Calcium-dependant binding proteins associated with human placental syncytiotrophoblast microvillous cytoskeleton

Isolated human placental syncytiotrophoblast microvillous plasma membrane vesicles were extracted with Triton X-100 to yield a detergent-insoluble residue. The residue contained approx. 50% of the total membrane protein and was qualitatively different from untreated trophoblast on SDS-polyacrylamide gel electrophoresis, Western blots and dot-immunobinding assay. Three major proteins, with molecular weights of 68, 36 and 34 kDa, dissociated from this non-ionic detergent-insoluble submembranous cytoskeletal fraction in the presence of calcium chelators. They were immunologically related to human lymphocyte cytoskeletal calcium-binding proteins, and the 36 kDa component reacted with antisera to the phospholipase A2 inhibitor, lipocortin II. Anti-lipocortin I sera did not recognise the 34 kDa protein, but did react with a series of trophoblast cytoskeletal proteins in the 34-37 kDa region. Incubation of epidermal growth factor with isolated trophoblast membrane vesicles stimulated the phosphorylation of a 36 kDa protein on tyrosine residues. Immunoprecipitation studies further showed there was no phosphorylation of the 34 kDa protein, but the 68 kDa protein was a major phosphorylated component of isolated syncytiotrophoblast membranes. p68 was principally phosphorylated on serine with slight tyrosine phosphorylation which showed an apparent increase after epidermal growth factor treatment. These results indicate a family of calcium-dependant binding proteins, some of which are phosphorylated, associated with the submembranous cytoskeleton of syncytiotrophoblast microvilli.     

Biocalorimetric and respirometric studies on metabolic activity of aerobically grown batch culture of<Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis>

Biocalorimetry has proved to be a useful tool for scale up and control of bioreactors. The findings reported here are fundamental data required for scale up and control of a reactor for the treatment of saline tannery wastewater. The study deals with biokinetics of a halo-tolerant bacteriaPseudomonas aeruginosa isolated from tannery saline wastewater (soak liquor). Batch experiments were performed in a biocalorimeter and the isolated strain was grown in a glucose-limited mineral salt medium (MSM) at optimized growth conditions. Tessier model is found to fit well for the growth ofP. aeruginosa in biocalorimeter. Biokinetic constants are evaluated and simulation is done to validate experimental results with theoretical values. Respirogram and heat profiles are seen to follow the biomass growth curve. Oxycalorific coefficient is validated with the theoretical values and those noticed in the published literature. There is a good correlation between experimentally determined heat yields and the theoretical values predicted by elemental and enthalpy balances. The heat yield and biomass yield values indicated the behavior of the isolated organism in a substrate-limited well defined growth media (MSM)     

Biological treatment of tannery wastewater by using salt-tolerant bacterial strains

Background  

High salinity (1–10% w/v) of tannery wastewater makes it difficult to be treated by conventional biological treatment. Salt tolerant microbes can adapt to these saline conditions and degrade the organics in saline wastewater.     

Homepeptide Repeats:Implications for Protein Structure,Function and Evolution

Analysis of protein sequences from Mycobacterium tuberculosis H37Rv(Mtb H37Rv) was performed to identify homopeptide repeatcontaining proteins(HRCPs).Functional annotation of the HRCPs showed that they are preferentially involved in cellular metabolism.Furthermore,these homopeptide repeats might play some specific roles in protein-protein interaction.Repeat length differences among Bacteria,Archaea and Eukaryotes were calculated in order to identify the conservation of the repeats in these divergent kingdoms.From the results,it was evident that these repeats have a higher degree of conservation in Bacteria and Archaea than in Eukaryotes.In addition,there seems to be a direct correlation between the repeat length difference and the degree of divergence between the species.Our study supports the hypothesis that the presence of homopeptide repeats influences the rate of evolution of the protein sequences in which they are embedded.Thus,homopeptide repeat may have structural,functional and evolutionary implications on proteins.     

Structural analysis of NADPH depleted bovine liver catalase and its inhibitor complexes

To study the functional role of NADPH during mammalian catalase inhibition, the X-ray crystal structures of NADPH-depleted bovine liver catalase and its inhibitor complexes, cyanide and azide, determined at 2.8Å resolution. From the complex structures it is observed that subunits with and without an inhibitor/catalytic water molecule are linked by N-terminal domain swapping. Comparing mammalian- and fungal- catalases, we speculate that NADPH-depleted mammalian catalases may function as a domain-swapped dimer of dimers, especially during inactivation by inhibitors like cyanide and azide. We further speculate that in mammalian catalases the N-terminal hinge-loop region and α-helix is the structural element that senses NADPH binding. Although the above arguments are speculative and need further verification, as a whole our studies have opened up a new possibility, viz. that mammalian catalase acts as a domain-swapped dimer of dimers, especially during inhibitor binding. To generalize this concept to the formation of the inactive state in mammalian catalases in the absence of tightly bound NADPH molecules needs further exploration. The present study adds one more intriguing fact to the existing mysteries of mammalian catalases.     

Media replenishment: A tool for the analysis of high-cell density perfusion systems

A simple method to analyze media consumption and by-product build-up in high-cell-density perfusion cultures has been developed. This technique makes use of media replenishment strategy which can be used to decouple the two phenomena. Also, the replenished media method can be used to identify limitation by complex and undefined media components such as bovine serum.     

Anin vitro test using cholesterol metabolism of macrophages to determine drug sensitivity and resistance ofMycobacterium leprae

Macrophages that have ingested liveMycobacterium leprae show a preferential accumulation of cholesterol ester. Such an accumulation is not seen, on the ingestion of dead bacteria. Among the macrophages that ingest liveMycobacterium leprae, the presence of dapsone or rifampicin prevents largely the alteration in the anticipated increase in the cholesterol ester indicating the sensitivity of the bacteria to the drug. In the small number of relapsed patients, the presence of dapsone did not reduce the cholesterol ester increase, suggesting that theMycobacterium leprae present are either resistant or escaped detection. The method provides a rapid drug screening system foranti-Mycobacterium leprae activity of known and unknown compounds     

Importance of determining viability ofMycobacterium leprae inside macrophages—anin vitro method using uracil

It has been demonstrated thatMycobacterium leprae, are caPable of taking uP uracil and incorPorating it into trichloroacetic acid-insoluble materials, both as free susPension of bacteria, as well as when they are inside the macrophages, a host cell for theirin vivo survival. Same amount of bacteria show better incorPoration inside macroPhages than as free bacterial susPension. Both tyPes of incorPoration are inhibited by rifamPicin an antileProsy drug and an RNA synthesis inhibitor. Thus uracil uPtake byMycobacterium lePrae inside macroPhages has been used for standardising a raPidin vitro viability assay for the leProsy causing bacteria.