Fructanolytic and saccharolytic enzymes of Treponema zioleckii strain kT

Enzymes in the newly described rumen bacterium, Treponema zioleckii strain kT, capable of digesting Timothy grass fructan, inulin, and sucrose were identified and characterized. Two specific endolevanases and one non-specific β-fructofuranosidase were found in a cell-free extract. The molecular weight of the endolevanases were estimated to be 60 and 36 kDa, whereas that of β-fructofuranosidase, 87 kDa. The former of the specific enzymes was associated with the outer membrane, while the latter and the non-specific β-fructofuranosidase, with the periplasm or cytosol. The K_m and V_max for Timothy grass fructan degradation by endolevanase were 0.27% and 15.75 μM fructose equivalents × mg protein^?1 × min^?1, those for sucrose and inulin digestion by β-fructofuranosidase were 1.35 × 10^?3 M and 1.73 μM hexoses × mg protein^?1 × min^?1 and 1.77% and 1.83 μM hexoses × mg protein^?1 × min^?1, respectively.     

Muskrat (Ondatra zibethicus) decline after the expansion of American mink (Neovison vison) in Poland

Field survey data in Central Poland revealed that the proportion of sites inhabited by muskrats decreased from 44% to 7% over one decade. This corresponded to the decline in hunting bags of muskrat over the whole of Poland. The largest hunting harvest of muskrat was recorded in 1987/1988 (66,416 individuals), the smallest in 2007/2008 (4,567 individuals). The decline in hunting bags occurred in all regions analysed; however, it was most rapid in the north and north-east. Before the expansion of mink, which started in northern Poland at the beginning of the 1980s, muskrat densities in particular regions depended on the availability of aquatic habitats. A comparison of hunting bags of muskrat and American mink in years 2002–2008 indicated a significant negative correlation between the numbers of these two species harvested in seven regions of Poland. The negative correlation between numbers of muskrat and mink suggests that mink predation is one of the most important factors in the decline of the muskrat population in Poland.     

Are linear‐leaved Potamogeton hybrids really so rare? Molecular evidence for multiple hybridizations between P. acutifolius and P. compressus in central Europe

Due to insufficient morphological differentiation, hybridization among linear‐leaved species of Potamogeton is rarely observed and recognized. Here, we applied molecular tools (sequencing of nrITS and rpl32‐trnL cpDNA intergenic spacer) to study several natural, central European populations of morphologically intermediate forms between two closely related species, P. acutifolius and P. compressus, to examine their taxonomical status and test whether they represented the variation range of the species or are results of interspecific hybridization. Both DNA regions provided distinguishing characters between putative parental species. The ITS sequences from all the morphologically intermediate specimens displayed an additive pattern combining the variation of both parental species, providing evidence for the hybrid origin of all these plants. This case study suggests that hybrids of linear‐leaved Potamogeton species are more common than usually believed, but have been difficult to identify without molecular tools.     

The role of hydrophobic interactions in ankyrin-spectrin complex formation

Spectrin and ankyrin are the key components of the erythrocyte cytoskeleton. The recently published crystal structure of the spectrin-ankyrin complex has indicated that their binding involves complementary charge interactions as well as hydrophobic interactions. However, only the former is supported by biochemical evidence. We now show that nonpolar interactions are important for high affinity complex formation, excluding the possibility that the binding is exclusively mediated by association of distinctly charged surfaces. Along these lines we report that substitution of a single hydrophobic residue, F917S in ankyrin, disrupts the structure of the binding site and leads to complete loss of spectrin affinity. Finally, we present data showing that minimal ankyrin binding site in spectrin is formed by helix 14C together with the loop between helices 15 B/C.     

Real time PCR hybridization for the rapid and specific identification of Francisella tularensis

Tularemia is highly infectious and fatal zoonotic disease caused by Gram negative bacteria Francisella tularensis. The necessity to undergo medical treatment in early phase of illness in humans and possibility of making use of bacterial aerosol by terrorists in an attack create an urgent need to implement a rapid and effective method which enables to identify the agent. In our study two primers FopA F/R and hybridization probes FopA S1/S2 designed from fopA gene sequence, were tested for their potential applicability to identify F. tularensis. In this research 50 strains of F. tularensis were used and the test gave positive results. Reaction specificity was confirmed by using of non-Francisella tularensis bacterial species. The results obtained in the real-time PCR reaction with primers Tul4 F/R and hybridization probes Tul4 S1/S2, designed from tul4 gene, were comparable to the results from previous experiment with fopA - primers set. Investigation of fopA and tul4 primers and hybridization probes properties revealed characteristic Tm (melting temperature) value of the products--61 degrees C and 60 degrees C, respectively. Detection sensitivity was remarkably higher when fopA primers set was used 1 fg/microl, and for tul4 primers set, minimal detectable concentration is 10 fg/microl.     

