Ssn6, a yeast protein that comprises 10 tandem tetratricopeptide repeat (TPR) motifs, associates with Tup1 repressor protein and acts as a transcriptional corepressor. In this report we identify point mutations in the TPR1 of Ssn6 that disrupt Tup1 interaction. Furthermore, we construct a 3D model of the TPR domain of Ssn6, which is responsible for Tup1 binding, based on the known structure of protein phosphatase 5. According to this model all selected mutations reduce the ability of Ssn6 to interact with Tup1 by affecting the structural integrity of TPR1 and/or the correct spatial arrangement of TPR1 relative to TPR2 and TPR3.