Cloning,sequencing and partial characterisation of sorbitol transporter (srlT) gene encoding phosphotransferase system,glucitol/sorbitol-specific IIBC components of Erwinia herbicola ATCC 21998 – Cloning,sequencing and partial characterisation of sorbitol specific transporter (srlT) gene of Erwinia herbicola ATCC 21998

A DNA fragment of approximately 1500 bp, harbouring the sorbitol transport gene (srlT), was amplified from the chromosomal DNA of Erwinia herbicola ATCC 21998 by PCR and cloned in Escherichia coli JM109. Degenerate oligonucleotide primers used were designed based on the conserved regions in the gene sequences within the gut operon of E. coli (Gene Bank accession no. J02708) and the srl operon of Erwinia amylovora (Gene Bank accession no. Y14603). The cloned DNA fragment was sequenced and found to contain an open reading frame of 1473 nucleotides coding for a protein of 491 amino acids, corresponding to a mass of 52410 Da. The nucleotide sequence of this ORF was highly homologous to that of the gutA gene of Escherichia coli gut operon, the srlE gene of Shigella flexrni and the sorbitol transporter gene sequence of Escherichia coli K12 (Gene Bank accession nos. J02708, AE016987 and D90892 respectively). The protein sequence showed significant homology to that of the phosphotransferase system i.e. the glucitol/sorbitol-specific IIBC components of Escherichia coli and Erwinia amylovora (P56580, O32522). The cloned DNA fragment was introduced into a pRA90 vector and the recombinant was used for developing srlT mutants of Erwinia herbicola, by homologous recombination. Mutants obtained were unable to grow on minimal medium with sorbitol. The insertion of the pRA90 vector inside the srlT gene sequence of the mutants was confirmed by DNA-DNA hybridisation.     

Role of chelating agents and antioxidants in beryllium induced toxicity

The present study was conducted to evaluate the therapeutic effectiveness of chelating agents [glutathione, 2,3 dimercapto propane sulfonic acid (DMPS) and D-penicillamine (DPA)] in combination with antioxidant (sodium selenite) in beryllium induced toxicity in female rats. A bolus dose of 50mg/kg-beryllium nitrate was administered singly followed by chelation therapy with GSH, DMPS + Se and DPA + Se at various durations of 1,3 and 7 days respectively. Results revealed a significant fall in the glycogen content, whereas, a marginal fall in the protein was also observed. The enzymatic activity of alkaline phosphatase and adenosine triphosphatase was depleted; on the contrary, there was a significant rise in the acid phosphatase and glucose-6-phosphatase pattern. A rise in the hepatic lipid peroxidation activity is a direct indication of oxidative damage resulting in free radical generation. The distribution of the metal by atomic absorption spectrophotometry revealed an increased concentration of beryllium in liver and kidney, followed by lung and uterus. The relative ability of three chelating agents to act as antagonists, for acute beryllium poisoning, have been examined in liver, kidney, lungs and uterus. The appreciable change in the beryllium concentration in various organs is duration dependent during the entire period being highly significant at 7 days regimen. Biochemical and distribution studies reveal that DPA + Se was the most effective therapeutic agent followed by DMPS + Se and GSH.     

Cell mediated immunity during experimental ancylostomiasis: a review

Cell-mediated immunity is usually recognized only in a very small number of infections caused by helminths. The present review is engaged with some aspects of the cell-mediated immunity in infections caused by Ancylostoma caninum. Special attention is paid to the role of passive transfer of sensitized cells and serum. The paper is based on a series of own original investigations of the authors.     

Degradation of alpha, beta, gamma and delta-hexachlorocyclohexanes by Sphingomonas paucimobilis

Sphingomonas paucimobilis degrades aerobically , , and -hexachlorocyclohexane. With -HCH, complete degradation occurred after 3 days but with and , and with -HCH, 98 and 56 % degradation occurred after 12 and 8 days of incubation, respectively. Pentachlorocyclohexene was formed as the primary metabolite during the degradation of all the HCH isomers. © Rapid Science Ltd. 1998     

Screening of monoclonal antibodies to scarce and labile enzymes: A functional immunoassay for isolating monoclonal antibodies to NADPH:nitrate reductase

A functional immunoassay, that has proved very useful, is described for screening and identifying monoclonal antibodies (McAbs) against scarce and labile enzymes. This method does not require purified enzyme or antigen and it has been successfully applied to isolate three hybridomas secreting McAbs to NADPH:nitrate reductase from the chloronema cells of the mossFunaria hygrometrica. Briefly, the protocol involves: adsorption of murine antibodies from hybridoma supernatants by rabbit antimouse IgG antibody pre-adsorbed toStaphylococcus aureus cells (SAC), reaction with crude extract for 15 min, sedimentation of the SAC complex by centrifugation and measurement of residual enzymatic activity in the supernatant. A depletion indicates the presence of antibodies that bind to the active enzyme. The method is rapid, sensitive and versatile enough to be used to isolate McAbs with exquisite specificities. The three isolated McAbs recognized nitrate reductase protein in a conformation-independent and/or a conformation-dependent manner.     

Formation and activities of xylanhydrolysing enzymes of Humicola grisea var thermoidea

A thermophilic fungus, Humicola grisea var thermoidea, produced in liquid culture two endoxylanases (1,4--d-xylan-xylanohydrolase, EC 3.2.1.8) with M _r of 95 (Xyl I) and 13 (Xyl II) kDa. PAGE of the crude culture filtrate and of each fraction obtained by gel filtration produced three and one band, respectively. Cross-reaction of the culture filtrate and each fraction with polyclonal antibodies prepared against Xyl II produced two and one precipitin bands, respectively. Hydrolysis of wheat straw and rice husk xylan was maximal using a combination of Xyl I and Xyl II. The products formed after hydrolysis, xylo-oligosaccharides and traces of xylose, indicated an endotype enzyme action and the co-operative activities of the xylanases.     

Immobilization of milk-clotting proteases

Traditionally, cheese manufacturing is a batch process and current practice is to use a milk-clotting enzyme in a soluble form. Immobilization of proteases for milk coagulation has received renewed interest and potential applications have recently been reported. Use of immobilized proteases would permit renneting of milk as a continuous process. In addition, it should be possible to recover and re-use the enzyme for coagulation of further batches of milk. This review elaborates on the recent developments in the area of immobilized proteases and their application in cheese-making.Paper No. 7334 through the Experiment Station, G.B. Pant University of Agriculture & Technology, Pantnagar-263 145, India.     

A possible explanation for the synergism between mixed T- and B- cell populations in mice adoptively immunized with Ancylostoma caninum

Ancylostoma caninum--mouse model was employed to study the cellular cooperation in the adoptive immune response. The syngeneic recipient mice were intraperitoneally injected once or twice with mixtures of thymus and bone marrow cells from infected (with 500 or 2000 larvae) and uninfected donors. The experimental recipients expelled and/or destroyed the challenge larval burden more readily and at a greater rate than the controls with unsensitized cells. The cooperation between sensitized thymus and bone marrow cells was, thus, found to be exposed in a better manifestation of adoptive immune response than either of these two alone. The cellular elements of delayed hypersensitivity after combining with the antibodies of humoral system could elicit a much better response in these recipients.