PDBsum entry: 6gat

Transcription/DNA

PDB id: 6gat

Contents

Protein chain

66 a.a. *

DNA/RNA

Metals

_ZN

* Residue conservation analysis

PDB id:

6gat

Name:

Transcription/DNA

Title:

Solution nmr structure of the l22v mutant DNA binding domain of area complexed to a 13 bp DNA containing a tgata site, regularized mean structure

Structure:

DNA (5'- d( Cp Ap Gp Tp Gp Ap Tp Ap Gp Ap Gp Ap C)-3'). Chain: b. Engineered: yes. DNA (5'- d( Gp Tp Cp Tp Cp Tp Ap Tp Cp Ap Cp Tp G)-3'). Chain: c. Engineered: yes. Nitrogen regulatory protein area.

Source:

Synthetic: yes. Emericella nidulans. Organism_taxid: 162425. Gene: potential. Expressed in: escherichia coli. Expression_system_taxid: 562

NMR struc:

1 models

Authors:

G.M.Clore,M.Starich,M.Wikstrom,A.M.Gronenborn

Key ref:

M.R.Starich et al. (1998). The solution structure of the Leu22-->Val mutant AREA DNA binding domain complexed with a TGATAG core element defines a role for hydrophobic packing in the determination of specificity. J Mol Biol, 277, 621-634. PubMed id: 9533884 DOI: 10.1006/jmbi.1997.1626

Date:

07-Nov-97 Release date: 28-Jan-98

Protein chain

P17429  (AREA_EMENI) -  Nitrogen regulatory protein areA

Seq: 876 a.a.

Struc: 66 a.a.*

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

 Gene Ontology (GO) functional annotation 

• Biological process: regulation of transcription, DNA-dependent (1 term)
• Biochemical function: transcription factor activity (3 terms)

DOI no: 10.1006/jmbi.1997.1626

J Mol Biol 277:621-634 (1998)

PubMed id: 9533884

The solution structure of the Leu22-->Val mutant AREA DNA binding domain complexed with a TGATAG core element defines a role for hydrophobic packing in the determination of specificity.

M.R.Starich, M.Wikström, S.Schumacher, H.N.Arst, A.M.Gronenborn, G.M.Clore.

ABSTRACT

The seemingly innocuous leucine-to-valine mutation at position 22 of the AREA DNA binding domain results in dramatic changes in the in vivo expression profile of genes controlled by this GATA transcription factor. This is associated with a preference of the Leu22-->Val mutant for TGATAG sites over (A/C)GATAG sites. Quantitative gel retardation assays confirm this observation and show that the Leu22-->Val mutant AREA DNA binding domain has a approximately 30-fold lower affinity than the wild-type domain for a 13 base-pair oligonucleotide containing the wild-type CGATAG target. To gain insight into the measured affinity data and further explore sequence specificity of the AREA protein, the solution structure of a complex between the Leu22-->Val mutant AREA DNA binding domain and a 13 base-pair oligonucleotide containing its physiologically relevant TGATAG target sequence has been determined by multidimensional nuclear magnetic resonance spectroscopy. Comparison of this structure with that of the wild-type AREA DNA binding domain complexed to its cognate CGATAG target site shows how subtle changes in amino acid side-chain length and hydrophobic packing can affect affinity and specificity for GATA-containing sequences, and how changes in DNA sequence can be compensated for by changes in protein sequence.

Selected figure(s)

Figure 5.
Figure 5. Two views showing a comparison of the wtAREA (red worm) and Leu22→Val mutAREA (gold worm) DBDs complexed to DNA. Residues 10 to 61 of both domains are superimposed in the views shown. The surfaces representation of only one of the DNA models (the T GATAG site for the mutant complex) is shown for clarity. The surfaces of the bases in the major groove are colored in light blue while those in the minor groove are in light red. Side-chains for the wtAREA and mutAREA DBDs are shown in blue and green, respectively. The zinc atoms and their coordinating ligands are shown in purple (wtAREA) and pink (mutAREA). The coordinates of the wtAREA DBD complex are taken from [Starich et al 1998].

Figure 6.
Figure 6. Comparison of the Leu22→Val mutAREA DBD·T GATAG and wtAREA DBD·C GATAG complexes. A, Superposition of the protein backbone and DNA for the wild-type and mutant AREA DBD complexes. The protein backbones are depicted as red (wtAREA) and gold (mutAREA) worms. Bonds between heavy atoms of the DNA (base-pairs 3 to 11) are represented as light red (CGATA site) or tan (TGATA site) sticks. B, Stereoview showing protein-DNA interactions for residues at positions 22 and 24. The wtAREA DBD backbone and side-chains are shown in red and blue, respectively, while the mutAREA DBD backbone and side-chains are shown in gold and green, respectively. The G3-C4-G5 element of the C GATAG site is shown in light red, and the G3-T4-G5 element of the T GATAG site is shown in tan. Only bonds between heavy atoms for the protein and DNA are represented with the exception of C---H bonds for all methyl groups shown (Leu22 δs, Val22 γs and the methyl group of T4). The coordinates of the wtAREA DBD complex are taken from [Starich et al 1998].

The above figures are reprinted by permission from Elsevier: J Mol Biol (1998, 277, 621-634) copyright 1998.

Figures were selected by the author.