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. 2013 Sep 1;19(7):665–682. doi: 10.1089/ars.2012.4859

FIG. 4.

FIG. 4.

ISC formation on trypanothione. (A) One mM T(SH)2 was incubated with the individual components of the reconstitution assay (5 mM dithiothreitol, 10 μM pyridoxal-5′-phosphate, 500 μM Fe2+, 5 μM EcIscS, and 500 μM cysteine) as described under the Material and Methods section, but in the absence of Grxs. The UV–visible spectrum of each mixture was recorded after centrifugation. (B) The ISC reconstitution mixture containing all components (black solid line in A) was subjected to gel chromatography. The major peak (star) elutes with a retention time corresponding to an LMM component. EcIscS indicates the elution peak of E. coli cysteine desulfurase. (Abs. 280 nm: black solid line, Abs. 420 nm: gray dashed line). (C) The UV–visible spectrum of the peak fraction from B (star) containing the T(SH)2-ISC complex. (D) The CD Spectra of the T(SH)2-ISC complex (gray line) and free T(SH)2 (black line). CD, circular dichroism; LMM, low molecular mass.