PDBsum entry: 1ckt

Gene regulation/DNA

PDB id: 1ckt

Contents

Protein chain

71 a.a. *

DNA/RNA

Ligands

CPT

Waters ×74

* Residue conservation analysis

PDB id:

1ckt

Name:

Gene regulation/DNA

Title:

Crystal structure of hmg1 domain a bound to a cisplatin-modi duplex

Structure:

DNA (5'-d( Cp Cp (5Iu) p Cp Tp Cp Tp Gp Gp Ap Cp Cp Tp Tp Cp C)-3'). Chain: b. Engineered: yes. DNA (5'-d( Gp Gp Ap Ap Gp Gp Tp Cp Cp Ap Gp Ap Gp 3'). Chain: c. Engineered: yes. High mobility group 1 protein.

Source:

Synthetic: yes. Rattus norvegicus. Norway rat. Organism_taxid: 10116. Strain: soragye-dawley. Tissue: liver. Expressed in: escherichia coli bl21(de3). Expression_system_taxid: 469008.

Biol. unit:

Trimer (from PQS)

Resolution:

2.50Å R-factor: 0.238 R-free: 0.239

Authors:

U.-M.Ohndorf,M.A.Rould,C.O.Pabo,S.J.Lippard

Key ref:

U.M.Ohndorf et al. (1999). Basis for recognition of cisplatin-modified DNA by high-mobility-group proteins. Nature, 399, 708-712. PubMed id: 10385126 DOI: 10.1038/21460

Date:

23-Apr-99 Release date: 30-Jun-99

Protein chain

P63159  (HMGB1_RAT) -  High mobility group protein B1

Seq: 215 a.a.

Struc: 71 a.a.

 Gene Ontology (GO) functional annotation 

• Cellular component: nucleus (1 term)
• Biochemical function: protein binding (2 terms)

DOI no: 10.1038/21460

Nature 399:708-712 (1999)

PubMed id: 10385126

Basis for recognition of cisplatin-modified DNA by high-mobility-group proteins.

U.M.Ohndorf, M.A.Rould, Q.He, C.O.Pabo, S.J.Lippard.

ABSTRACT

The anticancer activity of cis-diamminedichloroplatinum(II) (cisplatin) arises from its ability to damage DNA, with the major adducts formed being intrastrand d(GpG) and d(ApG) crosslinks. These crosslinks bend and unwind the duplex, and the altered structure attracts high-mobility-group domain (HMG) and other proteins. This binding of HMG-domain proteins to cisplatin-modified DNA has been postulated to mediate the antitumour properties of the drug. Many HMG-domain proteins recognize altered DNA structures such as four-way junctions and cisplatin-modified DNA, but until now the molecular basis for this recognition was unknown. Here we describe mutagenesis, hydroxyl-radical footprinting and X-ray studies that elucidate the structure of a 1:1 cisplatin-modified DNA/HMG-domain complex. Domain A of the structure-specific HMG-domain protein HMG1 binds to the widened minor groove of a 16-base-pair DNA duplex containing a adduct. The DNA is strongly kinked at a hydrophobic notch created at the platinum-DNA crosslink and protein binding extends exclusively to the 3' side of the platinated strand. A phenylalanine residue at position 37 intercalates into a hydrophobic notch created at the platinum crosslinked d(GpG) site and binding of the domain is dramatically reduced in a mutant in which alanine is substituted for phenylalanine at this position.

Selected figure(s)

Figure 1.
Figure 1: Primary structure and features of the complex between the non-sequence-specific domain A of HMG1 and cisplatin-modified DNA. a, Sequence of DNA used here; asterisks, nucleotides crosslinked by cisplatin. Nucleotides contacted by the protein are in bold type. b, Alignment of selected members of the two subfamilies of HMG-domain proteins^30, based on primary sequence and structural superposition. Shaded boxes represent -helices and solid lines indicate loops. The top group are a representative set of structure-specific HMG domains and the bottom group are sequence-specific proteins. The HMG1 domain A sequence and numbering shown correspond to the residues located and refined in the crystal structure. Residues that contact DNA in structurally characterized complexes are highlighted. Amino acids at positions equivalent to residue 37 of HMG1 are boxed. c, A simulated annealing omit map (starting temperature, 2,000 K), contoured at the 1.1 level, in the region of the cisplatin–DNA adduct and showing the localized bend at the platination site and the intercalated Phe 37 side chain. For computing this (F[o] - F[c]) electron density map, residues 36–38 and bases T[7]–G[9], C[24]–A[26] were omitted. d, Overall structure of the complex. The protein backbone is shown in yellow, the intercalating Phe 37 residue as van der Waals spheres, and the DNA in red and blue with the cis -[Pt(NH[3])[2]{d(GpG)-N7(G[8]),-N7(G[9])}] intrastrand adduct in green. Numbers indicate the first (N terminus) and last (C terminus) ordered residues in the crystal structure. Main-chain torsion angles for all non-glycine residues fall within energetically favourable Ramachandran boundaries.

Figure 3.
Figure 3: The platinum–DNA crosslink and protein–DNA interactions. a, Diagram summarizing all HMG domain–DNA contacts. Solid lines, hydrogen bonds or salt bridges; dashed lines, van der Waals contacts; mc, main chain; and w, water molecule. The wedge indicates the Phe-37 intercalation site. b, Expanded view ofprotein–DNA interactions at the site of platination. The aromatic rings of Phe 37and G[8] pack edge-to-face with a dihedral angle of 74°. The main-chain carbonyl group of Phe 37 is within hydrogen-bonding distance (2.9 Å) of the exocyclic amino group of G[9]. As predicted^7, Ser 41 is involved in a very tight hydrogen-bonding contact with N3 of A[10]. c, Superposition of crystallographically determined cisplatin–{d(GpG)} intrastrand crosslinks in the present structure (yellow), the equivalent site in the structure of a single-stranded stranded cis -[Pt(NH[3])[2]{d(pGpG)}] platinated dinucleotide^17 (blue, left), and the site in a cisplatin-modified DNA dodecamer duplex^14 (red, right), portraying dihedral angles between the two coordinated guanine bases of 75°, 77° and 30°, respectively, as calculated by the program QUANTA 4.1 (Molecular Simulations).

The above figures are reprinted by permission from Macmillan Publishers Ltd: Nature (1999, 399, 708-712) copyright 1999.

Figures were selected by the author.