PDBsum entry 1owr

Transcription/DNA

PDB id: 1owr

Contents

Protein chains

284 a.a. *

DNA/RNA

* Residue conservation analysis

PDB id:

1owr

Name:

Transcription/DNA

Title:

Crystal structure of human nfat1 bound monomerically to DNA

Structure:

Nfat1 monomeric binding site, plus strand. Chain: a, c, e, g. Engineered: yes. Nfat1 monomeric binding site, minus strand. Chain: b, d, f, h. Engineered: yes. Nuclear factor of activated t-cells, cytoplasmic 2. Chain: m, n, p, q. Synonym: t cell transcription factor nfat1, nfat pre-existing

Source:

Synthetic: yes. Other_details: the plus strand of the nfat1 monomeric binding site was chemically synthesized. Other_details: the minus strand of the nfat1 monomeric binding site homo sapiens. Human. Organism_taxid: 9606. Gene: nfatc2 or nfat1 or nfatp. Expressed in: escherichia coli.

Biol. unit:

Trimer (from PQS)

Resolution:

3.00Å R-factor: 0.241 R-free: 0.273

Authors:

J.C.Stroud,L.Chen

Key ref:

J.C.Stroud and L.Chen (2003). Structure of NFAT bound to DNA as a monomer. J Mol Biol, 334, 1009-1022. PubMed id: 14643663 DOI: 10.1016/j.jmb.2003.09.065

Date:

29-Mar-03 Release date: 10-Feb-04

Protein chains

Q13469  (NFAC2_HUMAN) -  Nuclear factor of activated T-cells, cytoplasmic 2 from Homo sapiens

Seq: 925 a.a.

Struc: 284 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

DNA/RNA chains

T-T-G-C-T-G-G-A-A-A-A-A-T-A-G 15 bases

A-A-C-T-A-T-T-T-T-T-C-C-A-G-C 15 bases

T-T-G-C-T-G-G-A-A-A-A-A-T-A-G 15 bases

A-A-C-T-A-T-T-T-T-T-C-C-A-G-C 15 bases

T-T-G-C-T-G-G-A-A-A-A-A-T-A-G 15 bases

A-A-C-T-A-T-T-T-T-T-C-C-A-G-C 15 bases

T-T-G-C-T-G-G-A-A-A-A-A-T-A-G 15 bases

A-A-C-T-A-T-T-T-T-T-C-C-A-G-C 15 bases

Enzyme reactions

• Enzyme class: E.C.?

DOI no: 10.1016/j.jmb.2003.09.065

J Mol Biol 334:1009-1022 (2003)

PubMed id: 14643663

Structure of NFAT bound to DNA as a monomer.

J.C.Stroud, L.Chen.

ABSTRACT

The nuclear factor of activated T cells (NFAT) is a calcium-dependent transcription factor that cooperates with a myriad of partner transcription factors to regulate distinct transcription programs. Transcription activation by NFAT without the cooperation of co-stimulatory signals in lymphocytes can also impose a genetic program of anergy. Although the ternary NFAT1/Fos-Jun/DNA complex has been structurally characterized, how NFAT1 recognizes DNA in the absence of cooperative partners and how such a binary NFAT/DNA complex may lead to the assembly of distinct high-order NFAT transcription complexes are still poorly understood. We have determined the crystal structure of the entire Rel homology region (RHR) of human NFAT1 (NFATc2) bound to DNA as a monomer. We also present footprinting evidence that corroborates the protein-DNA contacts observed in the crystal structure. Our structural and biochemical studies reveal the mechanism by which the monomeric Rel protein NFAT recognizes its cognate DNA site. A remarkable feature of the binary NFAT/DNA complex is the conformational flexibility exhibited by NFAT1 in the four independent copies of the NFAT/DNA complex in the crystal structure, which may reflect a mechanism by which NFAT1 interacts with a variety of protein partners as it mediates disparate biological responses.

Selected figure(s)

Figure 2.
Figure 2. Interactions between the RHR-N and RHR-C in Complexes 1 and 2. (a) Comparison of Complexes 1 and 2 with the NFAT/Fos-Jun/DNA ternary complex. All complexes have been oriented for comparison looking down the DNA axis with the RHR-C projecting to the left. Fos is colored red and Jun is colored blue. (b) Detailed view of the D464-R541-Q669 triad from Complexes 1 and 2 and the ternary complex. This triad is representative of the malleability seen in the RHR-N/RHR-C interface. Orientations are similar to (a) except for some minor changes for clarity.

Figure 4.
Figure 4. EMSA and footprinting of the binary and ternary complexes in solution. (a) Titration of full-length NFAT RHR and the NFAT RHR-N with the DNA used for crystallographic analysis. Left panel, binding curve of full-length NFAT RHR binding to the DNA fragment (K[d]=2.7(±0.4) nM, see Methods). Center panel, EMSA gel of full-length NFAT RHR binding to the DNA fragment. Right panel, EMSA gel of the NFAT RHR-N binding to the DNA fragment. (b) Methylation interference footprinting of NFAT1 and NFAT1/Fos-Jun binding to ARRE2. F, free (unbound) DNA; L, lower complex containing NFAT1 alone; U, upper complex containing NFAT1 and Fos-Jun. Lanes 1-3, coding strand; 4-6, non-coding strand. Fiducial lines connect certain residues to their corresponding bands for clarity. (c) Ethyl nitroso-urea (ENU) ethylation interference footprinting of NFAT1 and NFAT1/Fos-Jun binding to ARRE2. F, free (unbound) DNA; L, lower complex containing NFAT1 alone; U, upper complex containing NFAT1 and Fos-Jun; G, (lanes 1 and 11) Maxam-Gilbert G lane; lanes 2-4, coding strand using the full-length NFAT RHR; lanes 5-7, non-coding strand using only the NFAT RHR-N; lanes 8-10, non-coding strand using the full-length NFAT RHR. Fiducial lines connect certain residues to their corresponding bands for clarity. On the right of the gel is a histogram showing quantitative analysis of the footprints on the non-coding strand for the NFAT1 RHR/DNA complex (open) and NFAT1 RHR/Fos-Jun/DNA complex (shaded). Values are expressed as fractions of the intensity at the same positions in the free probe after normalization for loading.

The above figures are reprinted by permission from Elsevier: J Mol Biol (2003, 334, 1009-1022) copyright 2003.

Figures were selected by an automated process.