Nuclease P1

 

P1 nuclease is a 36 kDa glycoprotein that cleaves its substrate at every position yielding nucleoside 5' monophosphates, and it does not recognize or act on double-stranded DNA [PMID: 9726413]. It is useful at removing single stranded strands hanging off the end of double stranded DNA and at completely cleaving melted DNA for simple DNA composition analysis. The enzyme is a phosphodiesterase cleaving the P-O3' bond with inversion of configuration at the phosphorus, but also acts as a phosphomonesterase removing 3'-terminal phosphate groups. It has no apparent specificity for bases.

 

Reference Protein and Structure

Sequence
P24289 UniProt (3.1.30.1) IPR003154 (Sequence Homologues) (PDB Homologues)
Biological species
Penicillium citrinum (Fungus) Uniprot
PDB
1ak0 - P1 NUCLEASE IN COMPLEX WITH A SUBSTRATE ANALOG (1.8 Å) PDBe PDBsum 1ak0
Catalytic CATH Domains
1.10.575.10 CATHdb (see all for 1ak0)
Cofactors
Zinc(2+) (3)
Click To Show Structure

Enzyme Reaction (EC:3.1.30.1)

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: Aspergillus oryzae S(1) nuclease, Neurospora crassa single-strand specific endonuclease, S(1) nuclease, Deoxyribonuclease S(1), Endonuclease S(1) (Aspergillus), Nuclease S(1), Single strand-specific DNase, Single-strand endodeoxyribonuclease, Single-strand-specific endodeoxyribonuclease, Single-stranded DNA specific endonuclease, Single-stranded-nucleate endonuclease, Endonuclease S1, Deoxyribonuclease S1, Aspergillus nuclease S1,

Enzyme Mechanism

Introduction

Two zinc ions coordinate and activate a water molecule, with the help of Asp45, to attack the phosphate in line with the P-O3' bond. Another zinc ion stabilises the leaving oxyanion, while an arginine residue stabilises the negative charge of the transition state.

Catalytic Residues Roles

UniProt PDB* (1ak0)
Arg48 Arg48A Stabilises negatively charged transition state. In several homologues, this residue is a lysine, rather than argenine. However, the proposed function remains the same. electrostatic stabiliser
Asp45 Asp45A Helps activate the nucleophilic water molecule. activator, metal ligand
His6, Trp1 (N-term), Asp120 His6A, Trp1A (N-term), Asp120A Forms part of the zinc 3 binding site. metal ligand
His60, Asp45, His116, Asp120 His60A, Asp45A, His116A, Asp120A Forms part of the zinc 1 binding site. metal ligand
His126, His149, Asp153 His126A, His149A, Asp153A Forms part of the zinc 2 binding site. 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

References

  1. Romier C et al. (1998), Proteins, 32, 414-424. Recognition of single-stranded DNA by nuclease P1: High resolution crystal structures of complexes with substrate analogs. DOI:10.1002/(sici)1097-0134(19980901)32:4<414::aid-prot2>3.0.co;2-g. PMID:9726413.
  2. Kovaľ T et al. (2016), PLoS One, 11, e0168832-. Structural and Catalytic Properties of S1 Nuclease from Aspergillus oryzae Responsible for Substrate Recognition, Cleavage, Non–Specificity, and Inhibition. DOI:10.1371/journal.pone.0168832. PMID:28036383.
  3. Yu TF et al. (2014), PLoS One, 9, e105821-. Structural Insights of the ssDNA Binding Site in the Multifunctional Endonuclease AtBFN2 from Arabidopsis thaliana. DOI:10.1371/journal.pone.0105821. PMID:25157844.
  4. Chou T et al. (2013), Biocatal Agric Biotechnol, 2, 191-195. Mechanistic insights to catalysis by a zinc-dependent bi-functional nuclease from Arabidopsis thaliana. DOI:10.1016/j.bcab.2013.03.006.
  5. Liao RZ et al. (2010), Inorg Chem, 49, 6883-6888. Phosphate Mono- and Diesterase Activities of the Trinuclear Zinc Enzyme Nuclease P1—Insights from Quantum Chemical Calculations. DOI:10.1021/ic100266n. PMID:20604512.
  6. Naik AK et al. (2008), DNA Repair (Amst), 7, 1384-1391. P1 nuclease cleavage is dependent on length of the mismatches in DNA. DOI:10.1016/j.dnarep.2008.04.010. PMID:18524693.

Step 1.

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Catalytic Residues Roles

Residue Roles
Arg48A electrostatic stabiliser
Asp45A activator
Trp1A (N-term) metal ligand
His6A metal ligand
Asp120A metal ligand
Asp45A metal ligand
His60A metal ligand
His116A metal ligand
His126A metal ligand
His149A metal ligand
Asp153A metal ligand

Chemical Components

Step 1.

Contributors

Stuart Lucas, Craig Porter, Gemma L. Holliday