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...

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Bibliographic Details
Published in:The Journal of biological chemistry 1997-12, Vol.272 (50), p.31855-31864
Main Authors: Zheng, Guoxing, Blumenthal, Kenneth M., Ji, Yonggang, Shardy, Deborah L., Cohen, Steven B., Stavnezer, Edward
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Animals
Chickens
Circular Dichroism
Dimerization
DNA-Binding Proteins - chemistry
Molecular Sequence Data
Protein Conformation
Protein Structure, Secondary
Proto-Oncogene Proteins - chemistry
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Summary: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 (Kd = 4 × 10−6m), whereas LZ dimerizes poorly (Kd > 2 × 10−5m). However, the entire TR-LZ domain dimerizes efficiently (Kd = 2 × 10−8m), 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.”
ISSN:0021-9258
1083-351X
DOI:10.1074/jbc.272.50.31855