Characterization of 3-chlorobenzoate degrading aerobic bacteria isolated under various environmental conditions

The rates of bacterial growth in nature are often restricted by low concentrations of oxygen or carbon substrates. In the present study the metabolic properties of 24 isolates that had been isolated using various concentrations of 3-chlorobenzoate, benzoate and oxygen as well as using continuous culture at high and low growth rates were determined to investigate the effects of these parameters on the metabolism of monoaromatic compounds. Bacteria were enriched from different sampling sites and subsequently isolated. In batch culture this was done both under low oxygen (2% O(2)) and air-saturated concentrations. Chemostat enrichments were performed under either oxygen or 3-chlorobenzoate limiting conditions. Bacteria metabolizing aromatics with gentisate or protocatechuate as intermediates (gp bacteria) as well as bacteria metabolizing aromatic compounds via catechols (cat bacteria) were isolated from batch cultures when either benzoate or 3CBA were used as C sources, regardless of the enrichment conditions applied. In contrast, enrichments performed in chemostats at low dilution rates resulted in gp-type organisms only, whereas at high dilution rates cat-type organisms were enriched, irrespective of the oxygen and 3-chlorobenzoate concentration used during enrichment. It is noteworthy that the gp-type of bacteria possessed relatively low μ(max) values on 3CBA and benzoate along with relatively high substrate and oxygen affinities for these compounds. This is in contrast with cat-type of bacteria, which seemed to be characterized by high maximum specific growth rates on the aromatic substrates and relatively high apparent half saturation constants. In contrast, bacteria degrading chlorobenzoate via gentisate or protocatechuate may possibly be better adapted to conditions leading to growth at reduced rates such as low oxygen and low substrate concentrations.     

Bacillus stearothermophilus PcrA monomer is a single-stranded DNA translocase but not a processive helicase in vitro

Structural studies of the Bacillus stearothermophilus PcrA protein along with biochemical studies of the single-stranded (ss) DNA translocation activity of PcrA monomers have led to the suggestion that a PcrA monomer possesses processive helicase activity in vitro. Yet definitive studies testing whether the PcrA monomer possesses processive helicase activity have not been performed. Here we show, using single turnover kinetic methods, that monomers of PcrA are able to translocate along ssDNA, in the 3' to 5' direction, rapidly and processively, whereas these same monomers display no detectable helicase activity under the same solution conditions in vitro. The PcrA monomer ssDNA translocation activity, although necessary, is not sufficient for processive helicase activity, and thus the translocase and helicase activities of PcrA are separable. These results also suggest that the helicase activity of PcrA needs to be activated either by self-assembly or through interactions with accessory proteins. This same behavior is displayed by both the Escherichia coli Rep and UvrD monomers. Hence, all three of these SF1 enzymes are ssDNA translocases as monomers but do not display processive helicase activity in vitro unless activated. The fact that the translocase and helicase activities are separable suggests that each activity may be used for different functions in vivo.     

Changes in behavior are unable to disrupt a trophic cascade involving a specialist herbivore and its food plant

Changes in ecological conditions can induce changes in behavior and demography of wild organisms, which in turn may influence population dynamics. Black brant (Branta bernicla nigricans) nesting in colonies on the Yukon–Kuskokwim Delta (YKD) in western Alaska have declined substantially (~50%) since the turn of the century. Black brant are herbivores that rely heavily on Carex subspathacea (Hoppner's sedge) during growth and development. The availability of C. subspathacea affects gosling growth rates, which subsequently affect pre‐ and postfledging survival, as well as size and breeding probability as an adult. We predicted that long‐term declines in C. subspathacea have affected gosling growth rates, despite the potential of behavior to buffer changes in food availability during brood rearing. We used Bayesian hierarchical mixed‐effects models to examine long‐term (1987–2015) shifts in brant behavior during brood rearing, forage availability, and gosling growth rates at the Tutakoke River colony. We showed that locomotion behaviors have increased (β = 0.05, 95% CRI: 0.032–0.068) while resting behaviors have decreased (β = ?0.024, 95% CRI: ?0.041 to ?0.007), potentially in response to long‐term shifts in forage availability and brood density. Concurrently, gosling growth rates have decreased substantially (β = ?0.100, 95% CRI: ?0.191 to ?0.016) despite shifts in behavior, mirroring long‐term declines in the abundance of C. subspathacea (β = ?0.191, 95% CRI: ?0.355 to ?0.032). These results have important implications for individual fitness and population viability, where shifts in gosling behavior putatively fail to mitigate long‐term declines in forage availability.     

Cryptic species as a window on diversity and conservation

The taxonomic challenge posed by cryptic species (two or more distinct species classified as a single species) has been recognized for nearly 300 years, but the advent of relatively inexpensive and rapid DNA sequencing has given biologists a new tool for detecting and differentiating morphologically similar species. Here, we synthesize the literature on cryptic and sibling species and discuss trends in their discovery. However, a lack of systematic studies leaves many questions open, such as whether cryptic species are more common in particular habitats, latitudes or taxonomic groups. The discovery of cryptic species is likely to be non-random with regard to taxon and biome and, hence, could have profound implications for evolutionary theory, biogeography and conservation planning.     

