High affinity dimerization by Ski involves parallel pairing of a novel bipartite alpha-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 alpha-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 (Kd = 4 x 10(-6) M), whereas LZ dimerizes poorly (Kd > 2 x 10(-5) M). However, the entire TR-LZ domain dimerizes efficiently (Kd = 2 x 10(-8) M), showing a cooperative effect of the two motifs. CD analyses indicate that all three proteins contain predominantly alpha-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."
Zheng
G
G
Department of Biochemistry, Case Western Reserve University, School of Medicine, Cleveland, Ohio 44106-4935, USA.
Blumenthal
K M
KM
Ji
Y
Y
Shardy
D L
DL
Cohen
S B
SB
Stavnezer
E
E
D000595:Amino Acid Sequence; D000818:Animals; D002645:Chickens; D002942:Circular Dichroism; D004268:DNA-Binding Proteins / Q000737:chemistry*; D019281:Dimerization; D008969:Molecular Sequence Data; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D011518:Proto-Oncogene Proteins / Q000737:chemistry*
D004268:DNA-Binding Proteins; D011518:Proto-Oncogene Proteins