Secondary structure assignment of mouse SOCS3 by NMR defines the domain boundaries and identifies an unstructured insertion in the SH2 domain.
SOCS3 is a negative regulator of cytokine signalling that inhibits Janus kinase-signal transduction and activator of transcription (JAK-STAT) mediated signal tranduction by binding to phosphorylated tyrosine residues on intracellular subunits of various cytokine receptors, as well as possibly the JAK proteins. SOCS3 consists of a short N-terminal sequence followed by a kinase inhibitory region, an extended SH2 domain and a C-terminal suppressor of cytokine signalling (SOCS) box. SOCS3 and the related protein, cytokine-inducible SH2-containing protein, are unique among the SOCS family of proteins in containing a region of mostly low complexity sequence, between the SH2 domain and the C-terminal SOCS box. Using NMR, we assigned and determined the secondary structure of a murine SOCS3 construct. The SH2 domain, unusually, consists of 140 residues, including an unstructured insertion of 35 residues. This insertion fits the criteria for a PEST sequence and is not required for phosphotyrosine binding, as shown by isothermal titration calorimetry. Instead, we propose that the PEST sequence has a functional role unrelated to phosphotyrosine binding, possibly mediating efficient proteolytic degradation of the protein. The latter half of the kinase inhibitory region and the entire extended SH2 subdomain form a single alpha-helix. The mapping of the true SH2 domain, and the location of its C terminus more than 50 residues further downstream than predicted by sequence homology, explains a number of previously unexpected results that have shown the importance of residues close to the SOCS box for phosphotyrosine binding.
Babon
Jeffrey J
JJ
Walter and Eliza Hall Institute, Parkville, Victoria, Australia. babon@wehi.edu.au
Yao
Shenggen
S
DeSouza
David P
DP
Harrison
Christopher F
CF
Fabri
Louis J
LJ
Liepinsh
Edvards
E
Scrofani
Sergio D
SD
Baca
Manuel
M
Norton
Raymond S
RS
D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D019000:Phosphotyrosine / Q000378:metabolism; D017433:Protein Structure, Secondary; D016415:Sequence Alignment; D015398:Signal Transduction / Q000502:physiology; D000071223:Suppressor of Cytokine Signaling 3 Protein; D050826:Suppressor of Cytokine Signaling Proteins / Q000737:chemistry* / Q000378:metabolism; D018909:src Homology Domains
C491411:Socs3 protein, mouse; D000071223:Suppressor of Cytokine Signaling 3 Protein; D050826:Suppressor of Cytokine Signaling Proteins; D019000:Phosphotyrosine