DNASE1

Deoxyribonuclease I, customarily called DNase I, is a specific DNA endonuclease covering a single glycosylated polypeptide chain with two disulfide bonds and is coded by the human gene DNASE1. The optimal pH of DNase I is around 7.5. The activity and structural integrity of DNase I is dependent on divalent cations, where Mg²⁺ or Mn²⁺ activates DNase I, and Ca²⁺ is required to exert full DNase I activity. DNase I binds to actin monomers with very high (sub-nanomolar) affinity and to actin polymers with lower affinity, both of which could inactivate the actin-bound DNase I. Herein, the DNase-actin complex might be an effective storage form of DNase I, which restrains the damage of genetic information. Generally, DNase I is stored in the zymogen granules of the nuclear envelope.

Genomics

In genomics, hypersensitive sites of DNase I are assumed to be characterized by open, accessible chromatin and have been shown to be capable of labeling many different types of genetic regulatory elements, including enhancers, promoters, insulators, silencers, locus control regions, and histone modifications. Consequently, DNase I sensitivity assay is widely adopted in genomics to identify which regions of the genome may contain active genes. The sequence of DNase I is evolutionarily related to that of human muscle-specific DNase-like protein and human proteins DHP1 and DHP2, where yet the first disulphide bond of DNase I is not conserved. Alternate transcriptional splice variants of DNASE1 have been noticed but have not been thoroughly characterized.

Specificity

DNase I has recently been reported to show some levels of sequence specificity that may depend on experimental conditions. Unlike other enzymes with high substrate specificity, DNase I certainly could not work with an absolute sequence specificity. However, cleavage at sites containing C or G at their 3' terminal is found to be less efficient. It shows partiality to cleave at the 5’ side of pyrimidines, and is predominantly pronounced in alternative copolymers. It has been shown that variations in the twist angle are recognized by DNase I. The specificity of DNase I also rests with the present divalent cations. The presence of Ca²⁺ and Mg²⁺ leads to single-strand breaks, and the appearance of Mn²⁺ could be related with double-strand breaks.

Action Mode

DNase I indiscriminately digests phosphodiester linkages next to a pyrimidine nucleotide to produce oligonucleotide, trinucleotide, or dinucleotide fragments with 5’-phosphorylated and 3’-hydroxylated ends. It can cleave single- and double-stranded DNA, and DNA-RNA hybrids in an endonucleolytic manner. DNase I results in single-strand nicks in duplex DNA, when a proton acceptor-donor chain containing a glutamic acid and a histidine generates a nucleophilic hydroxyl ion to cleave the 3'-P-O bond. It also facilitates the breakdown of chromatin during apoptosis. DNase I is normally secreted outside of the cell, but likely to be able to enter into the nucleus where it is associated with apoptosis to induce programmed cell death. The action manner of DNase I is neither base- nor sequence-specific, while the cleavage pattern and the frequency of cutting are clearly sequence-dependent, demonstrating that DNase I could recognize variations in the geometry of the DNA backbone. DNase I also functions at a rate depending strongly upon its chromatin environment.

Figure 1. Catalytic mechanism of DNase I.

Functions

DNase I is usually applied in tissue dissociation protocols to digest DNA that has leaked into the tissue culture on account of cell damage. DNase I is also employed for the removal of membrane-bound DNA fragments. Apart from its role as a waste-management endonuclease, DNase I has been recommended to be responsible for DNA fragmentation during apoptosis. The digestion of human tissues such as cartilage, microglia, epithelium, colon, lung, liver, neural, and stem cells can be accomplished at the disposal of DNase I. Active DNase I is critical to prevent immune stimulation, and the decrease in degrading potency may increase the risk for production of anti-nucleosome antibodies, which signifies the development of systemic lupus erythematosus. The formation of a complex from with G-actin results in the inhibition of DNase activity and loss of the capability of G-actin to polymerise into fibres. DNase I has been employed to control lung problems in patients with cystic fibrosis by destroying DNA found in purulent lung secretions with an attempt to reduce the viscosity and make it easier for the patient to breathe. Transient myocardial ischaemia occurring during percutaneous coronary intervention could induce a prominent elevation of serum DNase I activity, which may be valuable as a sensitive indicator for the detection of transient myocardial ischaemia. It is also noteworthy that the application of a mammalian DNase may overcome the limitations of immunotoxin therapy with less toxicity and less immunogenity than currently available immunotoxins.

Reference