Changes in cyclin and cyclin-dependent protein kinase expression in the long-tailed ground squirrel (<Emphasis Type="Italic">Spermophilus undulatus</Emphasis>) brain during hibernation and awakening

We have studied the expression of cyclins (Cicl A and Cicl B1) and cyclin-dependent protein kinases (Cdk1, Cdk2, Cdk4, and Cdk5) in the brain of the long-tailed ground squirrel (Spermophilus undulatus) during different phases of their yearly cycle of life activities. We found that the expression of protein kinases in the frontal neocortex, hippocampus, and caudal brainstem differed by from three to five times, which indicates the regional specificity of the activity of cell-cycle proteins in the brain of a hibernating animal. During the end of winter hibernation, a significant increase in the expression of Cdk1, Cdk2, and Cdk4 were found in the hippocampus, which is due to the presence of progenitor neural cells in the subgranular region of the dentate gyrus. These cells are able to produce new neurons during all of ontogenesis. Our results show that during winter hibernation and awakening, region-specific changes in the expression of cell cycle proteins occur in the brain of a long-tail ground squirrel, which provides the appropriate activity of the cell cycle during the new functional state of a hibernating animal.     

Ubiquitin Ser65 phosphorylation affects ubiquitin structure,chain assembly and hydrolysis

The protein kinase PINK1 was recently shown to phosphorylate ubiquitin (Ub) on Ser65, and phosphoUb activates the E3 ligase Parkin allosterically. Here, we show that PINK1 can phosphorylate every Ub in Ub chains. Moreover, Ser65 phosphorylation alters Ub structure, generating two conformations in solution. A crystal structure of the major conformation resembles Ub but has altered surface properties. NMR reveals a second phosphoUb conformation in which β5-strand slippage retracts the C-terminal tail by two residues into the Ub core. We further show that phosphoUb has no effect on E1-mediated E2 charging but can affect discharging of E2 enzymes to form polyUb chains. Notably, UBE2R1- (CDC34), UBE2N/UBE2V1- (UBC13/UEV1A), TRAF6- and HOIP-mediated chain assembly is inhibited by phosphoUb. While Lys63-linked poly-phosphoUb is recognized by the TAB2 NZF Ub binding domain (UBD), 10 out of 12 deubiquitinases (DUBs), including USP8, USP15 and USP30, are impaired in hydrolyzing phosphoUb chains. Hence, Ub phosphorylation has repercussions for ubiquitination and deubiquitination cascades beyond Parkin activation and may provide an independent layer of regulation in the Ub system.     

Optimization of mycophenolic acid production in solid state fermentation using response surface methodology

Mycophenolic acid (MPA) can be produced in solid state fermentation. An isolate of Penicillium brevi-compactum ATCC 16024 grown on moist wheat bran produced a titre of 425 mg per kg of wheat bran. Central composite rotatable design and response surface methodology were employed to derive a statistical model for media optimization towards production of mycophenolic acid. Five levels with a five factorial design were adopted. The correlation coefficient was 0.82, ensuring a satisfactory adjustment of the model to the experimental values. This statistical design was very effective in improving the titre of mycophenolic acid up to 3286 mg per kg of wheat bran. Received 24 July 1998/ Accepted in revised form 4 December 1998     

Spermine Condenses DNA,but Not RNA Duplexes

Interactions between the polyamine spermine and nucleic acids drive important cellular processes. Spermine condenses DNA and some RNAs, such as poly(rA):poly(rU). A large fraction of the spermine present in cells is bound to RNA but apparently does not condense it. Here, we study the effect of spermine binding to short duplex RNA and DNA, and compare our findings with predictions of molecular-dynamics simulations. When small numbers of spermine are introduced, RNA with a designed sequence containing a mixture of 14 GC pairs and 11 AU pairs resists condensation relative to DNA of an equivalent sequence or to 25 bp poly(rA):poly(rU) RNA. A comparison of wide-angle x-ray scattering profiles with simulation results suggests that spermine is sequestered deep within the major groove of mixed-sequence RNA. This prevents condensation by limiting opportunities to bridge to other molecules and stabilizes the RNA by locking it into a particular conformation. In contrast, for DNA, simulations suggest that spermine binds externally to the duplex, offering opportunities for intermolecular interaction. The goal of this study is to explain how RNA can remain soluble and available for interaction with other molecules in the cell despite the presence of spermine at concentrations high enough to precipitate DNA.     

