High Affinity Dimerization by Ski Involves Parallel Pairing of a Novel Bipartite α-Helical Domain

c-Ski protein possesses a C-terminal dimerization domain that was deleted during the generation of v- ski , and has been implicated in the increased potency of c- ski in cellular transformation compared with the viral gene. The domain is predicted to consist of an extended α-helical segment made up of two motifs: a tandem repeat (TR) consisting of five imperfect repeats of 25 residues each and a leucine zipper (LZ) consisting of six heptad repeats. We have examined the structure and dimerization of TR or LZ individually or the entire TR-LZ domain. Using a quenched chemical cross-linking method, we show that the TR dimerizes with moderate efficiency ( K d = 4 × 10 −6 m ), whereas LZ dimerizes poorly ( K d > 2 × 10 −5 m ). However, the entire TR-LZ domain dimerizes efficiently ( K d = 2 × 10 −8 m ), showing a cooperative effect of the two motifs. CD analyses indicate that all three proteins contain predominantly α-helices. Limited proteolysis of the TR-LZ dimer indicates that the two helical motifs are linked by a small loop. Interchain disulfide bond formation indicates that both the LZ and TR helices are oriented in parallel. We propose a model for the dimer interface in the TR region consisting of discontinuous clusters of hydrophobic residues forming “leucine buttons.”