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. 2007 Nov 15;26(24):4935–4945. doi: 10.1038/sj.emboj.7601915

Figure 1.

Figure 1

TEV protease-mediated Nyv1p inactivation prevents homotypic yeast vacuole fusion in vitro. (A) Schematic representation of wild-type Nyv1p and Nyv1p-TEV. (B) TEV protease specifically inactivates Nyv1p-TEV vacuoles for fusion. BJ3505 (NYV1) and DKY6281 (NYV1) vacuoles or BJ3505 NYV1-TEV and DKY6281 NYV1-TEV vacuoles were incubated at 27°C in fusion reactions (see Materials and methods) in the presence of indicated concentrations of TEV protease. After 90 min, a portion was assayed for fusion (B), and the rest centrifuged to sediment vacuoles, which were resuspended in SDS sample buffer and analyzed by SDS–PAGE and immunoblotting (C). (C) Fusion inactivation by TEV protease correlates with the removal of Nyv1p-TEV from vacuoles. The anti-Pep4p blot serves as a loading control. Data are mean±s.e.m. (n=3).