Human Tat-specific factor 1 binds the HIV-1 genome and selectively transports HIV-1 RNAs

Molecular Biology Reports - Human immunodeficiency virus type 1 (HIV-1) propagation requires many human cofactors. Multiple groups have demonstrated that Tat-specific factor 1 (Tat-SF1) is an HIV-1...     

Direct measurement of DNA bending by type IIA topoisomerases: implications for non-equilibrium topology simplification

Type IIA topoisomerases modify DNA topology by passing one segment of duplex DNA (transfer or T–segment) through a transient double-strand break in a second segment of DNA (gate or G–segment) in an ATP-dependent reaction. Type IIA topoisomerases decatenate, unknot and relax supercoiled DNA to levels below equilibrium, resulting in global topology simplification. The mechanism underlying this non-equilibrium topology simplification remains speculative. The bend angle model postulates that non-equilibrium topology simplification scales with the bend angle imposed on the G–segment DNA by the binding of a type IIA topoisomerase. To test this bend angle model, we used atomic force microscopy and single-molecule Förster resonance energy transfer to measure the extent of bending imposed on DNA by three type IIA topoisomerases that span the range of topology simplification activity. We found that Escherichia coli topoisomerase IV, yeast topoisomerase II and human topoisomerase IIα each bend DNA to a similar degree. These data suggest that DNA bending is not the sole determinant of non-equilibrium topology simplification. Rather, they suggest a fundamental and conserved role for DNA bending in the enzymatic cycle of type IIA topoisomerases.     

Interrelationship between Cytoplasmic Retroviral Gag Concentration and Gag–Membrane Association

The early events in the retrovirus assembly pathway, particularly the timing and nature of Gag translocation from the site of protein translation to the inner leaflet of the plasma membrane, are poorly understood. We have investigated the interrelationship between cytoplasmic Gag concentration and plasma membrane association using complementary live-cell biophysical fluorescence techniques in real time with both human T-cell leukemia virus type 1 (HTLV-1) and human immunodeficiency virus type 1 (HIV-1) Gag proteins. In particular, dual-color, z-scan fluorescence fluctuation spectroscopy in conjunction with total internal reflection fluorescence and conventional, epi-illumination imaging were utilized. Our results demonstrate that HTLV-1 Gag is capable of membrane targeting and particle assembly at low (i.e., nanomolar) cytoplasmic concentrations and that there is a critical threshold concentration (approaching micromolar) prior to the observation of HIV-1 Gag associated with the plasma membrane. These observations imply fundamental differences between HIV-1 and HTLV-1 Gag trafficking and membrane association.     

Mutational analysis of the helicase-like domain of Thermotoga maritima reverse gyrase

Reverse gyrase is a unique type IA topoisomerase that is able to introduce positive supercoils into DNA in an ATP-dependent process. ATP is bound to the helicase-like domain of the enzyme that contains most of the conserved motifs found in helicases of the SF1 and SF2 superfamilies. In this paper, we have investigated the role of the conserved helicase motifs I, II, V, VI, and Q by generating mutants of the Thermotoga maritima reverse gyrase. We show that mutations in motifs I, II, V, and VI completely eliminate the supercoiling activity of reverse gyrase and that a mutation in the Q motif significantly reduces this activity. Further analysis revealed that for most mutants, the DNA binding and cleavage properties are not significantly changed compared with the wild type enzyme, whereas their ATPase activity is impaired. These results clearly show that the helicase motifs are tightly involved in the coupling of ATP hydrolysis to the topoisomerase activity. The zinc finger motif located at the N-terminal end of reverse gyrases was also mutated. Our results indicate that this motif plays an important role in DNA binding.     

Inhaled adenosine A(2A) receptor agonists for the treatment of chronic obstructive pulmonary disease

COPD is a major cause of mortality in the western world. A(2A) agonists are postulated to reduce the lung inflammation that causes COPD. The cardiovascular effects of A(2A) agonists dictate that a compound needs to be delivered by inhalation to be therapeutically useful. A strategy of minimizing side-effect liability by maximizing systemic clearance was followed and pharmacological and pharmacokinetic SAR of a series of inhaled A(2A) agonists described. A sevenfold improvement in potency and 150-fold reduction in side-effect liability over the lead compound CGS-21680, were obtained.     

Relationship between sprint times and the strength/power outputs of a machine squat jump

Strength testing is often used with team-sport athletes, but some measures of strength may have limited prognostic/diagnostic value in terms of the physical demands of the sport. The purpose of this study was to investigate relationships between sprint ability and the kinetic and kinematic outputs of a machine squat jump. Thirty elite level rugby union and league athletes with an extensive resistance-training background performed bilateral concentric-only machine squat jumps across loads of 20% to 90% 1 repetition maximum (1RM), and sprints over 10 meters and 30 or 40 meters. The magnitudes of the relationships were interpreted using Pearson correlation coefficients, which had uncertainty (90% confidence limits) of approximately +/-0.3. Correlations of 10-meter sprint time with kinetic and kinematic variables (force, velocity, power, and impulse) were generally positive and of moderate to strong magnitude (r = 0.32-0.53). The only negative correlations observed were for work, although the magnitude was small (r = -0.18 to -0.26). The correlations for 30- or 40-meter sprint times were similar to those for 10-meter times, although the correlation with work was positive and moderate (r = 0.35-0.40). Correlations of 10-meter time with kinetic variables expressed relative to body mass were generally positive and of trivial to small magnitude (r = 0.01-0.29), with the exceptions of work (r = -0.31 to -0.34), and impulse (r = -0.34 to -0.39). Similar correlations were observed for 30- and 40-meter times with kinetic measures expressed relative to body mass. Although correlations do not imply cause and effect, the preoccupation with maximizing power output in this particular resistance exercise to improve sprint ability appears problematic. Work and impulse are potentially important strength qualities to develop in the pursuit of improved sprinting performance.     

Wrist splint effects on muscle activity and force during a handgrip task

Wrist splints are commonly prescribed to limit wrist motion and provide support at night and during inactive periods but are often used in the workplace. In theory, splinting the wrist should reduce wrist extensor muscle activity by stabilizing the joint and reducing the need for co-contraction to maintain posture. Ten healthy volunteers underwent a series of 24 10-s gripping trials with surface electromyography on 6 forearm muscles. Trials were randomized between splinted and nonsplinted conditions with three wrist postures (30 degrees flexion, neutral, and 30 degrees extension) and four grip efforts. Custom-made Plexiglas splints were taped to the dorsum of the hand and wrist. It was found that when simply holding the dynamometer, use of a splint led to a small (<1% MVE) but significant reduction in activity for all flexor muscles and extensor carpi radialis (all activity <4% maximum). At maximal grip, extensor muscle activity was significantly increased with the splints by 7.9-23.9% MVE. These data indicate that splinting at low-to-moderate grip forces may act to support the wrist against external loading, but appears counterproductive when exerting maximal forces. Wrist bracing should be limited to periods of no to light activity and avoided during tasks that require heavy efforts.