Molecular dynamics simulation of human LOX-1 provides an explanation for the lack of OxLDL binding to the Trp150Ala mutant

<p>Abstract</p> <p>Background</p> <p>Dimeric lectin-like oxidized low-density lipoprotein receptor-1 LOX-1 is the target receptor for oxidized low density lipoprotein in endothelial cells. <it>In vivo </it>assays revealed that in LOX-1 the basic spine arginine residues are important for binding, which is lost upon mutation of Trp150 with alanine. Molecular dynamics simulations of the wild-type LOX-1 and of the Trp150Ala mutant C-type lectin-like domains, have been carried out to gain insight into the severe inactivating effect.</p> <p>Results</p> <p>The mutation does not alter the dimer stability, but a different dynamical behaviour differentiates the two proteins. As described by the residues fluctuation, the dynamic cross correlation map and the principal component analysis in the wild-type the two monomers display a symmetrical motion that is not observed in the mutant.</p> <p>Conclusion</p> <p>The symmetrical motion of monomers is completely damped by the structural rearrangement caused by the Trp150Ala mutation. An improper dynamical coupling of the monomers and different fluctuations of the basic spine residues are observed, with a consequent altered binding affinity.</p

Effect on DNA relaxation of the single Thr718Ala mutation in human topoisomerase I: a functional and molecular dynamics study

The functional and dynamical properties of the human topoisomerase I Thr718Ala mutant have been compared to that of the wild-type enzyme using functional assays and molecular dynamics (MD) simulations. At physiological ionic strength, the cleavage and religation rates, evaluated on oligonucleotides containing the preferred topoisomerase I DNA sequence, are almost identical for the wild-type and the mutated enzymes, as is the cleavage/religation equilibrium. On the other hand, the Thr718Ala mutant shows a decreased efficiency in a DNA plasmid relaxation assay. The MD simulation, carried out on the enzyme complexed with its preferred DNA substrate, indicates that the mutant has a different dynamic behavior compared to the wild-type enzyme. Interestingly, no changes are observed in the proximity of the mutation site, whilst a different flexibility is detected in regions contacting the DNA scissile strand, such as the linker and the V-shaped α helices. Taken together, the functional and simulation results indicate a direct communication between the mutation site and regions located relatively far away, such as the linker domain, that with their altered flexibility confer a reduced DNA relaxation efficiency. These results provide evidence that the comprehension of the topoisomerase I dynamical properties are an important element in the understanding of its complex catalytic cycle

The different cleavage DNA sequence specificity explains the camptothecin resistance of the human topoisomerase I Glu418Lys mutant

Yeast cells expressing the Glu418Lys human topoisomerase I mutant display a camptothecin resistance that slowly decreases as a function of time. Molecular characterization of the single steps of the catalytic cycle of the purified mutant indicates that it has a relaxation activity identical to the wild-type protein but a different DNA sequence specificity for the cleavage sites when compared to the wild-type enzyme, as assayed on several substrates. In particular the mutant has a low specificity for CPT sensitive cleavable sites. In fact, the mutant has, at variance of the wild-type enzyme, a reduced preference for cleavage sites having a thymine base in position −1 of the scissile strand. This preference, together with the strict requirement for a thymine base in position −1 for an efficient camptothecin binding, explains the temporary camptothecin resistance of the yeast cell expressing the mutant and points out the importance of the DNA sequence in the binding of the camptothecin drug

Effect of a standardized meal on the threshold of exercise-induced myocardial ischemia in patients with stable angina

AbstractObjectives. This study was undertaken to determine the effect of a standardized meal on the ischemie threshold and exercise capacity in a series of 20 patients with stable angina, exerciseinduced ischemia and reversible exercise-induced perfusion defects.Background. It is generally accepted that exercise tolerance in patients with angina is reduced after a meal. However, studies that have addressed this phenomenon have yielded results that are contradictory and inconclusive.Methods. Two exercise tests using the Brace protocol with technetium-99m (99mTc)-sestamibi were performed on consecutive days in a randomized order. One test was performed in the fasting state and the other 30 min after a 1,000-calorle meal.Results. In the postprandial state, exercise time to ischemia was reduced by 20% from 248 ± 93 s to 197 ±87 s (p = 0.0007), time to angina by 15% from 340 ± 82 s to 287 ± 94 s (p = 0.002) and exercise tolerance by 9% from 376 ± 65 s to 344 ± 86 s (p = 0.002). Rate-pressure products at these exercise test end points were not significantly different in the fasting and postprandial tests, and the quantitative 99mTc-sestamibi ischemia score was unchanged.Conclusions. In patients with stable angina, a 1,000-calorie meal significantly reduced tine to ischemia, time to angina and exercise tolerance because of a more rapid increase in myocardial oxygen demand with exercise. The extent and severity of exerciseinduced ischemia were unchanged