Active displacement of RecA filaments by UvrD translocase activity

The UvrD helicase has been implicated in the disassembly of RecA nucleoprotein filaments in vivo and in vitro. We demonstrate that UvrD utilizes an active mechanism to remove RecA from the DNA. Efficient RecA removal depends on the availability of DNA binding sites for UvrD and/or the accessibility of the RecA filament ends. The removal of RecA from DNA also requires ATP hydrolysis by the UvrD helicase but not by RecA protein. The RecA-removal activity of UvrD is slowed by RecA variants with enhanced DNA-binding properties. The ATPase rate of UvrD during RecA removal is much slower than the ATPase activity of UvrD when it is functioning either as a translocase or a helicase on DNA in the absence of RecA. Thus, in this context UvrD may operate in a specialized disassembly mode.     

A preliminary investigation into the relationship between functional movement screen scores and athletic physical performance in female team sport athletes

There is little research investigating relationships between the Functional Movement Screen (FMS) and athletic performance in female athletes. This study analyzed the relationships between FMS (deep squat; hurdle step [HS]; in-line lunge [ILL]; shoulder mobility; active straight-leg raise [ASLR]; trunk stability push-up; rotary stability) scores, and performance tests (bilateral and unilateral sit-and-reach [flexibility]; 20-m sprint [linear speed]; 505 with turns from each leg; modified T-test with movement to left and right [change-of-direction speed]; bilateral and unilateral vertical and standing broad jumps; lateral jumps [leg power]). Nine healthy female recreational team sport athletes (age = 22.67 ± 5.12 years; height = 1.66 ± 0.05 m; body mass = 64.22 ± 4.44 kilograms) were screened in the FMS and completed the afore-mentioned tests. Percentage between-leg differences in unilateral sit-and-reach, 505 turns and the jumps, and difference between the T-test conditions, were also calculated. Spearman''s correlations (p ≤ 0.05) examined relationships between the FMS and performance tests. Stepwise multiple regressions (p ≤ 0.05) were conducted for the performance tests to determine FMS predictors. Unilateral sit-and-reach positive correlated with the left-leg ASLR (r = 0.704-0.725). However, higher-scoring HS, ILL, and ASLR related to poorer 505 and T-test performance (r = 0.722-0.829). A higher-scored left-leg ASLR related to a poorer unilateral vertical and standing broad jump, which were the only significant relationships for jump performance. Predictive data tended to confirm the correlations. The results suggest limitations in using the FMS to identify movement deficiencies that could negatively impact athletic performance in female team sport athletes.     

The importance of coulombic end effects: experimental characterization of the effects of oligonucleotide flanking charges on the strength and salt dependence of oligocation (L8+) binding to single-stranded DNA oligomers.

Binding constants Kobs, expressed per site and evaluated in the limit of zero binding density, are quantified as functions of salt (sodium acetate) concentration for the interactions of the oligopeptide ligand KWK6NH2 (designated L8+, with ZL = 8 charges) with three single-stranded DNA oligomers (ss dT-mers, with |ZD| = 15, 39, and 69 charges). These results provide the first systematic experimental information about the effect of changing |ZD| on the strength and salt dependence of oligocation-oligonucleotide binding interactions. In a comparative study of L8+ binding to poly dT and to a short dT oligomer (|ZD| = 10),. Proc. Natl. Acad. Sci. USA. 93:2511-2516) demonstrated the profound thermodynamic effects of phosphate charges that flank isolated nonspecific L8+ binding sites on DNA. Here we find that both Kobs and the magnitude of its power dependence on salt activity (|SaKobs|) increase monotonically with increasing |ZD|. The dependences of Kobs and SaKobs on |ZD| are interpreted by introducing a simple two-state thermodynamic model for Coulombic end effects, which accounts for our finding that when L8+ binds to sufficiently long dT-mers, both DeltaGobso = -RT ln Kobs and SaKobs approach the values characteristic of binding to poly-dT as linear functions of the reciprocal of the number of potential oligocation binding sites on the DNA lattice. Analysis of our L8+-dT-mer binding data in terms of this model indicates that the axial range of the Coulombic end effect for ss DNA extends over approximately 10 phosphate charges. We conclude that Coulombic interactions cause an oligocation (with ZL < |ZD|) to bind preferentially to interior rather than terminal binding sites on oligoanionic or polyanionic DNA, and we quantify the strong increase of this preference with decreasing salt concentration. Coulombic end effects must be considered when oligonucleotides are used as models for polyanionic DNA in thermodynamic studies of the binding of charged ligands, including proteins.     

Immortalization and characterization of Nijmegen Breakage syndrome fibroblasts.

Nijmegen Breakage Syndrome (NBS) is a very rare autosomal recessive chromosomal instability disorder characterized by microcephaly, growth retardation, immunodeficiency and a high incidence of malignancies. Cells from NBS patients are hypersensitive to ionizing radiation (IR) and display radioresistant DNA synthesis (RDS). NBS is caused by mutations in the NBS1 gene on chromosome 8q21 encoding a protein called nibrin. This protein is a component of the hMre11/hRad50 protein complex, suggesting a defect in DNA double-strand break (DSB) repair and/or cell cycle checkpoint function in NBS cells. We established SV40 transformed, immortal NBS fibroblasts, from primary cells derived from a Polish patient, carrying the common founder mutation 657del5. Immortalized NBS cells, like primary cells, are X-ray sensitive (2-fold) and display RDS following IR. They show an increased sensitivity to bleomycin (3.5-fold), etoposide (2.5-fold), camptothecin (3-fold) and mitomycin C (1.5-fold), but normal sensitivity towards UV-C. Despite the clear hypersensitivity towards DSB-inducing agents, the overall rates of DSB-rejoining in NBS cells as measured by pulsed field gel electrophoresis were found to be very similar to those of wild type cells. This indicates that the X-ray sensitivity of NBS cells is not directly caused by an overt defect in DSB repair.