The Araucaria Project: A study of the classical Cepheid in the eclipsing binary system OGLE LMC562.05.9009 in the Large Magellanic Cloud

We present a detailed study of the classical Cepheid in the double-lined, highly eccentric eclipsing binary system OGLE-LMC562.05.9009. The Cepheid is a fundamental mode pulsator with a period of 2.988 days. The orbital period of the system is 1550 days. Using spectroscopic data from three 4-8-m telescopes and photometry spanning 22 years, we were able to derive the dynamical masses and radii of both stars with exquisite accuracy. Both stars in the system are very similar in mass, radius and color, but the companion is a stable, non-pulsating star. The Cepheid is slightly more massive and bigger (M_1 = 3.70 +/- 0.03M_sun, R_1 = 28.6 +/- 0.2R_sun) than its companion (M_2 = 3.60 +/- 0.03M_sun, R_2 = 26.6 +/- 0.2R_sun). Within the observational uncertainties both stars have the same effective temperature of 6030 +/- 150K. Evolutionary tracks place both stars inside the classical Cepheid instability strip, but it is likely that future improved temperature estimates will move the stable giant companion just beyond the red edge of the instability strip. Within current observational and theoretical uncertainties, both stars fit on a 205 Myr isochrone arguing for their common age. From our model, we determine a value of the projection factor of p = 1.37 +/- 0.07 for the Cepheid in the OGLE-LMC562.05.9009 system. This is the second Cepheid for which we could measure its p-factor with high precision directly from the analysis of an eclipsing binary system, which represents an important contribution towards a better calibration of Baade-Wesselink methods of distance determination for Cepheids.Comment: Accepted to be published in Ap

Examination of thermal stability of the Ts2631 endolysin.

<p>(A) Samples of Ts2631 endolysin (5 μg) and hen egg white lysozyme (HEWL, 5 μg) were incubated in 10 mM potassium phosphate buffer pH 8.0 at 95°C (0–4 h). Then samples were placed on ice before measuring their activity against <i>T</i>. <i>thermophilus</i> HB8 cells at optimal temperature (60°C for Ts2631 endolysin and 37°C for HEWL). The activity of Ts2631 endolysin and HEWL lysozyme is indicated as percentage relative to untreated samples. Each experiment was repeated in triplicate; error bars indicate the standard deviation. (B) Heat-capacity curves of Ts2631 endolysin determined by: DSC (black dotted line) and the fit to the two-state model [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0137374#pone.0137374.ref031" target="_blank">31</a>] (black solid line) recorded in MES buffer, pH 6.0.</p

Substrate specificity of Ts2631 endolysin.

<p>Relative activities are expressed as the percentage of activity to <i>T</i>. <i>thermophilus</i> HB8 DSM579. Values represent the mean ± standard deviation (n = 3).</p

(A) Effect of pH, (B) NaCl and (C) temperature on the lytic activity of Ts2631 endolysin.

<p>Relative activity against <i>T</i>. <i>thermophilus</i> HB8 cells was calculated by comparing the lytic activity at specific condition with the maximal lytic activity among the dataset. Each experiment was repeated in triplicate; error bars indicate the standard deviation.</p

Thermostability of Ts2631 endolysin substitution variants.

<p>(A) Clear lysates of Ts2631 endolysin substitution variants (1) and supernatants after heat treatment (2) were mixed with Laemmli buffer and loaded on 12.5% SDS-PAGE. Ts2631 endolysin was used as an experiment positive control. Gels were stained with Coomassie Blue R-250 and visualized with use of Gel Doc XR+ Imager (BioRad). (B) Ts2631 endolysin and its substitution variants (20 μl) at concentration 0.5 mg ml^-1 were incubated in thermocycler at indicated temperature gradient for 15 min. Samples were subsequently centrifuged (10,000 × g; 20 min; 4°C) to remove aggregated proteins. Supernatant (6 μl) from each fraction was mixed with Laemmli buffer and loaded on 12.5% SDS-PAGE. Densitometric analysis was performed with use of Image Lab 5.1 BETA software (Bio-Rad). At 75°C 73% of H131N and more than 32% of H30N and C139S aggregate. At 85°C native enzyme and T137K variant were soluble, while at 90.4°C only Ts2631 endolysin remained in the supernatant.</p

Lytic activity of Ts2631 endolysin.

<p>Different concentrations of Ts2631 endolysin were added to chloroform treated <i>T</i>. <i>thermophilus</i> HB8 DSM 579 cell suspension in 200 μl of 10 mM potassium phosphate buffer pH 8.0 and a decrease in OD₆₀₀ was measured for 40 min in 5 min intervals. Results of the control (buffer only) were subtracted from the sample values. The experiment was repeated in triplicate; error bars indicate the standard deviation.</p

Effect of divalent metal cations on lytic activity of Ts2631 endolysin.

<p>^a Lytic activities under standard conditions before EDTA treatment of Ts2631 endolysin.</p><p>^b Lytic activities of EDTA treated endolysins.</p><p>^c Lytic activities of EDTA treated and dialyzed endolysins against chloroform treated <i>T</i>. <i>thermophilus</i> HB8 cells supplemented with different divalent metal ions at 0.1 and 1 mM concentrations.</p><p>Activities are expressed as percentage in relation to the non-treated endolysin control. Values represent the mean ± standard deviation (n = 3).</p