Effect of Solvents and Surfactants on the Catalytic Activity of Trigonopsis Variabilis

D-Amino acid oxidase present in the cells of Trigonopsis variabilis became accessible to cephalosporin C, its substrate, when treated with either butyl acetate or Triton-X-100. The enzyme was not leached out enabling use of these cells as a biocatalyst.     

Isoforms of Linamarase in Cassava (Manihot esculenta)

Linamarase (EC. 3.2.1.21) was purified from different tissues of cassava (leaf, rind and tuber) to compare the kinetic properties and characteristics of the enzyme in these tissues. Purified enzyme preparation appeared as single band of average molecular size 70 kD in SDS-PAGE gels. The kinetic properties of linamarase with respect to pH and temperature indicated that tuber linamarase possessed a broader pH optimum and higher temperature stability as compared to leaf and rind enzymes. Differences in Km values for linamarin were observed with leaf linamarase having the highest Km value (500 μM) followed by rind (400 μM) and then tuber (250 μM) linamarases. Rind enzyme appeared to be less susceptible to urea denaturation than the leaf enzyme. Comparison of elution profiles from DEAE-Cellulose indicated that the relative amounts of the various ionic forms of the enzyme differed in the case of each tissue. Elution patterns of the enzyme from Con A-Sepharose also differed, suggesting difference in glycosylation among leaf, rind and tuber enzymes. This was confirmed by carbohydrate analysis which showed that the tuber linamarase contained significantly higher amount of protein bound carbohydrate. These results suggest the possible occurrence of different forms of linamarase in cassava.     

Initiation of altered heparan sulphate on beta-D-xyloside in rat hepatocytes

Inhibition of protein synthesis by cycloheximide 10(-3)M reduced the incorporation of [35S]sulphate into heparan sulphate to about 5% of untreated hepatocytes. Addition of rho-nitrophenyl beta-D-xyloside could partially revert this inhibitory effect. The sulphated material isolated from the cell layer or secretions of hepatocytes grown in presence of cycloheximide and rho-nitrophenyl beta-D-xyloside were shown to be mostly free heparan sulphate chains not bound to core protein. Covalent association of beta-xylosides to the heparan sulphates was demonstrated for heparan sulphate synthetized in the presence of [35S]sulphate, cycloheximide and the fluorogenic 4-methylumbelliferyl beta-D-xyloside. Beta-Xylosides served as an initiator of heparan sulphate chain synthesis in rat hepatocytes only in the absence of protein synthesis. Heparan sulphates primed on artificial beta-xylosides are slightly smaller in molecular size and are more sulphated than chains linked to core protein.     

Negative modulation of eNOS by laminin involving post‐translational phosphorylation

Nitric oxide (NO) regulates the vascular tone, and influences survival and apoptosis of endothelial cells (ECs). NO is produced by nitric oxide synthase (NOS) and eNOS is the constitutive enzyme in the endothelium. Though the extracellular matrix (ECM) has been reported to regulate various EC functions, the role of ECM in the regulation of eNOS is not clear. The present study was designed to analyze if laminin‐1 (Ln‐1), the major glycoprotein of the basement membrane, can regulate eNOS. The activity of eNOS was significantly low in ECs maintained on Ln‐1 as compared to those on Col I and polylysine. Reversal of the effect of Ln‐1 on treatment with inhibitor of p38 MAPK and changes in Thr and Ser phosphorylation in purified eNOS suggested that eNOS activity in cells maintained on Ln‐1 is negatively regulated by post‐translational phosphorylation at Ser and Thr residues by recruiting p38 MAPK pathway. Increase in eNOS activity and induction of apoptosis upon inhibition of p38 MAPK and reversal of this on inhibition of NOS by L ‐NAME suggested that increased NO induced apoptosis in ECs maintained on Ln‐1 when p38 MAPK was inhibited. These results suggest that Ln contributes to survival of ECs by negatively modulating eNOS in a p38 MAPK dependent pathway. J. Cell. Physiol. 219: 123–131, 2009. © 2008 Wiley‐Liss, Inc.     

Biofuels: opportunities and challenges in India

Energy plays a vital role in the economic growth of any country. Current energy supplies in the world are unsustainable from environmental, economic, and societal standpoints. All over the world, governments have initiated the use of alternative sources of energy for ensuring energy security, generating employment, and mitigating CO₂ emissions. Biofuels have emerged as an ideal choice to meet these requirements. Huge investments in research and subsidies for production are the rule in most of the developed countries. India started its biofuel initiative in 2003. This initiative differs from other nations’ in its choice of raw material for biofuel production—molasses for bioethanol and nonedible oil for biodiesel. Cyclicality of sugar, molasses, and ethanol production resulted in a fuel ethanol program which suffered from inconsistent production and supply. The restrictive policies, availability of molasses, and cost hampered the fuel ethanol program. Inconsistent policies, availability of land, choice of nonnative crops, yield, and market price have been major impediments for biodiesel implementation. However, a coherent, consistent, and committed policy with long-term vision can sustain India’s biofuel effort. This will provide energy security, economic growth, and prosperity and ensure a higher quality of life for India.     

