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. 2008 Aug 11;182(3):531–542. doi: 10.1083/jcb.200711151

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© 2008 Ju et al.

This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.jcb.org/misc/terms.shtml). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/).

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Figure 7. — Cosmc directly interacts with T-synthase. The N-terminal His-tagged soluble Cosmc were expressed in Hi-5 insect cells. The human N-terminal HPC4 epitope-tagged soluble T-synthase was expressed by cotransfecting WT Cosmc in Hi-5 insect cells. Proteins were purified directly from the media and used in coprecipitation experiments to test physical association. Galectin-3 was used as a negative control for specificity. Cosmc, T-synthase, Cosmc and T-Synthase, Galectin-3, Cosmc and Galactin-3 were incubated with Ni-NTA Superflow. Ni-NTA Superflow beads were washed and proteins were eluted and analyzed by Western blot with mAb to the HPC4, anti-Galectin-3, and anti-Cosmc. Cosmc interactions with T-synthase were determined (A and B). As a control, Cosmc and Galectin-3 interactions were determined (C and D). Black line indicates that intervening lanes have been spliced out. L, loading; U, unbound; B, bound.