Changes in the hydrodynamic properties of DmChd64 and TcChd64 induced by GdmCl.

<p>(A), (C) SEC analysis of DmChd64 (A) and TcChd64 (B) in the presence of GdmCl. The curves represent profiles of DmChd64 and TcChd64 chromatographed at different GdmCl concentrations. The blue lines represent elution peaks of renatured proteins chromatographed in buffer A. Purified DmChd64 and TcChd64 (0.5 mg/ml in buffer A with an appropriate GdmCl concentration) were injected into a Superdex 200 10/300 GL column equilibrated with buffer A of the same denaturant concentration for each sample at room temperature, with a flow rate of 0.5 ml/min. (B), (D) Using the calibration curve (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0137074#sec002" target="_blank">Materials and Methods</a>) and the elution volumes determined in (A) and (C), the R_s values of DmCh64 (B) and TcChd64 (D) were calculated and plotted against the GdmCl concentration.</p

Fluorescence analysis of the effect of GdmCl on DmChd64 and TcChd64.

<p>Fluorescence emission spectra (<i>λ</i>_<i>ex</i> = 280 nm) of DmChd64 (A) and TcChd64 (B) in the presence of different GdmCl concentrations (a-l). The spectra were recorded between 300 and 400 nm under native (a) or denaturing conditions in the presence of 0.2 M (b), 0.5 M (c), 0.7 M (d), 1.0 M (e), 1.2 M (f), 1.5 M (g), 1.7 M (h), 2.0 M (i), 2.2 M (j), 2.5 M (k), 3.0 M (l) GdmCl. The blue lines represent spectra of proteins recorded after renaturation. Inset: changes in fluorescence intensity monitored at 346 nm for DmChd64 (A) and TcChd64 (B) titrated with GdmCl. Data points were taken from the fluorescence spectra.</p

Analytical gel filtration experiments to determine the MvaI oligomeric state and the stoichiometry of DNA binding for the oligoduplex 1 shown in ()

<p><b>Copyright information:</b></p><p>Taken from "Restriction endonuclease MvaI is a monomer that recognizes its target sequence asymmetrically"</p><p></p><p>Nucleic Acids Research 2007;35(6):2035-2046.</p><p>Published online 7 Mar 2007</p><p>PMCID:PMC1874612.</p><p>© 2007 The Author(s)</p> Elution profiles were recorded simultaneously at 260 and 280 nm and deconvoluted to obtain separate curves for the MvaI (blue) and DNA (red) concentration. () MvaI alone, () DNA alone, () mixture with a 2:1 molar excess of MvaI over DNA, () stoichiometric mixture, () mixture with a 2:1 molar excess of DNA over MvaI, () calibration curve for Superose ™ 12 HR 10/30 column (Amersham Biosciences) with standards from Biorad (vitamin B-12, 1.35 kDa; myoglobin, 17 kDa; ovalbumin, 44kDa; IgG, 150 kDa and thyroglobin, 670 kDa)