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. 2013 Jul 19;368(1622):20120259. doi: 10.1098/rstb.2012.0259

Figure 3.

Figure 3.

Cofactors and activity of AtTAH18 and AtDRE2. (a) UV-visible absorbance spectrum of purified AtTAH18 (3 µM) in 25 mM Tris–HCl, pH 8.0, 150 mM NaCl. (b) Electron transfer from NADPH to horse heart cytochrome c (41 µM) by purified AtTAH18. Best fits of experimental points to the Michaelis Menten equation (solid lines) are for a turnover number of 0.074 or 0.046 s−1 (at infinite cytochrome c concentration) and Km of 50 μM or 28 µM for NADPH and NADH, respectively. Buffer as in (a). (c) UV-visible absorbance spectra of 30 µM purified AtDRE2 before (as isolated) and after chemical reconstitution and desalting (reconstituted). Buffer as in (a). (d) X band EPR spectra of purified (as isolated) AtDRE2 (100 μM) reduced with 2 mM sodium dithionite in buffer. EPR conditions: microwave frequency, 9.458 GHz; microwave power, 1.3 mW (10 K) or 20 mW (84 K); modulation amplitude, 1.25 mT; modulation frequency, 100 kHz. Simulation parameters for the EPR signal: gz, gy and gx; 1.998, 1.959 and 1.916 with line widths of 2.3, 4.1 and 3.4 mT, respectively.