PDBsum entry: 1g88

Transcription

PDB id: 1g88

Contents

Protein chains

221 a.a. *

242 a.a. *

* Residue conservation analysis

PDB id:

1g88

Name:

Transcription

Title:

S4afl3arg515 mutant

Structure:

Smad4. Chain: a, b, c. Fragment: smad4 active fragment. Synonym: dpc4. Mutation: yes

Source:

Homo sapiens. Human. Organism_taxid: 9606

Biol. unit:

Trimer (from PQS)

Resolution:

3.00Å R-factor: 0.203 R-free: 0.263

Authors:

B.M.Chako,B.Qin,S.S.Lam,J.J.Correia,K.Lin

Key ref:

B.M.Chacko et al. (2001). The L3 loop and C-terminal phosphorylation jointly define Smad protein trimerization. Nat Struct Biol, 8, 248-253. PubMed id: 11224571 DOI: 10.1038/84995

Date:

16-Nov-00 Release date: 29-Nov-00

Protein chain

Q13485  (SMAD4_HUMAN) -  Mothers against decapentaplegic homolog 4

Seq: 552 a.a.

Struc: 221 a.a.*

Protein chains

Q13485  (SMAD4_HUMAN) -  Mothers against decapentaplegic homolog 4

Seq: 552 a.a.

Struc: 242 a.a.*

* PDB and UniProt seqs differ at 2 residue positions (black crosses)

 Gene Ontology (GO) functional annotation 

• Cellular component: intracellular (1 term)
• Biological process: regulation of transcription, DNA-dependent (1 term)

DOI no: 10.1038/84995

Nat Struct Biol 8:248-253 (2001)

PubMed id: 11224571

The L3 loop and C-terminal phosphorylation jointly define Smad protein trimerization.

B.M.Chacko, B.Qin, J.J.Correia, S.S.Lam, M.P.de Caestecker, K.Lin.

ABSTRACT

Smad proteins mediate the transforming growth factor beta responses. C-terminal phosphorylation of R-Smads leads to the recruitment of Smad4 and the formation of active signaling complexes. We investigated the mechanism of phosphorylation-induced Smad complex formation with an activating pseudo-phosphorylated Smad3. Pseudo-phosphorylated Smad3 has a greater propensity to homotrimerize, and recruits Smad4 to form a heterotrimer containing two Smad3 and one Smad4. The trimeric interaction is mediated through conserved interfaces to which tumorigenic mutations map. Furthermore, a conserved Arg residue within the L3 loop, located near the C-terminal phosphorylation sites of the neighboring subunit, is essential for trimerization. We propose that the phosphorylated C-terminal residues interact with the L3 loop of the neighboring subunit to stabilize the trimer interaction.

Selected figure(s)

Figure 3.
Figure 3. The sulfate binding site within the L3 loop is critical for Smad heteromeric interaction. a, The sulfate binding sites shown in detail. The location of the sulfate ions in the context of the trimeric S4AF structure is shown in Fig. 2b. b, Mutation of the sulfate binding site at the L3 loop specifically reduces heteromeric interaction between S3LC(3E) and S4AF. S3LC(3E) and the sulfate binding site mutants of S4AF were mixed at a 1:1 mole ratio and loaded on the size exclusion column. The eluted fractions were analyzed by SDS-PAGE and the gels were stained with Coomassie blue. c, Stereo view of the F[o] - F[c] omit map at the L3 loop of the S4AF(R515S) mutant. The mutant and wild type coordinates are shown in black and gray, respectively. d, The Smad4 Arg 515 mutant only weakly activates Smad3/4 dependent transcriptional responses. Smad4 null MDA-MB468 cells were transfected with SBE-Lux along with the indicated FLAG-Smad3(3E) and Smad4-Myc mutant constructs. e, Homotrimerization of S3LC(3E) is abolished by mutation of residues at the trimer interface or the conserved Arg residue within the L3 loop. Proteins (50 M) were loaded onto the size-exclusion column and the eluted fractions were analyzed by SDS-PAGE and the gels were stained by Coomassie-Blue.

Figure 4.
Figure 4. Proposed model of phosphorylation induced Smad protein activation. Left, phosphorylation induced heterotrimer between Smad4 and R-Smad subunits. Right, phosphorylation induced homotrimer of R-Smad. The Smad4 subunit is shown in pink. The R-Smad subunits are shown in blue. The conserved L3 loops are shown in black. The phosphorylated C-terminal tails of the R-Smad subunits are represented by red arrows.

The above figures are reprinted by permission from Macmillan Publishers Ltd: Nat Struct Biol (2001, 8, 248-253) copyright 2001.

Figures were selected by an automated process.