A Peptide Bond from the Inter-lobe Segment in the Bilobal Lactoferrin Acts as a Preferred Site for Cleavage for Serine Proteases to Generate the Perfect C-lobe: Structure of the Pepsin Hydrolyzed Lactoferrin C-lobe at 2.28 Å Resolution

The Protein Journal - C-lobe represents the C-terminal half of lactoferrin which is a bilobal 80 kDa iron binding glycoprotein. The two lobes are designated as N-lobe (Ser1-Glu333) and...     

PMTER-ABE: a practical multi-authority CP-ABE with traceability,revocation and outsourcing decryption for secure access control in cloud systems

Cluster Computing - Attribute-based encryption (ABE) has evolved as an efficient and secure method for storage of data with fine-grained access control in cloud platforms. In recent years,...     

Targeted comparative RNA interference analysis reveals differential requirement of genes essential for cell proliferation

Differences in the genetic and epigenetic make up of cell lines have been very useful for dissecting the roles of specific genes in the biology of a cell. Targeted comparative RNAi (TARCOR) analysis uses high throughput RNA interference (RNAi) against a targeted gene set and rigorous quantitation of the phenotype to identify genes with a differential requirement for proliferation between cell lines of different genetic backgrounds. To demonstrate the utility of such an analysis, we examined 257 growth-regulated genes in parallel in a breast epithelial cell line, MCF10A, and a prostate cancer cell line, PC3. Depletion of an unexpectedly high number of genes (25%) differentially affected proliferation of the two cell lines. Knockdown of many genes that spare PC3 (p53-) but inhibit MCF10A (p53+) proliferation induces p53 in MCF10A cells. EBNA1BP2, involved in ribosome biogenesis, is an example of such a gene, with its depletion arresting MCF10A at G1/S in a p53-dependent manner. TARCOR is thus useful for identifying cell type-specific genes and pathways involved in proliferation and also for exploring the heterogeneity of cell lines. In particular, our data emphasize the importance of considering the genetic status, when performing siRNA screens in mammalian cells.     

Using defined finger–finger interfaces as units of assembly for constructing zinc-finger nucleases

Zinc-finger nucleases (ZFNs) have been used for genome engineering in a wide variety of organisms; however, it remains challenging to design effective ZFNs for many genomic sequences using publicly available zinc-finger modules. This limitation is in part because of potential finger–finger incompatibility generated on assembly of modules into zinc-finger arrays (ZFAs). Herein, we describe the validation of a new set of two-finger modules that can be used for building ZFAs via conventional assembly methods or a new strategy—finger stitching—that increases the diversity of genomic sequences targetable by ZFNs. Instead of assembling ZFAs based on units of the zinc-finger structural domain, our finger stitching method uses units that span the finger–finger interface to ensure compatibility of neighbouring recognition helices. We tested this approach by generating and characterizing eight ZFAs, and we found their DNA-binding specificities reflected the specificities of the component modules used in their construction. Four pairs of ZFNs incorporating these ZFAs generated targeted lesions in vivo, demonstrating that stitching yields ZFAs with robust recognition properties.     

Effect of UV irradiation on aflatoxin reduction: a cytotoxicity evaluation study using human hepatoma cell line

In this proof-of-concept study, the efficacy of a medium-pressure UV (MPUV) lamp source to reduce the concentrations of aflatoxin B₁, aflatoxin B₂, and aflatoxin G₁ (AFB_1, AFB₂, and AFG₁) in pure water is investigated. Irradiation experiments were conducted using a collimated beam system operating between 200 to 360 nm. The optical absorbance of the solution and the irradiance of the lamp are considered in calculating the average fluence rate. Based on these factors, the UV dose was quantified as a product of average fluence rate and treatment time. Known concentrations of aflatoxins were spiked in water and irradiated at UV doses ranging from 0, 1.22, 2.44, 3.66, and 4.88 J cm^?2. The concentration of aflatoxins was determined by HPLC with fluorescence detection. LC-MS/MS product ion scans were used to identify and semi-quantify degraded products of AFB₁, AFB₂, and AFG₁. It was observed that UV irradiation significantly reduced aflatoxins in pure water (p < 0.05). Irradiation doses of 4.88 J cm^?2 reduced concentrations 67.22% for AFG₁, 29.77% for AFB₂, and 98.25% for AFB₁ (p < 0.05). Using this technique, an overall reduction of total aflatoxin content of ≈95% (p < 0.05) was achieved. We hypothesize that the formation of ˙OH radicals initiated by UV light may have caused photolysis of AFB₁, AFB₂, and AFG₁ molecules. In cell culture studies, our results demonstrated that the increase of UV dosage decreased the aflatoxin-induced cytotoxicity in HepG₂ cells. Therefore, our research finding suggests that UV irradiation can be used as an effective technique for the reduction of aflatoxins.     

