Pore-forming Activity of the Escherichia coli Type III Secretion System Protein EspD

Enterohemorrhagic Escherichia coli is a causative agent of gastrointestinal and diarrheal diseases. Pathogenesis associated with enterohemorrhagic E. coli involves direct delivery of virulence factors from the bacteria into epithelial cell cytosol via a syringe-like organelle known as the type III secretion system. The type III secretion system protein EspD is a critical factor required for formation of a translocation pore on the host cell membrane. Here, we show that recombinant EspD spontaneously integrates into large unilamellar vesicle (LUV) lipid bilayers; however, pore formation required incorporation of anionic phospholipids such as phosphatidylserine and an acidic pH. Leakage assays performed with fluorescent dextrans confirmed that EspD formed a structure with an inner diameter of ∼2.5 nm. Protease mapping indicated that the two transmembrane helical hairpin of EspD penetrated the lipid layer positioning the N- and C-terminal domains on the extralumenal surface of LUVs. Finally, a combination of glutaraldehyde cross-linking and rate zonal centrifugation suggested that EspD in LUV membranes forms an ∼280–320-kDa oligomeric structure consisting of ∼6–7 subunits.     

Changes in chemical composition and caloric content of developing eggs of the shrimpcrangon crangon

Summary 1. Changes in chemical composition and caloric content as well as the cumulative efficiencies of yolk utilization have been studied in the developing eggs and freshly hatched protozoea of the shrimpCrangon crangon L.2. Per unit dry weight of the fresh egg the following relative increases were observed during the development: 16.8% water, 5.9% ash, 10.6% protein, and 0.5% non-protein nitrogen. During the same period (fresh egg to freshly hatched protozoea) fat content decreased from 32.6% to 15.6% and energy content from 6443 to 5287 cal/g dry organic substance.3. The cumulative efficiencies of yolk utilization for the different constituents varied; they were 70.3% for dry weight, 54.0% for total energy, 83.0% for protein, and 33.6% for fat.4. Of the 0.0453 cal expended on the metabolic processes of the embryo, only 20.8% was drawn from the oxidation of protein, while fat oxidation contributed as much as 75.0%.5. Considerable quantities of inorganic salts (0.29µg/egg) were absorbed from the surrounding sea-water by the egg during its development.

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Änderungen in der chemischen Zusammensetzung und im Kaloriengehalt sich entwickelnder Eier der GarneleCrangon crangon

Kurzfassung Pro Trockengewichtseinheit ergeben sich während der gesamten Eientwicklung vonCrangon crangon L. folgende relative Zunahmen: Wasser 16,8%, Asche 5,9%, Eiweiß 10,6%, Nicht-Eiweiß Stickstoff 0,5%. In gleichem Zeitraum sinken die entsprechenden Werte für den Fettgehalt von 32,6% auf 15,6% und für den Energiegehalt von 6443 auf 5287 cal/g organische Substanz. Für die Entwicklung der aus dem Ei geschlüpften Protozoea beträgt der Kumulativ-Nutzeffekt 70,3% für Trockengewicht, 54,0% für Energie, 83,0% für Eiweiß und 33,6% für Fett. Während der ganzen Entwicklung eines Eies werden im Mittel 0,0453 cal für den Stoffwechsel aufgewendet; davon stammen 20,8% aus dem Eiweiß und 75,0% aus der Fettoxydation. Im Verlauf der Eientwicklung werden offensichtlich beträchtliche Mengen anorganischer Salze (0,29µg pro Ei) aus dem umgebenden Meerwasser absorbiert.

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This paper is based on a lecture presented during the Annual meeting of the International Council for the Exploration of the Sea in Hamburg, October, 1967.

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Dedicated to Prof.R. V. Seshaiya, the founder of the Marine Biological Laboratory, Porto Nova, South India, on his 70^th birthday.     

Deciphering structural aspects of reverse cholesterol transport: mapping the knowns and unknowns

Atherosclerosis is a major contributor to the onset and progression of cardiovascular disease (CVD). Cholesterol-loaded foam cells play a pivotal role in forming atherosclerotic plaques. Induction of cholesterol efflux from these cells may be a promising approach in treating CVD. The reverse cholesterol transport (RCT) pathway delivers cholesteryl ester (CE) packaged in high-density lipoproteins (HDL) from non-hepatic cells to the liver, thereby minimising cholesterol load of peripheral cells. RCT takes place via a well-organised interplay amongst apolipoprotein A1 (ApoA1), lecithin cholesterol acyltransferase (LCAT), ATP binding cassette transporter A1 (ABCA1), scavenger receptor-B1 (SR-B1), and the amount of free cholesterol. Unfortunately, modulation of RCT for treating atherosclerosis has failed in clinical trials owing to our lack of understanding of the relationship between HDL function and RCT. The fate of non-hepatic CEs in HDL is dependent on their access to proteins involved in remodelling and can be regulated at the structural level. An inadequate understanding of this inhibits the design of rational strategies for therapeutic interventions. Herein we extensively review the structure–function relationships that are essential for RCT. We also focus on genetic mutations that disturb the structural stability of proteins involved in RCT, rendering them partially or completely non-functional. Further studies are necessary for understanding the structural aspects of RCT pathway completely, and this review highlights alternative theories and unanswered questions.     

