The role of C-terminus carbohydrate-binding domain of Vibrio cholerae haemolysin/cytolysin in the conversion of the pre-pore β-barrel oligomer to a functional diffusion channel.

Background & objectives: Vibrio cholerae cytolysin/hemolysin (VCC) is a 65 kDa pore-forming toxin (PFT) secreted by O1 El Tor and non-O1 strains. The purified toxin, which contains two C-terminus carbohydrate-binding domains in addition to the cytolytic domain at the core, causes lysis of a wide spectrum of eukaryotic cells at picomolar concentrations, apoptogenesis of intestinal and immune cells and accumulation of fluid in rabbit ligated ileal loop. Therefore, it may potentially complement the action of cholera toxin (CT) in diarrheagenic strains that do not produce CT. We showed earlier that β1-galactosyl-terminated glycoconjugates are strong inhibitors of its pore-forming activity, though carbohydrates are not functional receptors of VCC. Here, we investigate how the 15 kDa C-terminus β-prism lectin domain contributed to pore formation in erthrocytes. Methods: VCC was isolated from the culture supernatant of late log phase grown bacteria and purified to homogeneity by chromatography. The 50 kDa truncated variant was generated by restricted proteolysis. Liposome was prepared by sonication of a suspension of phospholipids and calceine release assay was done by spectrofluorometric monitoring of the released dye trapped in liposome. Formation of β-barrel oligomers in erythrocyte stroma was monitored by scanning electron microscopy. Results: Proteolytic truncation of the C-terminus β-prism lectin domain decreased hemolytic activity of the toxin by ~800-fold without causing a significant change in pore-forming activity toward synthetic lipid vesicles devoid of incorporated glycoproteins/glycolipids. Truncation at the C-terminus did not impair membrane-binding or assembly to the oligomeric pore. Interpretation & conclusions: Our data indicated that the C-terminus domain played a critical role in translocation of the pre-pore oligomeric assembly from the cell surface or lipid-water interface to the hydrocarbon core of the membrane bilayer, signaling the formation of functional diffusion channels.

Condom leukoderma.

Contact dermatitis from natural latex of condom has been reported and is attributed to latex sensitivity. Chemical leukoderma from rubber condom is probably not reported. Here we present a case of chemical leukoderma in a 32-year-old male who developed depigmentation around the shaft of the penis in a circumferential pattern. Since the lesion was solitary and the site corresponded to the point of maximum contact of the condom, a diagnosis of contact leukoderma due to latex condom was thought of. Patch testing was done with mercaptobenzothiazole (MBT), dusting powder present in the condom and condom latex as such. The patient tested positive (3+) with mercaptobenzothiazole and the condom latex. On discontinuation of condom use and with UVB phototherapy, lesions repigmented in eight weeks.

Amulet string contact leukoderma and its differentiation from vitiligo.

A young adult factory worker presented with a linear depigmented vitiliginous patch on his right arm at the site where a silver amulet had been fixed with a nylon thread. He claimed that it was occupational in origin and demanded compensation, but patch testing with the nylon thread of the amulet and its extracted dyes proved that the contact leukoderma was due to the thread.

Damodar river pollution and health hazards.

Damodar river is one of the most polluted rivers in this country. Chemical pollution and bacteriological pollution, both are high--leading to rising trend in different industrial health hazards in the Damodar basin.

Injuries and granuloma caused by water slider.

Bite of waters slider (Ranatra) producing granuloma, numbness of the arm followed by neurotoxicity in the form of acute abdomen is being reported.

Haemolysis of rabbit erythrocytes by a lectin from the seeds of Croton tiglium.

The kinetics of haemolysis of rabbit erythrocytes by Croton tiglium lectin was studied as a function of concentration of the lectin and erythrocytes. The length of the prelytic period decreased with increasing lectin concentrations, indicating that the secondary events at the membrane which follow the binding of the lectin to cell surface carbohydrate receptors are accelerated at higher surface concentrations of the lectin. The rate or extent of haemolysis was not affected by the inclusion of ions like K+, Ca2+ and Mg2+ in the medium or by the substitution of ionic medium by a non-ionic medium. The inhibition of haemagglutination and haemolysis of rabbit red cells by Croton tiglium lectin by antilectin rabbit serum was observed. A possible mechanism of haemolysis by the lectin is discussed.

Analysis of alphavirus polypeptides by two dimensional polyacrylamide gel electrophoresis.

Semliki Forest, Sindbis and Chikungunya viruses were grown and radiolabeled with [3H]-amino acids in Vero cells. Analysis of virus infected cell lysates by two dimensional polyacrylamide gel electrophoresis resulted in detection of polypeptides of molecular, weights corresponding to those of E1, P62, ns60, ns70/72 for Semliki Forest virus, the C, E1, 6K, 14Κ, ΡΕ2, P97, ns60, ns82 for Sindbis virus and E1. P62, P97, ns70/72 for Chikungunya virus. Charge and molecular weight heterogeneity in the precursor polypeptide P62 of Semliki Forest virus was detected. Structural polypeptides e.g. E1 and E2 of Semliki Forest virus and C, E1, E2 of Sindbis virus and E1 of Chikungunya virus were detected when purified radiolabeled virus preparations were analyzed by two dimensional polyacrylamide gel-electrophoresis. Membrane glycoprotein Ε1 and E2 of Semliki Forest and Ε1 of Sindbis and Chikungunya viruses exhibited charge heterogeneity. In contrast to the marked difference in isoelectric points of E1 and E2 of Sindbis virus; E1 and E2 of Semliki Forest virus had almost identical isoelectric points.