editorial preface ati 2018 energy procedia

Abstract The 73rd Conference of the Italian Thermal Machines Engineering Association (ATI) was held in Pisa (Italy) on September 12-14, 2018. The conference was organized by ATI and the University of Pisa. The main topic of this conference edition was: "Innovation and research for a sustainable energy future" In the last 10 years, the share of renewable sources in the energy mix of several countries has increased at a steady pace. This led to a revolution in the way energy conversion is conceived and distributed in comparison to a fossil based system. Nowadays, thinking about a 100% renewable energy system is no more just a dream. Nevertheless, the transition to this future implies several critical decisions from the technical and economic point of view. The 73rd ATI conference was the opportunity to discuss these topics, present new frontiers of the energy engineering research and promote the cooperation between researchers. The topics of the conference were: Heat Transfer and Fluid Dynamics, Energetics of Buildings, Air Conditioning and Refrigeration, Environmental Aspects of Energy Conversion, Renewable Energy, Advanced Energy Conversion and Storage Systems, Innovative Propulsion Systems and Internal Combustion Engines, Turbomachinery, Combustion and Fuels, Fluid Power. This issue of Energy Procedia includes all the papers presented at the conference, 143 in oral form and 20 in the poster session reserved to PhD students. The conference also included a plenary session and two panel sessions with outstanding keynote speakers from both academy and industry. The Guest Editors of the 73rd Conference of the Italian Thermal Machines Engineering Associatio

ATP independent type IB topoisomerase of Leishmania donovani is stimulated by ATP: an insight into the functional mechanism

Most type IB topoisomerases do not require ATP and Mg2+ for activity. However, as shown previously for vaccinia topoisomerase I, we demonstrate that ATP stimulates the relaxation activity of the unusual heterodimeric type IB topoisomerase from Leishmania donovani (LdTOP1L/S) in the absence of Mg2+. The stimulation is independent of ATP hydrolysis but requires salt as a co-activator. ATP binds to LdTOP1L/S and increases its rate of strand rotation. Docking studies indicate that the amino acid residues His93, Tyr95, Arg188 and Arg190 of the large subunit may be involved in ATP binding. Site directed mutagenesis of these four residues individually to alanine and subsequent relaxation assays reveal that the R190A mutant topoisomerase is unable to exhibit ATP-mediated stimulation in the absence of Mg2+. However, the ATP-independent relaxation activities of all the four mutant enzymes remain unaffected. Additionally, we provide evidence that ATP binds LdTOP1L/S and modulates the activity of the otherwise ATP-independent enzyme. This study establishes ATP as an activator of LdTOP1L/S in the absence of Mg2

Solvent Dependency of the UV-Vis Spectrum of Indenoisoquinolines: Role of Keto-Oxygens as Polarity Interaction Probes.

Indenoisoquinolines are the most promising non-campthotecins topoisomerase IB inhibitors. We present an integrated experimental/computational investigation of the UV-Vis spectra of the IQNs parental compound (NSC314622) and two of its derivatives (NSC724998 and NSC725776) currently undergoing Phase I clinical trials. In all the three compounds a similar dependence of the relative absorption intensities at 270 nm and 290 nm on solvent polarity is found. The keto-oxygens in positions 5 and 11 of the molecular scaffold of the molecule are the principal chromophores involved in this dependence. Protic interactions on these sites are also found to give rise to absorptions at wavelengthsolution, due to the stabilization of highly polarized tautomers of the molecule. These results suggest that the keto-oxygens are important polarizable groups that can act as useful interactors with the molecular receptor, providing at the same time an useful fingerprint for the monitoring of the drug binding to topoisomerase IB