High level of orexin A observed in the phenylketonuria mouse brain is due to the abnormal expression of prepro-orexin

Orexins/hypocretins are recently discovered neuropeptides, synthesized mainly in the lateral hypothalamus of the brain. Orexins regulate various functions including sleep and apetite. We recently reported increased amount of orexin A in the phenylketonuria (PKU) mouse brain. Whether this is caused by overexpression of the precursor for orexins, prepro-orexin was studied in the PKU mouse brain. Microarray expression analysis revealed overexpression of orexin gene in the brain of PKU mouse. Quantitative real-time RT-PCR showed increased level of prepro-orexin mRNA in the PKU mouse brain. In addition, expression of genes associated with cell signal and growth regulation was also affected in the PKU mouse brain, as observed by microarray analysis. These data suggest that up-regulation of orexin mRNA expression is the possible factor for inducing high orexin A in the brain of PKU mouse. The metabolic environment in the brain of PKU mouse affects normal expression of other genes possibly to result in pathophysiology seen in the PKU mouse, if documented also in patients with PKU.     

Unfolding of a Small Protein Proceeds via Dry and Wet Globules and a Solvated Transition State

Dissecting a protein unfolding process into individual steps can provide valuable information on the forces that maintain the integrity of the folded structure. Solvation of the protein core determines stability, but it is not clear when such solvation occurs during unfolding. In this study, far-UV circular dichroism measurements suggest a simplistic two-state view of the unfolding of barstar, but the use of multiple other probes brings out the complexity of the unfolding reaction. Near-UV circular dichroism measurements show that unfolding commences with the loosening of tertiary interactions in a native-like intermediate, N^∗. Fluorescence resonance energy transfer measurements show that N^∗ then expands rapidly but partially to form an early unfolding intermediate I_E. Fluorescence spectral measurements indicate that both N^∗ and I_E have retained native-like solvent accessibility of the core, suggesting that they are dry molten globules. Dynamic quenching measurements at the single tryptophan buried in the core suggest that the core becomes solvated only later in a late wet molten globule, I_L, which precedes the unfolded form. Fluorescence anisotropy decay measurements show that tight packing around the core tryptophan is lost when I_L forms. Of importance, the slowest step is unfolding of the wet molten globule and involves a solvated transition state.     

Identification of B- and T-cell epitopes on HtrA protein of Orientia tsutsugamushi

Orientia tsutsugamushi, a cause of scrub typhus is emerging as an important pathogen in several parts of the tropics. The control of this infection relies on rapid diagnosis, specific treatment, and prevention through vector control. Development of a vaccine for human use would be very important as a public health measure. Antibody and T-cell response have been found to be important in the protection against scrub typhus. This study was undertaken to predict the peptide vaccine that elicits both B- and T-cell immunity. The outer-membrane protein, 47-kDa high-temperature requirement A was used as the target protein for the identification of protective antigen(s). Using BepiPred2 program, the potential B-cell epitope PNSSWGRYGLKMGLR with high conservation among O. tsutsugamushi and the maximum surface exposed residues was identified. Using IEDB, NetMHCpan, and NetCTL programs, T-cell epitopes MLNELTPEL and VTNGIISSK were identified. These peptides were found to have promiscuous class-I major histocompatibility complex (MHC) binding affinity to MHC supertypes and high proteasomal cleavage, transporter associated with antigen processing prediction, and antigenicity scores. In the I-TASSER generated model, the C-score was −0.69 and the estimated TM-score was 0.63 ± 0.14. The location of the epitope in the 3D model was external. Therefore, an antibody to this outer-membrane protein epitope could opsonize the bacterium for clearance by the reticuloendothelial system. The T-cell epitopes would generate T-helper function. The B-cell epitope(s) identified could be evaluated as antigen(s) in immunodiagnostic assays. This cocktail of three peptides would elicit both B- and T-cell immune response with a suitable adjuvant and serve as a vaccine candidate.     

Reproductive characteristics of female Bengal tigers,in Ranthambhore Tiger Reserve,India

Reproductive characteristics of tigers (Panthera tigris) are important to understand population viability. We studied the reproductive parameters of female Bengal tigers (P. t. tigris) in a dry, tropical, deciduous habitat in Ranthambhore Tiger Reserve (RTR), western India, from April 2005 to March 2010. We monitored tigers by direct observation and with cameras placed throughout their habitat. The potential breeding population included 13 adult females. The average age at first reproduction was 3.3 years; 34 cubs were born during the study period (6.2?±?0.82 per year). Sixty-six percent of the births occurred between October and December. Mean litter size was 2.26?±?0.52 (n?=?13, range?=?1–3). The sex ratio of 32 cubs was 1.29 M:1.00 F. The survival rate of cubs (<12 months) was 85 % (95 % CI?=?0.68–0.94), whereas that of juveniles (12–24 months), and subadults (24–36 months) was 79 % (95 % CI?=?0.61–0.91). All breeding females were >3 years old. Only 2 of the 13 females reproduced twice during the 5 years of the study. The birth interval was 33.4?±?3.7 months (range 24–65 months). The mean reproductive rate was 0.59?±?0.23 cubs/female/year. Our study indicates that tiger populations can grow rapidly if the habitat provides adequate protection, an adequate population of prey, and minimal to no poaching.     

Pathogenic and Genetic Variation among the Isolates of Rhizoctonia solani (AG 1‐IA), the Rice Sheath Blight Pathogen

Sheath blight disease of rice caused by Rhizoctonia solani is one of the most dreaded plant diseases faced by the rice farmers all over the world. None of the commercially cultivated rice varieties have sufficient level of field resistance, and the disease is presently being managed by chemical pesticides. In this study, 40 isolates of rice sheath blight pathogen, collected from diverse rice ecosystems from 12 different states of India, were characterized for their morphological, pathological and genetic variation. The isolates showed wide morphological variation in terms of size of sclerotia and abundance of sclerotia production. The virulence of each pathogen isolate was studied on four rice varieties, that is TN1, IR 64, Tetep and Swarnadhan in glasshouse, and observations were taken by measuring the relative lesion height. The relative lesion heights produced by these isolates on four different rice varieties varied widely. Genetic variation of the isolates was analysed using ISSR markers. The primers based on AG, GA, AC and CA repeats were informative and revealed polymorphism among the isolates. The polymorphism information content (PIC) of the primers ranged from 0.80 to 0.96, while the resolving power (Rp) ranged from 3.7 to 15.35. Largely, grouping of the isolates happened based on their geographical origin. One isolate from Titabar, Assam, and another from Adialabad, Telangana, were quite distinct from rest of the isolates.     

Purification and Partial Characterization of Arginine Decarboxylase from Brassica campestris

Arginine decarboxylase (EC 4.1.1.19) has been purified and characterized from Brassica campestris cv B-9. The enzyme was purified 1120 fold and the recovery was 9%. The mol wt of the enzyme determined by gel filtration was 240 kD with identical subunits of 60 kD. The pH and temperature optima for the enzyme were 8.0 and 30°C respectively. The Km was 0.31mM. Polyamines inhibited the enzyme activity significantly. Immunodiffusion with ADC-specific antibodies showed cross reactivity against purified ADC from Brassica.