Deoxyribonuclease I

 

DNase I is a vertebrate enzyme that cleaves double stranded DNA to 5'-phosphodinucleotide and 5'-phospho-oligonucletide end-products. DNase I requires calcium and magnesium for full activity. It is a glycoprotein that causes single-stranded nicks on double-stranded DNA. DNase I shares structural homology to the Human Apurinic/Apyrimidinic endonuclease and Exonuclease III. These three enzymes also share similar mechanisms of DNA cleavage.

The overall reaction has been created using poly dAMP to show the chemical reaction.

 

Reference Protein and Structure

Sequence
P00639 UniProt (3.1.21.1) IPR016202 (Sequence Homologues) (PDB Homologues)
Biological species
Bos taurus (Cattle) Uniprot
PDB
1dnk - THE X-RAY STRUCTURE OF THE DNASE I-D(GGTATACC)2 COMPLEX AT 2.3 ANGSTROMS RESOLUTION (2.3 Å) PDBe PDBsum 1dnk
Catalytic CATH Domains
3.60.10.10 CATHdb (see all for 1dnk)
Cofactors
Magnesium(2+) (2) Metal MACiE
Click To Show Structure

Enzyme Reaction (EC:3.1.21.1)

water
CHEBI:15377ChEBI
+
single-stranded DNA
CHEBI:9160ChEBI
→
5'-end 2'-deoxyribonucleotide(2-) residue
CHEBI:136412ChEBI
+
2'-deoxynucleoside 3'-monophosphate(2-)
CHEBI:131705ChEBI
+
hydron
CHEBI:15378ChEBI
Alternative enzyme names: Escherichia coli endonuclease I, DNA depolymerase, DNA endonuclease, DNA nuclease, DNAase, DNase, DNase I, Alkaline DNase, Alkaline deoxyribonuclease, Deoxyribonuclease (pancreatic), Deoxyribonuclease A, Deoxyribonucleic phosphatase, Dornava, Dornavac, Endodeoxyribonuclease I, Pancreatic DNase, Pancreatic deoxyribonuclease, Pancreatic dornase, Thymonuclease, dornase, Thymonuclease,

Enzyme Mechanism

Introduction

The reaction involves an in-line attack at the DNA phosphorus group by a water molecule. His252 may act as a general base to abstract a proton from a water molecule opposite the O3' atom. Subsequent attack at the phosphorus group with inversion of its configuration gives rise to a penta-covalent state that may be stabilised by the metal ion. His134 is in an ideal position to protonate the leaving O3'. The metal ion may also be important for the correct positioning of the phosphate group.

Catalytic Residues Roles

UniProt PDB* (1dnk)
Asp234 Asp212A(C) Activates His252 to act as the general base. increase basicity, electrostatic stabiliser, increase acidity
Tyr98, Glu100 Tyr76A(C), Glu78A(C) Part of the His-Glu-Tyr triad that activates His134 as a general acid. electrostatic stabiliser
His156, His274 His134A(C), His252A(C) Acts as a general acid/base. proton acceptor, proton donor
Glu61, Asp190 Glu39A(C), Asp168A(C) Binds one of the Mg(II) ions. metal ligand
*PDB label guide - RESx(y)B(C) - RES: Residue Name; x: Residue ID in PDB file; y: Residue ID in PDB sequence if different from PDB file; B: PDB Chain; C: Biological Assembly Chain if different from PDB. If label is "Not Found" it means this residue is not found in the reference PDB.

Chemical Components

bimolecular nucleophilic substitution, overall reactant used, overall product formed, hydrolysis, proton transfer, rate-determining step, native state of enzyme regenerated, proton relay, inferred reaction step

References

  1. Jones SJ et al. (1996), J Mol Biol, 264, 1154-1163. Site-directed Mutagenesis of the Catalytic Residues of Bovine Pancreatic Deoxyribonuclease I. DOI:10.1006/jmbi.1996.0703. PMID:9000637.
  2. Chen WJ et al. (2007), Biochem Biophys Res Commun, 352, 689-696. Probing the catalytic mechanism of bovine pancreatic deoxyribonuclease I by chemical rescue. DOI:10.1016/j.bbrc.2006.11.078. PMID:17141190.
  3. Kabsch W et al. (1990), Nature, 347, 37-44. Atomic structure of the actin: DNase I complex. DOI:10.1038/347037a0. PMID:2395459.

Step 1. His252 deprotonates water, which initiates a nucleophilic attack on the phosphate of the DNA in a substitution reaction which eliminates the 5' end of the DNA, with concomitant deprotonation of His134.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Glu39A(C) metal ligand
Asp168A(C) metal ligand
Tyr76A(C) electrostatic stabiliser
Glu78A(C) increase acidity, electrostatic stabiliser
Asp212A(C) increase basicity, electrostatic stabiliser
His252A(C) proton acceptor
His134A(C) proton donor

Chemical Components

ingold: bimolecular nucleophilic substitution, overall reactant used, overall product formed, hydrolysis, proton transfer, rate-determining step

Step 2. His134 deprotonates water, which in turn deprotonates His252 in an inferred return step.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Glu39A(C) metal ligand
Asp168A(C) metal ligand
Tyr76A(C) electrostatic stabiliser
Glu78A(C) increase basicity, electrostatic stabiliser
Asp212A(C) increase acidity, electrostatic stabiliser
His252A(C) proton donor
His134A(C) proton acceptor

Chemical Components

native state of enzyme regenerated, proton relay, inferred reaction step, proton transfer
Download: Image, Marvin File

Step 1. His252 deprotonates water, which initiates a nucleophilic attack on the phosphate of the DNA in a substitution reaction which eliminates the 5' end of the DNA, with concomitant deprotonation of His134.

Step 2. His134 deprotonates water, which in turn deprotonates His252 in an inferred return step.

Products.

Contributors

Gemma L. Holliday, Gail J. Bartlett, Daniel E. Almonacid, James W. Murray, Craig Porter