Overproduced <Emphasis Type="Italic">Brucella abortus</Emphasis> PdhS-mCherry forms soluble aggregates in <Emphasis Type="Italic">Escherichia coli</Emphasis>, partially associating with mobile foci of IbpA-YFP

Background  

When heterologous recombinant proteins are produced in Escherichia coli, they often precipitate to form insoluble aggregates of unfolded polypeptides called inclusion bodies. These structures are associated with chaperones like IbpA. However, there are reported cases of "non-classical" inclusion bodies in which proteins are soluble, folded and active.     

Vacuolar ethylene formation does not depend on membrane potential

Enzymatic synthesis of ethylene in the vacuole is assumed to require membrane integrity. The possibility that this reflects dependence on the vacuolar membrane potential was investigated. Vacuoles were released from protoplasts isolated from leaves of Vicia faba L. cv. Cyprus. The dependence of the ethylene-forming activity on tonoplast integrity was re-examined by immobilization of the vacuoles in a cross-linked polymeric matrix and subsequent permeabilization of the tonoplast with toluene, a pore-forming reagent. The relationship between the vacuolar ethylene formation and the membrane potential of free vacuoles was investigated by following the uptake of thiocyanate using permeabilized, depolarized and hyperpolarized vacuoles. Toluene and the proton conductor carbonyl cyanide m -chlorophenylhydrazone (CCCP) caused loss of ethylene-forming activity and depolarized the vacuolar membrane potential. However, depolarization of the membrane potential with choline chloride and hyperpolarization by ATP did not affect ethylene biosynthesis. These conflicting results lead to the conclusion that vacuolar ethylene biosynthesis is not dependent on the vacuolar membrane potential. The possibility that the inhibition of ethylene biosynthesis by toluene and CCCP may result from direct hydrophobic interactions between these compounds and hydrophobic components of the ethylene-forming enzyme is discussed.     

Scale and structure of time-averaging (age mixing) in terrestrial gastropod assemblages from Quaternary eolian deposits of the eastern Canary Islands

Quantitative estimates of time-averaging (age mixing) in gastropod shell accumulations from Quaternary (the late Pleistocene and Holocene) eolian deposits of Canary Islands were obtained by direct dating of individual gastropods obtained from exceptionally well-preserved dune and paleosol shell assemblages. A total of 203 shells of the gastropods Theba geminata and T. arinagae, representing 44 samples (= stratigraphic horizons) from 14 sections, were dated using amino acid (isoleucine) epimerization ratios calibrated with 12 radiocarbon dates. Most samples reveal a substantial variation in shell age that exceeds the error that could be generated by dating imprecision, with the mean within-sample shell age range of 6670 years and the mean standard deviation of 2920 years. Even the most conservative approach (Monte Carlo simulations with a non-sequential Bonferroni correction) indicates that at least 25% of samples must have undergone substantial time-averaging (e.g., age variations within those samples cannot be explained by dating imprecision alone). Samples vary in shell age structure, including both left-skewed (17 out of 44) and right-skewed distributions (26 out of 44) as well as age distributions with a highly variable kurtosis. Dispersion and shape of age distributions of samples do not show any notable correlation with the stratigraphic age of samples, suggesting that the structure and scale of temporal mixing is time invariant. The statistically significant multi-millennial time-averaging observed here is consistent with previous studies of shell accumulations from various depositional settings and reinforces the importance of dating numerous specimens per horizon in geochronological studies. Unlike in the case of marine samples, typified by right-skewed age distributions (attributed to an exponential-like shell loss from older age classes), many of the samples analyzed here displayed left-skewed distributions, suggestive of different dynamics of age mixing in marine versus terrestrial shell accumulations.