PDBsum entry 4xqk

Hydrolase/DNA

PDB id: 4xqk

Contents

Protein chains

1517 a.a.

DNA/RNA

Metals

__K ×3

Waters ×148

PDB id:

4xqk

Name:

Hydrolase/DNA

Title:

Atp-dependent type isp restriction-modification enzyme llabiii bound to DNA

Structure:

Llabiii. Chain: a, b. Engineered: yes. DNA (28-mer). Chain: c, e. Engineered: yes. DNA (28-mer). Chain: d, f. Engineered: yes

Source:

Lactococcus lactis subsp. Cremoris. Organism_taxid: 1359. Strain: w10. Expressed in: escherichia coli. Expression_system_taxid: 562. Synthetic: yes. Synthetic construct. Organism_taxid: 32630. Organism_taxid: 32630

Resolution:

2.70Å R-factor: 0.221 R-free: 0.260

Authors:

M.K.Chand,K.Saikrishnan

Key ref:

M.K.Chand et al. (2015). Translocation-coupled DNA cleavage by the Type ISP restriction-modification enzymes. Nat Chem Biol, 11, 870-877. PubMed id: 26389736 DOI: 10.1038/nchembio.1926

Date:

19-Jan-15 Release date: 16-Sep-15

Protein chains

A0A0M3KL05  (A0A0M3KL05_LACLC) -  LlaBIII from Lactococcus lactis subsp. cremoris

Seq: 1578 a.a.

Struc: 1517 a.a.

DNA/RNA chains

C-A-C-C-T-C-G-G-C-T-C-A-G-T-C-T-A-T-T-A-G-C-T-A-G-A-G-C 28 bases

G-C-T-C-T-A-G-C-T-A-A-T-A-G-A-C-T-G-A-G-C-C-G-A-G-G-T-G 28 bases

C-A-C-C-T-C-G-G-C-T-C-A-G-T-C-T-A-T-T-A-G-C-T-A-G-A-G-C 28 bases

G-C-T-C-T-A-G-C-T-A-A-T-A-G-A-C-T-G-A-G-C-C-G-A-G-G-T-G 28 bases

DOI no: 10.1038/nchembio.1926

Nat Chem Biol 11:870-877 (2015)

PubMed id: 26389736

Translocation-coupled DNA cleavage by the Type ISP restriction-modification enzymes.

M.K.Chand, N.Nirwan, F.M.Diffin, K.van Aelst, M.Kulkarni, C.Pernstich, M.D.Szczelkun, K.Saikrishnan.

ABSTRACT

Production of endonucleolytic double-strand DNA breaks requires separate strand cleavage events. Although catalytic mechanisms for simple, dimeric endonucleases are known, there are many complex nuclease machines that are poorly understood. Here we studied the single polypeptide Type ISP restriction-modification (RM) enzymes, which cleave random DNA between distant target sites when two enzymes collide after convergent ATP-driven translocation. We report the 2.7-Å resolution X-ray crystal structure of a Type ISP enzyme-DNA complex, revealing that both the helicase-like ATPase and nuclease are located upstream of the direction of translocation, an observation inconsistent with simple nuclease-domain dimerization. Using single-molecule and biochemical techniques, we demonstrate that each ATPase remodels its DNA-protein complex and translocates along DNA without looping it, leading to a collision complex in which the nuclease domains are distal. Sequencing of the products of single cleavage events suggests a previously undescribed endonuclease model, where multiple, stochastic strand-nicking events combine to produce DNA scission.