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. Author manuscript; available in PMC: 2024 Nov 1.
Published in final edited form as: J Mol Biol. 2023 Sep 18;435(21):168283. doi: 10.1016/j.jmb.2023.168283

Figure 4. Kinetics of the tripartite complex formation.

Figure 4.

(A-D) BLI analysis of Isu1GST interaction with Ssq1* and Hsc20, and their variants. BLI sensors were loaded with Isu1GST or GST (background control). Association: loaded sensors were inserted into solutions containing Ssq1* and/or Hsc20. If two step was performed (B-D), 10 min later sensors were moved to a solution containing indicated combination of Ssq1* and/or Hsc20 for an additional 10 min. Dissociation phase was initiated by placing sensors into solutions without proteins. (A) One step association analysis of Ssq1* and Ssq1*/Hsc20 using Isu1GST and, as a control, GST. (B) Two step analysis of Isu1GST interaction with Ssq1* and Hsc20, assessing effect of order of addition of Hsc20 and Ssq1*. (C) Two step analysis of interaction of Isu1GST with Ssq1* and indicated variants defective in Ssq1* interaction: Hsc20 (R37A or D50A); Isu1LLY (LLY). (D) Two step analysis of Isu1GST interaction with Hsc20 and Ssq1* variant (F426S) defective in interaction with Isu1.