Change in absorbance at 360 nm plotted as a function of the [P_i] as a function of pH together with the reaction scheme.

<p>Change in absorbance at 360 nm plotted as a function of the [P_i] as a function of pH together with the reaction scheme.</p

Simplified schematic illustration of the dynamic behaviour of FtsZ during the cell division cycle.

<p>Cell growth occurs continuously and when the cell reaches an appropriate size, the plasma membrane and cell wall grow inwards to divide the cell in two between the two sets of daughter chromosomes.</p

FtsZ (11 µM) incubated in the polymerisation buffer (50 mM MES).

<p>(a) pH = 6.0, (b) pH = 6.5, (c) pH = 7, for 15 minutes prior to the addition of GTP (0.2 mM). The reactions were stopped at 4 mins after GTP addition and samples were negatively stained by the addition of 1% uranyl acetate. Selected areas were photographed at ×75,000. Scale bar represents 0.2 µm.</p

EM images of FtsZ polymerisation at different time points.

<p>FtsZ (11 µM) was incubated in the polymerisation buffer (pH = 6.5) for 15 minutes prior to the addition of GTP (0.2 mM). At the indicated times the reaction was stopped and the samples were negatively stained by the addition of 1% uranyl acetate. Scale bar represents 0.2 µm for ×75,000 magnification and 100 nm for ×120,000 magnification.</p

P_i released from the enzymatic conversion of GTP to GDP by FtsZ in the presence of MESG as a function of time, <i>t</i>.

<p>FtsZ (11 µM in MES buffer, pH = 6.0, 6.5 and 7.0); MESG (initial concentration 400 µM). GTP was added to the mixture at <i>t</i> = 0. The change in absorbance at 360 nm was measured in a 0.5 cm cuvette and converted to percentage P_i release with 100% denoting complete conversion of 0.2 mM GTP according to the calibration curves in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0019369#pone-0019369-g007" target="_blank">Figure 7</a>. The experimental error for the assay is in the range of 5%.</p

Polymerisation and depolymerisation of FtsZ.

<p>FtsZ (11 µM) in polymerisation buffer (50 mM MES buffer pH 6.5, 10 mM MgCl₂ and 50 mM KCl) monitored by light scattering and <i>LD</i>₂₁₀. Freshly thawed GTP (0.2 mM) was added at 0 minutes.</p

P_i release when GTP and GDP are sequentially added to FtsZ.

<p>(a) GDP (0.2 mM) and GTP (0.2 mM) and (b) GDP (0.2 mM), GDP (0.2 mM), GDP (0.67 mM), GTP (0.2 mM) are sequentially added as indicated to FtsZ (11 µM in MES buffer, pH 6.5) in the presence of MESG (initial concentration 400 µM) as a function of time. Figure (a) curve is overlaid in (b) in grey with an axis offset to align the first GDP addition. The change in absorbance at 360 nm was measured in a 0.5 cm cuvette and converted to percentage P_i release with 100% denoting complete conversion of 0.2 mM GTP according to the calibration curves in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0019369#pone-0019369-g006" target="_blank">Figure 6</a>. The experimental error for the assay is in the range of 5%.</p

Light scattering of FtsZ as a function of pH.

<p>FtsZ (11 µM) in (a) MES buffer (50 mM pH 6.5), KCl (50 mM) and MgCl₂ (10 mM) with addition GTP (0.2 mM) at <i>t</i> = 0 and at <i>t</i> = 16 min, and (b) in MES buffer (50 mM pH as indicated on the figure), KCl (50 mM) and MgCl₂ (10 mM) with addition of GTP (0.2 mM) at <i>t</i> = 0. Data were collected every second at room temperature in a 0.3 cm path length fluorimeter cuvette with excitation and emission wavelengths set at 450 nm.</p

<i>LD</i>_{210 nm} of FtsZ.

<p>FtsZ (11 µM) in MES buffer (50 mM pH as indicated in figure), KCl (50 mM) and MgCl₂ (10 mM) with addition GTP (0.2 mM).</p

<i>CD</i> spectrum of FtsZ

<p>(22 µM) from the average of 3 independent data collections in Tris-HCl (50 mM, pH 7.9), KCl (50 mM), EDTA (1 mM) and glycerol (10%). <i>CD</i> spectra were measured using a 0.012 mm demountable cuvette (determined by measuring the absorbance of potassium chromate) at room temperature and data converted to Δε using amino acid concentration.</p