Determination of subunit exchange rate constant of HSP18 subunits at different temperatures.

<p><b>(A)</b> Time-course alterations in the emission spectrum of Alexa fluor-350 and 488 labeled <i>M</i>. <i>leprae</i> HSP18 due to subunit exchange at 37°C. The emission spectra were recorded at different time points after mixing equal amount of Alexa fluor-350 labeled and Alexa fluor-488 labeled HSP18 (1 mg/ml each) at 37°C. The fluorescence spectra were recorded from 400 to 600 nm at 37°C using the excitation wavelength of 346 nm. The slit width of both excitation and emission monochromators was 5 nm each. The scan rate used for this assay was 240 nm/min. <b>(B)</b> Time-dependent decrease in fluorescence intensity at 440 nm for data shown in panel A. <b>(C)</b> Time-dependent increase in fluorescence intensity at 513 nm for data shown in panel A. <b>(D)</b> Temperature-dependent subunit exchange rate of <i>M</i>. <i>leprae</i> HSP18. Subunit exchange between Alexa fluor-350 labeled and Alexa fluor-488 labeled HSP18 was monitored at 25, 31, 37 and 43°C. Symbols in panel B, C and D represent the experimental data points and the solid lines in these pannels represent the best fit of the data according to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0129734#pone.0129734.e003" target="_blank">Eq 3</a>.