A novel gene of tumor necrosis factor ligand superfamily from kuruma shrimp,Marsupenaeus japonicus

A tumor necrosis factor (TNF) gene has been isolated and characterized in kuruma shrimp, Marsupenaeus japonicus, providing the first conclusive evidence for the existence of the TNF ligand in shrimp. The kuruma shrimp TNF (MjTNF) cDNA was composed of 1868 bp with a 262 bp 5′-untranslated region (UTR) and a 220 bp 3′-UTR, which was translated into a protein of 462 amino acid residues that included a predicted transmembrane domain of 23 amino acid residues (Trp20–Val42) and the TNF family signature (Pro321–Leu448). Homology analysis of MjTNF showed 30.7% and 26.7% identities with fruit fly (Drosophila melanogaster) Eiger and human (Homo sapiens) ectodysplasin A, respectively. The MjTNF gene was constitutively expressed in unstimulated organs of shrimp such as the muscle, stomach, brain and gill. In lymphoid organ cells, an enhanced expression of the MjTNF gene was observed following stimulation with peptidoglycan and polycytidylic acid. A high expression level of MjTNF was observed in vivo 2 h and 4 h after stimulation with lipopolysaccharide and Vibrio penaeicida, respectively. These observations suggest that MjTNF plays a role in the innate immune defense in kuruma shrimp. The discovery of shrimp TNF will allow a more complete and concrete understanding of shrimp inflammatory responses.     

Modulation of neutral matrix metalloproteinases of involuting rat mammary gland by different cations and glycosaminoglycans

The synthesis and regulation of the matrix metalloproteinases (MMPs) are important factors contributing to the involution of mammary gland. In order to understand the role of these MMPs in involution and in remodeling of the mammary gland, the different MMPs (130K, 68K, and 60K gelatinases) were partially purified by gel filtration and affinity chromatography over gelatin Sepharose and subjected to kinetic analysis. Comparative analysis of the different gelatinases showed that the 130K that appears at the early involuntary phase and the constitutive 68K enzyme are more specific for Col IV of the basement membrane, while the inducible 60K that appeared at the later phase of involution degraded Col I more efficiently. These neutral proteinases required Ca2+/Zn2+ for their activity and the analysis of cation dependence revealed that Ca2+ at 10 mM concentration and above completely inhibited the enzyme. The 60K was active at very low concentration of Zn2+ (5 microM); but at higher concentration of Zn2+ (2 mM), where the 68K and 130K were active, the 60K gelatinase was inhibited, indicating a difference in the cation dependence of these enzymes. Chondroitin sulfate A and chondroitin sulfate C caused inhibition of the 130K, 68K, and 60K, while hyaluronic acid and heparin did not show any effect, suggesting that the chondroitin sulfate proteoglycan that decorates collagen in the ECM can modu late the activity of the collagenases in vivo. These results suggest that the 130K gelatinase expressed during the early phase of involution degraded Col IV of the basement membrane, making the 60K gelatinase formed at a later stage of involution more accessible to its preferred substrate (Col I of the underlying stroma), highlighting the role of these MMPs in mammary gland involution.     

Soil Enzyme Activities and Their Relationship with Soil Physico-Chemical Properties and Oxide Minerals in Coastal Agroecosystem of Puducherry

The objectives of our research were to assess the soil enzyme activities in relation with soil physicochemical and oxide minerals in the coastal agroecosystem of Puducherry region, India. Soils from nine farms in organic (ORG), sustainable (SUS), and conventional (CON) farming were sampled. Organically managed farming system soils contain significantly higher amounts of soil total N, organic carbon, and a higher level of microbial biomass C and N. Urease, protease, β-glucosidase, cellulose, saccharase, xylanase, and alkaline phosphatase enzyme activities were higher in organic farming system soils compared to sustainable and conventional farming soils. In addition, silt, clay, Al₂O₃, CaO, Fe₂O₃, K₂O, MgO, MnO, Na₂O, and P₂O₅ oxides were higher in organic farming soil and they showed a significant positive correlation with soil enzyme activities. Our study revealed that soil enzyme activities and soil minerals were significantly affected by farm management practices. The organic farming system had improved the soil health, enzyme activities, and plant available nutrients in coastal agro-ecosystem.