File S1 - Structural Analysis of Intermolecular Interactions in the Kinesin Adaptor Complex Fasciculation and Elongation Protein Zeta 1/ Short Coiled-Coil Protein (FEZ1/SCOCO)

A single combined File S1 contains supplementary experimental procedures, five Figures S1 to S5, and Table S1. Figure S1 in File S1. Interaction between FEZ1 and SCOCO in an independent experiment. Purified recombinant proteins FEZ1 and SCOCO were incubated, chemically cross-linked, digested with trypsin, and analyzed by MS. MS/MS spectra were manually validated for b and y ion series of the α (peptide of FEZ1) and β (peptide of SCOCO) chains. The same peptides were found in two independent experiments. Figure S2 in File S1. Assay for β-galactosidase activity in yeast cells shows that FEZ1 (221-392) interacts with the coiled-coil region of SCOCO (2-82 and 2-65) and no interaction was detected with the empty vector pBTM116 (0). In the model, FEZ1 is shown in green and SCOCO in deep blue. The SCOCO region used in the two-hybrid assay is shown in light blue. The highlighted helix in cyan corresponds to the minimal interaction region in FEZ1/UNC-76 with SCOCO/UNC-69. Figure S3 in File S1. FEZ1-SCOCO docking. Relationship between binding energy and cross-linked lysine pair distance between FEZ1 and SCOCO. It was possible to obtain models from the docking routine that exhibit both optimal Cα-Cα distance between lysine pairs (>20 Å) and favorable binding energy (A). (B) General model of FEZ1-SCOCO in heterotetrameric fashion (ratio 2:2). FEZ1 is colored in green and SCOCO in deep blue. Figure S4 in File S1. Profile alignment of 20 templates for FEZ1 protein. Figure S5 in File S1. Per residue Z-score of FEZ1 protein modeling. Table S1 in File S1. Modeled loops in FEZ1 protein. (PDF)