A hydrophobic binding site of the tetrameric lectin, along with the presence of an Mn2+ ion, accounts for the observed LOX like activity. This is the first ever report of a protein exhibiting both haemagglutination and LOX-like activity. The two activities are associated with separate loci on the same protein. LOX activity associated with this molecule adds a new dimension to our understanding of lectin functions. This observation has wide implications for the understanding of plant defence mechanisms against pests and the cellular complexity in plant-pathogen interactions that may lead to the design of transgenics with potential to impart pest resistance to other crops.     

Smoke treatment triggers the release of a novel DNA damaging factor by lymphocytes

Abstact Organic fuel smoke is a hazardous agent, which pushes the cells towards“prooxidant state', leading to 4,46,400 strand breaks/cell/day as against 47,000 strand breaks/cell/day produced by constitutive oxygen radicals. This prooxidants scenario switches on a plethora of intercellular events. Here we report a novel DNA damaging factor released by lymphocytes, upon treatment with smoke condensate. Human lymphocytes, when exposed to cow dung cake smoke condensate, were found to release a low molecular weight factor into the media at 20 min of exposure. The conditioned media, displayed a propensity of inducing DNA damage in fresh, normal lymphocytes, which were not exposed to any damaging agent. The above DNA damaging effect of the conditioned media was not due to any residual presence of Polycyclic Aromatic Hydrocarbons, which were present in the smoke. The release of this factor was in correlation with the DNA damaging event, taking place in the cells. This secondary DNA damaging factor had a molecular weight less than 5 kd. The factor had the cell death inducing propensity when allowed to act on lymphocytes     

Characterization of an equine mannose-binding lectin and its roles in disease

The mannose-binding lectin (MBL), a pattern recognition serum protein, participates in the innate immune system of mammals as an opsonin. In humans, MBL plays a key role in first-line host defense against infection during the lag period prior to the development of a specific immune response. MBL also activates complement via the lectin pathway that requires a MBL-associated serine protease-2 (MASP-2). Homologues of human MBL (hMBL) have been identified in a variety of mammals, fish, and primitive animals such as ascidians. In this study, we report that equine MBL (eMBL) has properties that are similar to hMBL. In addition, we found low levels of MBL:MASP activity in sick horses compared to healthy horses. These results suggest that eMBL is involved in the immune response of the horse and that low MBL:MASP activity could be used to monitor immune function and clinical outcome.     

Plasmodium falciparum: chondroitin sulfate A is the major receptor for adhesion of parasitized erythrocytes in the placenta

Plasmodium falciparum parasites that sequester in the placenta bind to the molecule chondroitin sulfate A (CSA). Women become resistant to malaria during pregnancy as they acquire antibodies that inhibit parasite adhesion to CSA, suggesting that a vaccine against placental malaria is feasible. Hyaluronic acid (HA) and non-immune IgG have also been proposed as receptors for P. falciparum adhesion in the placenta, but evidence for their roles is inconclusive. In this study, CSA, HA, and IgG were simultaneously assessed for their relative contributions to placental adhesion. Placental parasites collected in Tanzania uniformly adhered to the molecule CSA, and soluble CSA completely inhibited adhesion of most samples to placental cryosections. Three of 46 placental parasite samples also adhered to immobilized HA, but HA failed to inhibit adhesion of any placental parasites to placental cryosections. Similarly, non-immune IgG and protein A failed to inhibit adhesion of parasite samples to placental cryosection. P. falciparum adhesion in the placenta appears to be a non-redundant process that requires CSA as a receptor. Vaccines that elicit functional antibodies against CSA-binding parasites may confer resistance to pregnancy malaria.     

Structure and activity of glycosylphosphatidylinositol anchors of Plasmodium falciparum

The glycosylphosphatidylinositol (GPI) anchors of Plasmodium falciparum are thought to be etiologic agents of malaria based on their ability to induce proinflammatory cytokine production by macrophages and cause symptoms that resemble severe malaria illness in animals. This review summarizes the published information on the structures of P. falciparum GPIs, structure-activity relationship, and anti-GPI antibodies in the host.     

The mitochondrial genome of the Cinnamon Bittern, Ixobrychus cinnamomeus (Pelecaniformes: Ardeidae): sequence, structure and phylogenetic analysis

Ixobrychus cinnamomeus is a member of the large wading bird family, known as Ardeidae. In the present study, we determined the complete mitochondrial genome of I. cinnamomeus for use in future phylogenetic analysis. This circular mitochondrial genome is 17,180 bp in length and composed of 13 protein-coding genes, 22 tRNA genes, two rRNA genes and one putative control region. Three conserved domains and a minisatellite of 17 nucleotides with 22 tandem repeats were detected at the end of the control region. Phylogenetic relationships were reconstructed using the nucleotide and corresponding amino acid datasets of 12 concatenated protein-coding genes from the mitochondrial genome. Using maximum likelihood, maximum parsimony and Bayesian inference methods, the monophyly of Ciconiidae, Ardeidae and Threskiornithidae were confirmed; however, the monophyly of traditional Ciconiiformes and Pelecaniformes failed to be recovered. Although further studies are recommended to clarify relationships among and within the orders of Ciconiiformes, Pelecaniformes, Suliformes and Phaethontiformes, our results provide preliminary exploratory results that can be useful in the current understanding of avian phylogenetics.