PDBsum entry 1ej9

Isomerase/DNA

PDB id: 1ej9

Contents

Protein chain

483 a.a. *

DNA/RNA

Waters ×353

* Residue conservation analysis

PDB id:

1ej9

Name:

Isomerase/DNA

Title:

Crystal structure of human topoisomerase i DNA complex

Structure:

DNA (5'- d( C Ap Ap Ap Ap Ap Gp Ap Cp Tp Cp Ap Gp Ap Ap Ap Ap Ap Tp Tp Tp Tp T )-3'). Chain: c. Engineered: yes. DNA (5'- d( C Ap Ap Ap Ap Ap Tp Tp Tp Tp Tp Cp Tp Gp Ap Gp Tp Cp Tp Tp Tp Tp T )-3'). Chain: d.

Source:

Synthetic: yes. Homo sapiens. Human. Organism_taxid: 9606. Expressed in: spodoptera frugiperda. Expression_system_taxid: 7108. Expression_system_cell: sf9 insect cells

Biol. unit:

Trimer (from PQS)

Resolution:

2.60Å R-factor: 0.214 R-free: 0.281

Authors:

M.R.Redinbo,J.J.Champoux,W.G.Hol

Key ref:

M.R.Redinbo et al. (2000). Novel insights into catalytic mechanism from a crystal structure of human topoisomerase I in complex with DNA. Biochemistry, 39, 6832-6840. PubMed id: 10841763 DOI: 10.1021/bi992690t

Date:

01-Mar-00 Release date: 03-Aug-00

Protein chain

P11387  (TOP1_HUMAN) -  DNA topoisomerase 1 from Homo sapiens

Seq: 765 a.a.

Struc: 483 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

DNA/RNA chains

A-A-A-A-A-G-A-C-T-C-A-G-A-A-A-A-A-T-T-T-T-T 22 bases

A-A-A-A-A-T-T-T-T-T-C-T-G-A-G-T-C-T-T-T-T-T 22 bases

Enzyme reactions

• Enzyme class: E.C.5.6.2.1 - Dna topoisomerase.

DOI no: 10.1021/bi992690t

Biochemistry 39:6832-6840 (2000)

PubMed id: 10841763

Novel insights into catalytic mechanism from a crystal structure of human topoisomerase I in complex with DNA.

M.R.Redinbo, J.J.Champoux, W.G.Hol.

ABSTRACT

Human topoisomerase I helps to control the level of DNA supercoiling in cells and is vital for numerous DNA metabolic events, including replication, transcription, and recombination. The 2.6 A crystal structure of human topoisomerase I in noncovalent complex with a DNA duplex containing a cytosine at the -1 position of the scissile strand rather than the favored thymine is reported. The hydrogen bond between the O2 position of this -1 base and the epsilon-amino of the conserved Lys-532 residue, the only base-specific contact observed previously in the human topoisomerase I-DNA interaction, is maintained in this complex. Several unique features of this structure, however, have implications for the DNA-binding and active-site mechanisms of the enzyme. First, the ends of the DNA duplex were observed to shift by up to 5.4 A perpendicular to the DNA helical axis relative to structures reported previously, suggesting a novel degree of plasticity in the interaction between human topoisomerase I and its DNA substrate. Second, 12 additional residues at the NH(2) terminus of the protein (Trp-203-Gly-214) could be built in this structure, and they were found to pack against the putative hinge region implicated in the clamping of the enzyme around duplex DNA. Third, a water molecule was observed adjacent to the scissile phosphate and the active-site residues; the potential specific base character of this solvent molecule in the active-site mechanism of the enzyme is discussed. Fourth, the scissile phosphate group was found to be rotated by 75 degrees, bringing Lys-532 into hydrogen-bonding distance of one of the nonbridging phosphate oxygens. This orientation of the scissile phosphate group implicates Lys-532 as a fifth active-site residue, and also mimics the orientation observed for the 3'-phosphotyrosine linkage in the covalent human topoisomerase I-DNA complex structure. The implications of these structural features for the mechanism of the enzyme are discussed, including the potential requirement for a rotation of the scissile phosphate group during DNA strand cleavage and covalent attachment.