Exploring protein native states and large-scale conformational changes with a modified generalized born model

Implicit solvation models provide, for many applications, a reasonably accurate and computationally effective way to describe the electrostatics of aqueous solvation. Here, a popular analytical Generalized Born (GB) solvation model is modified to improve its accuracy in calculating the solvent polarization part of free energy changes in large-scale conformational transitions, such as protein folding. In contrast to an earlier GB model (implemented in the AMBER-6 program), the improved version does not overstabilize the native structures relative to the finite-difference Poisson-Boltzmann continuum treatment. In addition to improving the energy balance between folded and unfolded conformers, the algorithm (available in the AMBER-7 and NAB molecular modeling packages) is shown to perform well in more than 50 ns of native-state molecular dynamics (MD) simulations of thioredoxin, protein-A, and ubiquitin, as well as in a simulation of Barnase/Barstar complex formation. For thioredoxin, various combinations of input parameters have been explored, such as the underlying gas-phase force fields and the atomic radii. The best performance is achieved with a previously proposed modification to the torsional potential in the Amber ff99 force field, which yields stable native trajectories for all of the tested proteins, with backbone root-mean-square deviations from the native structures being approximately 1.5 A after 6 ns of simulation time. The structure of Barnase/Barstar complex is regenerated, starting from an unbound state, to within 1.9 A relative to the crystal structure of the complex.     

Congenital deformity of the paw in a captive tiger: case report

Background

Although Morbillivirus and Toxoplasma gondii have emerged as important pathogens for several cetaceans populations over the last 20 years, they have never been identified together in a Mysticete. In particular, morbilliviral infection has been never described in the Mediterranean fin whale population.

Case presentation

On January 2011 an adult male of fin whale (Balaenoptera physalus) stranded along the Tyrrhenian coastline of Italy. During necropsy, tissue samples from heart, skeletal muscle, mesenteric lymph nodes, liver, spleen, lung, and kidney were collected and subsequently analyzed for Morbillivirus and Toxoplasma gondii by microscopic and molecular methods. Following the detailed necropsy carried out on this whale, molecular analysis revealed, for the first time, the simultaneous presence of a Dolphin Morbillivirus (DMV) and T. gondii infection coexisting with each other, along with high organochlorine pollutant concentrations, with special reference to DDT.

Conclusion

This report, besides confirming the possibility for Mysticetes to be infected with DMV, highlights the risk of toxoplasmosis in sea water for mammals, already immunodepressed by concurrent factors as infections and environmental contaminants.     

Analysis of basic clustering algorithms for numerical estimation of statistical averages in biomolecules.

In statistical mechanics, the equilibrium properties of a physical system of particles can be calculated as the statistical average over accessible microstates of the system. In general, these calculations are computationally intractable since they involve summations over an exponentially large number of microstates. Clustering algorithms are one of the methods used to numerically approximate these sums. The most basic clustering algorithms first sub-divide the system into a set of smaller subsets (clusters). Then, interactions between particles within each cluster are treated exactly, while all interactions between different clusters are ignored. These smaller clusters have far fewer microstates, making the summation over these microstates, tractable. These algorithms have been previously used for biomolecular computations, but remain relatively unexplored in this context. Presented here, is a theoretical analysis of the error and computational complexity for the two most basic clustering algorithms that were previously applied in the context of biomolecular electrostatics. We derive a tight, computationally inexpensive, error bound for the equilibrium state of a particle computed via these clustering algorithms. For some practical applications, it is the root mean square error, which can be significantly lower than the error bound, that may be more important. We how that there is a strong empirical relationship between error bound and root mean square error, suggesting that the error bound could be used as a computationally inexpensive metric for predicting the accuracy of clustering algorithms for practical applications. An example of error analysis for such an application-computation of average charge of ionizable amino-acids in proteins-is given, demonstrating that the clustering algorithm can be accurate enough for practical purposes.     

New coarse-grained DNA model

A new coarse-grained model of the DNA molecule has been proposed, which was elaborated on the basis of its all-atomic model analysis. The model has been shown to rather well reproduce the DNA structure under low and room temperatures. The Young’s and torsion moduli calculated using the coarse-grained model are in close agreement with experimental data and the theoretical results of other authors. The model can be used for DNA fragments of several hundreds base pairs for rather long time scales (of the order of μs) and for simulating their interactions with other structures.     

Effect of carnosine and 4-methyluracil on the development of experimental hepatitis in rats]

A comparative study of the hepatoprotective effect of carnosine and 4-methyluracil under CCl4-induced acute toxic hepatitis has been carried out. The extent of liver injury and its regeneration were established from morphological data as well as from changes in the activities of alanine aminotransferase (ALT) and histidase and the bilirubin content in blood serum. Hyperlipoperoxidation in the liver and serum was assessed by the amount of TBA-active products. It was found that by day 10 of experimental hepatitis ALT and histidase levels in blood sera of untreated animals exceeded the normal values 1.3- and 3.9-fold, whereas those in the carnosine-treated group approximated the values characteristic of intact animals. The activity of serum ALT in animals treated with vitamin B12 or 4-methyluracil exceeded normal values 1.5 and 1.6 times, whereas that of histidase was 2.5 and 2.7 times as high. Carnosine and 4-methyluracil inhibited (in approximately the same degree) the formation of TBA-active products in the liver. According to morphological dta, cessation of CCl4 injections was accompanied by rapid regeneration of liver tissues in all animal groups. Carnosine enhanced regenerative processes in parenchymatous and connective tissues in a far greater degree in comparison with other drugs. The mitotic index in the carnosine-treated group exceeded more than twofold the corresponding parameters in untreated animals. Possible mechanisms of carnosine action on liver repair are discussed.