Cloning, characterization and expression of the chitinase gene of Enterobacter sp. NRG4.

A chitinase producing bacterium Enterobacter sp. NRG4, previously isolated in our laboratory, has been reported to have a wide range of applications such as anti-fungal activity, generation of fungal protoplasts and production of chitobiose and N-acetyl D-glucosamine from swollen chitin. In this paper, the gene coding for Enterobacter chitinase has been cloned and expressed in Escherichia coli BL21(DE3). The structural portion of the chitinase gene comprised of 1686 bp. The deduced amino acid sequence of chitinase has high degree of homology (99.0%) with chitinase from Serratia marcescens. The recombinant chitinase was purified to near homogeneity using His-Tag affinity chromatography. The purified recombinant chitinase had a specific activity of 2041.6 U mg(-1). It exhibited similar properties pH and temperature optima of 5.5 and 45°C respectively as that of native chitinase. Using swollen chitin as a substrate, the K(m), k(cat) and catalytic efficiency (k(cat)/K(m)) values of recombinant chitinase were found to be 1.27 mg ml(-1), 0.69 s(-1) and 0.54 s(-1)M(-1) respectively. Like native chitinase, the recombinant chitinase produced medicinally important N-acetyl D-glucosamine and chitobiose from swollen chitin and also inhibited the growth of many fungi.

Synergistic antimicrobial activity of tea & antibiotics.

Tea leaves are known for its antibacterial activity against many microorganisms. In this study we attempted to describe the synergistic antimicrobial activity of tea and antibiotics against enteropathogens. Antimicrobial activity of boiled water tea extract and organic solvent extract were studied against Salmonella typhimurium 1402/84, S. typhi, S. typhi Ty2a, Shigella dysenteriae, Yersinia enterocolitica C770, and Escherichia coli (EPEC P2 1265) determining minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC) and death rate kinetics at MBC of tea extract in presence of subinhibitory concentration of antibiotic. Both green tea or black tea extracts effectively inhibited the growth of S. typhimurium 1402/84, S. typhi, S. typhi Ty2a, S. dysenteriae, Y. enterocolitica C770, and E.coli (EPEC P2 1265). However, the growth inhibitory concentration of tea extract was lower for green tea as compared to black tea extract. Antimicrobial activity of green tea tea methanol: water extract tea was better as compared to boiled water tea extract of green tea. Based on death rate kinetics results, S.typhi Ty2a appeared to be highly sensitive and Y. enterocolitica C770 the most resistant. Chloramphenicol and tea extract in combination inhibited the growth of S.dysenteriae at 2.5 microg/ml chloramphenicol (MIC 5 microg/ml) and 5.094 mg/ml black tea extract (MIC 9.089 mg/ml). Tea extract showed synergistic activity with chloramphenicol and other antibiotics like gentamycin, methicillin and nalidixic acid against test strains.

Microbial alkaline pectinases and their industrial applications: a review.

The biotechnological potential of pectinolytic enzymes from microorganisms has drawn a great deal of attention from various researchers worldwide as likely biological catalysts in a variety of industrial processes. Alkaline pectinases are among the most important industrial enzymes and are of great significance in the current biotechnological arena with wide-ranging applications in textile processing, degumming of plant bast fibers, treatment of pectic wastewaters, paper making, and coffee and tea fermentations. The present review features the potential applications and uses of microbial alkaline pectinases, the nature of pectin, and the vast range of pectinolytic enzymes that function to mineralize pectic substances present in the environment. It also emphasizes the environmentally friendly applications of microbial alkaline pectinases thereby revealing their underestimated potential. The review intends to explore the potential of these enzymes and to encourage new alkaline pectinase-based industrial technology.

Microbial xylanases and their industrial applications: a review.

Despite an increased knowledge of microbial xylanolytic systems in the past few years, further studies are required to achieve a complete understanding of the mechanism of xylan degradation by microorganisms and their enzymes. The enzyme system used by microbes for the metabolism of xylan is the most important tool for investigating the use of the second most abundant polysaccharide (xylan) in nature. Recent studies on microbial xylanolytic systems have generally focussed on induction of enzyme production under different conditions, purification, characterization, molecular cloning and expression, and use of enzyme predominantly for pulp bleaching. Rationale approaches to achieve these goals require a detailed knowledge of the regulatory mechanism governing enzyme production. This review will focus on complex xylan structure and the microbial enzyme complex involved in its complete breakdown, studies on xylanase regulation and production and their potential industrial applications, with special reference to biobleaching.

Properties of a thermostable and solvent stable extracellular lipase from a Pseudomonas sp. AG-8.

An extracellular lipase isolated from Pseudomonas sp. AG-8, had an optimal activity at 45 degrees C and pH 8.0-8.5. It retained more than 80% of its initial activity after keeping for 1 h at 65 degrees C. The enzyme was stable in 5 M NaCl and 6 M urea. Triton X-100 increased the lipase activity by 2.4 fold. Ca2+ ions activated the enzyme, while Zn2+, Fe2+, Fe3+ strongly inhibited its activity. Ethanol, methanol and acetone at 20% (v/v) enhanced the lipase activity by 2.9, 3.6 and 4.5 fold respectively. Dimethylsulphoxide at 90% (v/v) enhanced the enzyme activity up to 5.7 fold.

Isolation, purification and characterization of xylanasefrom Staphylococcus sp. SG-13 and its application in biobleaching of kraft pulp.

A haloalkalophilic Staphylococcus sp. SG-13 produced an alkalistable xylanase in wheat bran medium. A 12-fold purification was achieved by using standard purification techniques. The purified xylanase exhibited a dual pH optima of 7.5 and 9.2. The optimum temperature for enzyme activity was 50 degrees C. The enzyme was stable at 50 degrees C for more than 4 h. The xylanase exhibited Km and Vmax values of 4 mg ml-1, 90 micromol min-1 per mg for birchwood xylan and 7 mg ml-1, 55 micromol min-1 per mg for oatspelt xylan, respectively. The substrate binding affinity of xylanase was more for oatspelt xylan but birchwood xylan was hydrolysed more rapidly. The xylanase activity was stimulated by Fe2+, Ni2+, Cu2+ and dithiothreitol up to 60% and was strongly inhibited in the presence of Co2+, Hg2+, Pb2+, phenyl methane sulphonyl fluoride, ethylenediaminetetraacetic acid, and acetic anhydride up to 100%. The xylanase dose of 1.8 U g-1 moisture free pulp, exhibited bleach boosting of kraft pulps optimally at pH 9.5-10.0 and 50 degrees C after 4 h of reaction time. Pretreatment of pulp with xylanase and its subsequent treatment with 8% hypochlorite, reduced the kappa number by 30%, enhanced the brightness and viscosity by 11% and 1.8%, respectively, and improved the paper properties such as tensile strength and burst factor up to 10% and 17%, respectively.