PDBsum entry 3bep

Transferase, transcription/DNA

PDB id: 3bep

Contents

Protein chains

366 a.a. *

DNA/RNA

Ligands

5CY

Waters ×604

* Residue conservation analysis

PDB id:

3bep

Name:

Transferase, transcription/DNA

Title:

Structure of a sliding clamp on DNA

Structure:

DNA (5'-d(p Dcp Dcp Dcp Dap Dtp Dcp Dgp Dtp Dap Dt)-3'). Chain: c. Engineered: yes. Other_details: DNA labeled at 5' end by dye cy5 (labeled as residue name 5cy). Only half of dye is visible in density. DNA (5'- d( Dtp Dtp Dtp Dtp Dap Dtp Dap Dcp Dgp Dap Dtp Dgp Dgp Dg)-3'). Chain: d. Engineered: yes.

Source:

Synthetic: yes. Escherichia coli. Organism_taxid: 562. Gene: dnan. Expressed in: escherichia coli bl21. Expression_system_taxid: 511693.

Resolution:

1.92Å R-factor: 0.210 R-free: 0.268

Authors:

R.E.Georgescu,S.S.Kim,O.Yurieva,J.Kuriyan,X.-P.Kong,M.O'Donnell

Key ref:

R.E.Georgescu et al. (2008). Structure of a sliding clamp on DNA. Cell, 132, 43-54. PubMed id: 18191219 DOI: 10.1016/j.cell.2007.11.045

Date:

19-Nov-07 Release date: 29-Jan-08

Protein chains

P0A988  (DPO3B_ECOLI) -  Beta sliding clamp from Escherichia coli (strain K12)

Seq: 366 a.a.

Struc: 366 a.a.

DNA/RNA chains

C-C-C-A-T-C-G-T-A-T 10 bases

T-T-T-T-A-T-A-C-G-A-T-G-G-G 14 bases

Enzyme reactions

• Enzyme class: E.C.2.7.7.7 - DNA-directed Dna polymerase.
• Reaction: DNA(n) + a 2'-deoxyribonucleoside 5'-triphosphate = DNA(n+1) + diphosphate

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

Added reference

DOI no: 10.1016/j.cell.2007.11.045

Cell 132:43-54 (2008)

PubMed id: 18191219

Structure of a sliding clamp on DNA.

R.E.Georgescu, S.S.Kim, O.Yurieva, J.Kuriyan, X.P.Kong, M.O'Donnell.

ABSTRACT

The structure of the E. coli beta clamp polymerase processivity factor has been solved in complex with primed DNA. Interestingly, the clamp directly binds the DNA duplex and also forms a crystal contact with the ssDNA template strand, which binds into the protein-binding pocket of the clamp. We demonstrate that these clamp-DNA interactions function in clamp loading, perhaps by inducing the ring to close around DNA. Clamp binding to template ssDNA may also serve to hold the clamp at a primed site after loading or during switching of multiple factors on the clamp. Remarkably, the DNA is highly tilted as it passes through the beta ring. The pronounced 22 degrees angle of DNA through beta may enable DNA to switch between multiple factors bound to a single clamp simply by alternating from one protomer of the ring to the other.

Selected figure(s)

Figure 3.
Figure 3. Structure of the β-DNA Complex
(A) Ribbon representation of the β-DNA complex: front and side views. DNA is tilted vert, similar 22° from the C2 rotation axis of β. The Cy5 moiety is not shown for clarity but is shown in Figure S4.
(B) Detailed view of R24 (top) and Q149 (bottom) in the β-DNA complex compared to the apo β structure (blue).
(C) Replication assays using primed M13 ssDNA coated with SSB contain Pol III^* and the indicated amount of either WT β (blue diamonds), β[Q149A] (red squares), β[R24A] (orange triangles), or β[R24A/Q149A] (green circles).
(D) Polymerase extension rate was determined with primed M13 ssDNA to which β, or mutant β, is first assembled on the DNA, followed by initiating synchronous chain extension. Reactions were quenched at the indicated times, and products were analyzed in a native agarose gel.
(E) The scheme illustrates the bead conjugated primed DNA in which SSB blocks ^32P-β from sliding off the end of the DNA. Clamp loading rate was assessed in assays with either β[WT] (blue), β[Q149A] (red), β[R24A] (orange), or β[R24A,Q149A] (green).

Figure 4.
Figure 4. Interaction of β with the ssDNA Region of the Primed Site
(A) Crystal lattice showing two molecules of the β-DNA complex. The ssDNA makes a crystal contact between two molecules of β.
(B) Surface representation of the charged residues that line the channel directing ssDNA to the hydrophobic protein-binding site of β. Basic residues are colored blue. The protein-binding site is shaded purple and subsites I and II (defined in Burnouf et al., 2004) are indicated.
(C) View of ssDNA (orange) positioned inside the hydrophobic protein-binding pocket of β. Thy[11] and Thy[12] occupy subsite I of the β hydrophobic pocket; the exposed R246, R240 side chains interact with the DNA phosphate backbone. Y153 and Y154 (green) stack with Ade[15] and Thy[13], respectively.

The above figures are reprinted from an Open Access publication published by Cell Press: Cell (2008, 132, 43-54) copyright 2008.

Figures were selected by an automated process.