Micrococcal nuclease
Micrococcal nuclease (EC 3.1.31.1, S7 Nuclease, MNase, spleen endonuclease, thermonuclease, nuclease T, micrococcal endonuclease, nuclease T', staphylococcal nuclease, spleen phosphodiesterase, Staphylococcus aureus nuclease, Staphylococcus aureus nuclease B, ribonucleate (deoxynucleate) 3'-nucleotidohydrolase) is an endo-exonuclease that preferentially digests single-stranded nucleic acids. The rate of cleavage is 30 times greater at the 5' side of A or T than at G or C and results in the production of mononucleotides and oligonucleotides with terminal 3'-phosphates. The enzyme is also active against double-stranded DNA and RNA and all sequences will be ultimately cleaved.
| Micrococcal nuclease | |||||||||
|---|---|---|---|---|---|---|---|---|---|
 Ribbon schematic of micrococcal nuclease 3D structure, with Ca2+ and TdtP inhibitor | |||||||||
| Identifiers | |||||||||
| EC no. | 3.1.31.1 | ||||||||
| CAS no. | 9013-53-0 | ||||||||
| Databases | |||||||||
| IntEnz | IntEnz view | ||||||||
| BRENDA | BRENDA entry | ||||||||
| ExPASy | NiceZyme view | ||||||||
| KEGG | KEGG entry | ||||||||
| MetaCyc | metabolic pathway | ||||||||
| PRIAM | profile | ||||||||
| PDB structures | RCSB PDB PDBe PDBsum | ||||||||
| |||||||||
| Thermonuclease | |||||||
|---|---|---|---|---|---|---|---|
| Identifiers | |||||||
| Organism | |||||||
| Symbol | nuc | ||||||
| UniProt | P00644 | ||||||
| |||||||
| Staphylococcal nuclease | |||||||||
|---|---|---|---|---|---|---|---|---|---|
| Identifiers | |||||||||
| Symbol | ? | ||||||||
| Pfam | PF00565 | ||||||||
| Pfam clan | CL0049 | ||||||||
| InterPro | IPR016071 | ||||||||
| PROSITE | PDOC00865 | ||||||||
| CATH | 1tt2 | ||||||||
| SCOP2 | 1tt2 / SCOPe / SUPFAM | ||||||||
| |||||||||
Characteristics
The enzyme has a molecular weight of 16.9kDa.
The pH optimum is reported as 9.2. The enzyme activity is strictly dependent on Ca2+ and the pH optimum varies according to Ca2+ concentration.[1] The enzyme is therefore easily inactivated by EGTA.
Sources
This enzyme is the extracellular nuclease of Staphylococcus aureus. Two strains, V8 and Foggi, yield almost identical enzymes.[2] A common source is E.coli cells carrying a cloned nuc gene encoding Staphylococcus aureus extracellular nuclease (micrococcal nuclease).
Structure
The 3-dimensional structure of micrococcal nuclease (then called Staphyloccal nuclease) was solved very early in the history of protein crystallography, in 1969,[3] deposited as now-obsolete Protein Data Bank file 1SNS. Higher-resolution, more recent crystal structures are available for the apo form as Protein Data Bank file 1SNO: and for the thymidine-diphosphate-inhibited form as Protein Data Bank file 3H6M: or 1SNC: . As seen in the ribbon diagram above, the nuclease molecule has 3 long alpha helices and a 5-stranded, barrel-shaped beta sheet, in an arrangement known as the OB-fold (for oligonucleotide-binding fold) as classified in the SCOP database.
Applications
- CUT&RUN sequencing, antibody-targeted controlled cleavage by micrococcal nuclease for transcriptomic profiling.
- Hydrolysis of nucleic acids in crude cell-free extracts.
- Sequencing of RNA.
- Preparation of rabbit reticulocyte lysates.
- Studies of chromatin structure.
- Removal of nucleic acids from laboratory protein preparations allowing for protein folding and structure-function studies.
- Research on the mechanisms of protein folding.
See also
References
- Heins JN, Suriano JR, Taniuchi H, Anfinsen CB (1967). "Characterization of a nuclease produced by Staphylococcus aureus". J. Biol. Chem. 242 (5): 1016–20. doi:10.1016/S0021-9258(18)96225-3. PMID 6020427.
- Cusumano CL, Taniuchi H, Anfinsen CB (1968). "Staphylococcal nuclease (Foggi strain). I. Order of cyanogen bromide fragments and a "fourth" histidine residue". J. Biol. Chem. 243 (18): 4769–77. doi:10.1016/S0021-9258(18)93185-6. PMID 5687719.
- Arnone A, Bier J, et al. (1971). "A High Resolution Structure of an Inhibitor Complex of the Extracellular Nuclease of Staphylococcus aureus: I. Experimental Procedures and Chain Tracing". J. Biol. Chem. 246 (7): 2303–2316. doi:10.1016/S0021-9258(19)77221-4. PMID 5555571.
- http://www.thermoscientificbio.com/dna-and-rna-modifying-enzymes/micrococcal-nuclease/
- http://www.worthington-biochem.com/NFCP/default.html
- http://www.thermoscientificbio.com/uploadedFiles/Resources/en0181-usa-msds.pdf - A material and safety data sheet for the product
- http://www.thermoscientificbio.com/uploadedFiles/Resources/en018-product-information.pdf - A Product Information sheet
External links
- Micrococcal+Nuclease at the U.S. National Library of Medicine Medical Subject Headings (MeSH)
- EC 3.1.31.1