Isolation and characterization of biosurfactant producing <Emphasis Type="Italic">Bacillus</Emphasis> sp. from diesel fuel-contaminated site

Among hydrocarbon pollutants, diesel oil is a complex mixture of alkanes and aromatic compounds which are often encountered as soil contaminants leaking from storage tanks and pipelines or as result of accidental spillage. One of the best ecofriendly approaches is to restore contaminated soil by using microorganisms able to degrade those toxic compounds in a bioremediation process. In the present study, nineteen bacteria were isolated by enrichment culture technique from diesel spilled soil collected from electric generator shed of NBAIM, Mau. All the isolates were subjected to screening for lipase production and twelve isolates were found to be positive for lipase. When the isolates were screened for biosurfactant production using CTAB-methylene blue agar plates, only one isolate viz. 2NBDSH3 was found positive which was found to be phylogenetically closely related with Bacillus flexus. Despite having low emulsification index, the bacterium could degrade 88.6% of diesel oil in soil. Biosurfactant from the isolate was extracted and characterized through infra-red spectroscopy which indicated its possible lipopeptide nature which was further supported by strong absorption in UV range in the UV-Vis spectrum. The results of the present study indicated that the isolate either does not produce any bioemulsifier or produces very low amount of emulsifier rather it produces a lipopeptide biosurfactant which helps in degradation of diesel oil by lowering the surface tension. The bacterium thus isolated and characterized can serve as a promising solution for ecofriendly remediation of bacterium diesel contaminated soils.     

Clinical proteomics of the neglected human malarial parasite Plasmodium vivax

Recent reports highlight the severity and the morbidity of disease caused by the long neglected malaria parasite Plasmodium vivax. Due to inherent difficulties in the laboratory-propagation of P. vivax, the biology of this parasite has not been adequately explored. While the proteome of P. falciparum, the causative agent of cerebral malaria, has been extensively explored from several sources, there is limited information on the proteome of P. vivax. We have, for the first time, examined the proteome of P. vivax isolated directly from patients without adaptation to laboratory conditions. We have identified 153 proteins from clinical P. vivax, majority of which do not show homology to any previously known gene products. We also report 29 new proteins that were found to be expressed in P. vivax for the first time. In addition, several proteins previously implicated as anti-malarial targets, were also found in our analysis. Most importantly, we found several unique proteins expressed by P. vivax.This study is an important step in providing insight into physiology of the parasite under clinical settings.     

Enhanced Expression of Rabies Virus Surface G-Protein in Escherichia coli using SUMO Fusion

Fusion systems are known to increase the expression of difficult to express recombinant proteins in soluble form to facilitate their purification. Rabies glycoprotein was also tough to express at sufficient level in soluble form in both E. coli and plant. The present work was aimed to over-express and purify this membrane protein from soluble extract of E. coli. Fusion of Small Ubiqutin like Modifier (SUMO) with rabies glycoprotein increased ~1.5 fold higher expression and ~3.0 fold solubility in comparison to non-fused in E. coli. The SUMO fusion also simplified the purification process. Previously engineered rabies glycoprotein gene in tobacco plants provides complete protection to mice, but the expression was very low for purification. Our finding demonstrated that the SUMO-fusion was useful for enhancing expression and solubility of the membrane protein and again proves to be a good alternative technology for applications in biomedical and pharmaceutical research.