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 x 10^-6 M), whereas LZ dimerizes poorly (K(d) > 2 x 10^-5 M). However, the entire TR-LZ domain dimerizes efficiently (K(d) = 2 x 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'.