Selective incorporation of 5-hydroxytryptophan blocks long range electron transfer in oxalate decarboxylase

Oxalate decarboxylase from Bacillus subtilis is a binuclear Mn-dependent acid stress response enzyme that converts the mono-anion of oxalic acid into formate and carbon dioxide in a redox neutral unimolecular disproportionation reaction. A π-stacked tryptophan dimer, W96 and W274, at the interface between two monomer subunits facilitates long-range electron transfer between the two Mn ions and plays an important role in the catalytic mechanism. Substitution of W96 with the unnatural amino acid 5-hydroxytryptophan leads to a persistent EPR signal which can be traced back to the neutral radical of 5-hydroxytryptophan with its hydroxyl proton removed. 5-Hydroxytryptophan acts as a hole sink preventing the formation of Mn(III) at the N-terminal active site and strongly suppresses enzymatic activity. The lower boundary of the standard reduction potential for the active site Mn(II)/Mn(III) couple can therefore be estimated as 740 mV against the normal hydrogen electrode at pH 4, the pH of maximum catalytic efficiency. Our results support the catalytic importance of long-range electron transfer in oxalate decarboxylase while at the same time highlighting the utility of unnatural amino acid incorporation and specifically the use of 5-hydroxytryptophan as an energetic sink for hole hopping to probe electron transfer in redox proteins.     

Treatment of periodontal disease using xanthan based chlorhexidine gel

People of all ages are suffering from periodontal disease. It causes indirect damage in the oral cavity. It is of interest to evaluate the efficacy of xanthan-based chlorhexidine gel (Xan-CHX) in patients with mild-severe chronic periodontitis. Five patients with 60 sites were divided in two groups. Group A (treated with SRP) and group B (treated with Chlosite i.e., SRP + CHL). The recorded clinical parameters were Plaque index (PI), Gingival index (GI), Bleeding index (BI), and Clinical attachment Level (CAL) with sub gingival plaque subjected to microbial analysis. Significant reduction was observed in both groups. However, group B (treated with Chlosite i.e., SRP + CHL) showed statistically significant improvement on above mentioned parameters as compared to group A. Data suggest that in the treatment of periodontal disease (viz. PI, GI, BI and CAL) combination of SRP and Chlosite showed added benefits over only SRP.     

Chemically characterised Artemisia nilagirica (Clarke) Pamp. essential oil as a safe plant-based preservative and shelf-life enhancer of millets against fungal and aflatoxin contamination and lipid peroxidation

Abstract

The study recommends the Artemisia nilagirica (Clarke) Pamp. essential oil (ANEO) as plant-based shelf-life enhancer of millets against fungal, aflatoxin B₁ (AFB₁) contamination and lipid peroxidation with favourable safety profile. Chemical characterisation of ANEO through GC-MS, recorded 1,5-heptadiene-4-one,3,3,6-trimethyl (32.72%)as the main compound, followed by Artemisia alcohol (13.40%), alpha lonone (4.55%), benzene, methyl (1-methylethyl) (2.97%) and 1-cyclohexene-1-carboxaldehyde,4-(1-methylethyenyl) (2.23%). The mycoflora analysis of millet samples showed Aspergillus flavus strain[LHP(R)-5] as the most AFB₁ secreting strain. The ANEO inhibited growth and AFB₁ production by the toxigenic strain at 1.4 and 1.0?µL?mL^?1, respectively, and also possess broad fungitoxic spectrum. The decrement in membrane ergosterol content, enhanced leakage of cellular Ca²⁺, K⁺ and Mg²⁺ ions along with SEM and TEM study of ANEO-treated cell denotes plasma membrane as antifungal site of action. The ANEO also showed strong antioxidant activity as determined by DPPH^? and ABTS^?+ assays having IC₅₀ value 2.51 and 1.07?µL?mL^?1, respectively. More than 70.78% protection of Ragi samples from fungal contamination was observed during in situ trial. The ANEO showed favourable safety profile with high LD₅₀ value (7528.10?µL?kg^?1) for male mice and also exhibited non-phytotoxicity for Ragi seeds germination.     

Assessment of genetic diversity and population structure in pomegranate (Punica granatum L.) using hypervariable SSR markers

Physiology and Molecular Biology of Plants - The present study investigates the genetic diversity and population structure among 42 diverse pomegranate genotypes using a set of twenty one class I...     

Comparative analysis of metabolites in contrasting chickpea cultivars

Journal of Plant Biochemistry and Biotechnology - Chickpea (Cicer arietinum L.) is a good source of nutrients for animals and human consumption. In the present study, we analyzed the anthocyanin...