PDBsum entry 1dh3

Transcription/DNA

PDB id: 1dh3

Contents

Protein chains

55 a.a. *

DNA/RNA

Metals

_MG

Waters ×17

* Residue conservation analysis

PDB id:

1dh3

Name:

Transcription/DNA

Title:

Crystal structure of a creb bzip-cre complex reveals the basis for creb faimly selective dimerization and DNA binding

Structure:

DNA (5'- d( Cp Cp Tp Tp Gp Gp Cp Tp Gp Ap Cp Gp Tp Cp Ap Gp Cp Cp Ap Ap G)- 3'). Chain: b, d. Engineered: yes. Transcription factor creb. Chain: a, c. Fragment: residues 201-255. Engineered: yes

Source:

Synthetic: yes. Mus musculus. House mouse. Organism_taxid: 10090. Expressed in: escherichia coli. Expression_system_taxid: 562.

Biol. unit:

Tetramer (from PQS)

Resolution:

3.00Å R-factor: 0.218 R-free: 0.280

Authors:

M.A.Schumacher,R.H.Goodman,R.G.Brennan

Key ref:

M.A.Schumacher et al. (2000). The structure of a CREB bZIP.somatostatin CRE complex reveals the basis for selective dimerization and divalent cation-enhanced DNA binding. J Biol Chem, 275, 35242-35247. PubMed id: 10952992 DOI: 10.1074/jbc.M007293200

Date:

27-Nov-99 Release date: 27-Nov-00

Protein chains

Q01147  (CREB1_MOUSE) -  Cyclic AMP-responsive element-binding protein 1 from Mus musculus

Seq: 327 a.a.

Struc: 55 a.a.*

* PDB and UniProt seqs differ at 3 residue positions (black crosses)

DNA/RNA chains

C-C-T-T-G-G-C-T-G-A-C-G-T-C-A-G-C-C-A-A-G 21 bases

C-C-T-T-G-G-C-T-G-A-C-G-T-C-A-G-C-C-A-A-G 21 bases

Enzyme reactions

• Enzyme class: E.C.?

DOI no: 10.1074/jbc.M007293200

J Biol Chem 275:35242-35247 (2000)

PubMed id: 10952992

The structure of a CREB bZIP.somatostatin CRE complex reveals the basis for selective dimerization and divalent cation-enhanced DNA binding.

M.A.Schumacher, R.H.Goodman, R.G.Brennan.

ABSTRACT

The cAMP responsive element-binding protein (CREB) is central to second messenger regulated transcription. To elucidate the structural mechanisms of DNA binding and selective dimerization of CREB, we determined to 3.0 A resolution, the structure of the CREB bZIP (residues 283-341) bound to a 21-base pair deoxynucleotide that encompasses the canonical 8-base pair somatostatin cAMP response element (SSCRE). The CREB dimer is stabilized in part by ionic interactions from Arg(314) to Glu(319') and Glu(328) to Lys(333') as well as a hydrogen bond network that links the carboxamide side chains of Gln(322')-Asn(321)-Asn(321')-Gln(322). Critical to family selective dimerization are intersubunit hydrogen bonds between basic region residue Tyr(307) and leucine zipper residue Glu(312), which are conserved in all CREB/CREM/ATF-1 family members. Strikingly, the structure reveals a hexahydrated Mg(2+) ion bound in the cavity between the basic region and SSCRE that makes a water-mediated DNA contact. DNA binding studies demonstrate that Mg(2+) ions enhance CREB bZIP:SSCRE binding by more than 25-fold and suggest a possible physiological role for this ion in somatostatin cAMP response element and potentially other CRE-mediated gene expression.

Selected figure(s)

Figure 3.
Fig. 3. Electron density maps of the CREB bZIP divalent cation-binding pocket. A, (2F[o] F[c]) omit electron density map (gray wire mesh) contoured at 1 and calculated with phases from the final model in which residues 307 to 321 of each monomer, the hexahydrated magnesium ion and the water molecules involved in DNA binding were deleted and followed by xyzb refinement until convergence, which removes phase bias. Labeled are the hexahydrated magnesium ion, residues Tyr307 and Glu312', and Lys304 and Arg301 from each subunit. B, (F[o] F[c]) omit electron densities using coefficients F[o](BaCl[2]) F[o](MgCl[2]) (contoured at 6 and displayed as red wire mesh) or F[o](Na[2]WO[4]) F[o]((NH[4])[2]SO[4]) (contoured at 4 and displayed as blue wire mesh) confirming unequivocally the identity of the density in A to be a divalent cation. The slight off-centering of the Ba^2+ ion is very likely the result of its preferred hepta aquo coordination versus the hexa aquo coordination of a Mg2+ ion. A and B were generated with O (15).

Figure 4.
Fig. 4. CREB-DNA affinity and its divalent cation enhancement. A, representative fluorescence anisotropy-derived DNA binding isotherm of the CREB bZIP-SSCRE measured in the presence of 10 m M MgCl[2] (diamonds) or 10 mM BaCl[2] (boxes) in buffer A or buffer A alone (circles). Plotted are the normalized measured millipolarization units (mP) versus the CREB bZIP concentration (nM). B, representative divalent cation binding isotherm. Measurements were made by titrating CaCl[2] into a 0.990-ml reaction buffer of 25 mM Tris, pH 7.5, 5% glycerol, 50 mM NaCl, 6 µg of bovine serum albumin, 10 µg of poly d(I-C) containing 1 nM fluoresceinated DNA and 20 nM CREB bZIP. Plotted are the measured millipolarization units (mP) versus the Ca^2+ ion concentration. Essentially identical binding isotherms are generated by titration with MgCl[2] or BaCl[2]. C, CREB bZIP:SSCRE phosphate contacts. Residues making phosphate contacts are shown as light blue sticks and labeled in black. The DNA is shown as sticks. The carbon atoms are colored white, oxygen atoms are red, nitrogen atoms are blue, and phosphate atoms are shown as yellow. Arg289, which does not make any direct phosphate contacts, is also shown. This residue is in position to make potential water-mediated base contacts to bases outside the consensus SSCRE or, alternatively, may contribute to binding through a general electrostatic effect. The two identical DNA strands are differentiated as strand 1 (S1) and strand 2 (S2) to distinguish cross-strand phosphate contacts made by each CREB bZIP subunit. D, specific interactions between CREB and the SSCRE DNA. Only DNA base pairs 1 to 4 of the consensus SSCRE, are shown. DNA atoms are colored as in C. CREB residues involved in base specific contacts are shown as light blue sticks. Only residue Arg301' from the second subunit is shown as the remaining base-specific contacts are identical between subunits. Two water molecules (W1 and W2) and the hexahydrated magnesium ion are shown as ball-and-sticks. Oxygen atoms are colored red and the magnesium atom is colored light blue. Hydrogen bonds are indicated by black dashed lines and hydrophobic contacts by light blue dashed lines.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2000, 275, 35242-35247) copyright 2000.

Figures were selected by an automated process.