View source for Biology:Deoxyribonuclease

{{Short description|Enzymes that break down DNA}}
'''Deoxyribonuclease''' ('''DNase''', for short) refers to a group of [[Chemistry:Glycoprotein|glycoprotein]] [[Biology:Endonuclease|endonuclease]]s which are [[Biology:Enzymes|enzymes]] that catalyze the [[Chemistry:Hydrolysis|hydrolytic cleavage]] of phosphodiester linkages in the [[Biology:DNA|DNA]] backbone, thus degrading DNA. The role of the DNase enzyme in cells includes breaking down extracellular DNA (ecDNA) excreted by [[Biology:Apoptosis|apoptosis]], [[Biology:Necrosis|necrosis]], and [[Biology:Neutrophil extracellular traps|neutrophil extracellular traps]] (NET) of cells to help reduce inflammatory responses that otherwise are elicited. A wide variety of deoxyribonucleases are known and fall into one of two families ([[Biology:Deoxyribonuclease I|DNase I]] or [[Biology:Deoxyribonuclease II|DNase II]]), which differ in their [[Biology:Substrate (biochemistry)|substrate]] specificities, chemical mechanisms, and biological functions. Laboratory applications of DNase include [[Biology:Protein purification|purifying proteins]] when extracted from [[Biology:Prokaryote|prokaryotic]] organisms. Additionally, DNase has been applied as a treatment for diseases that are caused by ecDNA in the blood plasma. Assays of DNase are emerging in the research field as well.

== Types ==
The two main types of DNase found in [[Biology:Animal|animals]] are known as deoxyribonuclease I (DNase I) and deoxyribonuclease II (DNase II). These two families have subcategories within them.

=== The DNase I Family: DNase I, DNase1L1, DNase 1L2, DNase1L3 ===
The first set of DNases is [[Biology:Deoxyribonuclease I|DNase I]]. This family consisted of DNase I, [[Biology:DNASE1L1|DNase1L1]], DNase 1L2, and DNase1L3. DNase I cleaves DNA to form two [[Chemistry:Oligonucleotide|oligonucleotide]]-end products with 5’-phospho and 3’-hydroxy ends and is produced mainly by organs of the digestive system. The DNase I family requires Ca2+ and Mg2+ [[Physics:Cations|cations]] as activators and selectively expressed.<ref>{{cite journal |last1=Junowicz |first1=E. |title=Studies on bovine pancreatic deoxyribonuclease A. II. The effect of different bivalent metals on the specificity of degradation of DNA. |journal=Biochim. Biophys. Acta |year=1973 |volume=312 |issue=1 |pages=85–102 |doi=10.1016/0005-2787(73)90054-3|pmid=4353710 }}</ref> In terms of pH, the DNAses I family is active in normal pH of around 6.5 to 8.

=== The DNase II Family: DNase II ɑ and DNase II ꞵ ===
The second set of DNAases is DNase II. This family consisted of DNase II ɑ and DNase II ꞵ. Like DNAase I, DNAase II cleaves DNA to form two oligonucleotide-end products with 5’-hydroxy and 3’-phospho ends. This type of DNAase is more widely expressed in tissues due to high expression in macrophages but limited cell-type expression. Unlike DNAase I, they do not need Ca2+ and Mg2+ cations as activators.<ref>{{cite journal |last1=Ohkouchi |first1=S. |last2=Shibata |first2=M |last3=Sasaki |first3=M |last4=Koike |first4=M |last5=Safig |first5=P |last6=Peters |first6=C |last7=Nagata |first7=S |last8=Uchiyama |first8=Y |title=Biogenesis and proteolytic processing of lysosomal DNase II |journal=PLOS ONE |date=2013 |volume=8 |issue=3 |pages=e59148 |doi=10.1371/journal.pone.0059148|pmid=23516607 |pmc=3596287 |bibcode=2013PLoSO...859148O |doi-access=free }}</ref>  In terms of [[Chemistry:PH|pH]], the DNAase II family is expressed in acidic pH. The cleavage pattern of DNase II is altered in the presence of Dimethyl sulfoxide([[Chemistry:Dimethyl sulfoxide|DMSO]]), which significantly affects the structure of DNA.

== Structure ==
Although both DNase I and II are glycoprotein endonucleases, DNase I has a [[Chemistry:Monomer|monomer]]ic sandwich-type structure with a carbohydrate side chain whereas DNase II has a [[Biology:Protein dimer|dimeric]] [[Biology:Protein quaternary structure|quaternary structure]]. [[File:DNase 1 image 1.png|thumb|Glycoprotein DNase I 3D structure PDB: 3DNI  |211x211px]]'''DNase I Structure:''' DNase I is a glycoprotein with a molecular weight of 30,000 Da and a carbohydrate chain of 8-10 residues attached to Asn18 (orange).<ref name=":0">{{Cite journal |last1=Suck |first1=D. |last2=Oefner |first2=C. |last3=Kabsch |first3=W. |date=1984 |title=Three-dimensional structure of bovine pancreatic DNase I at 2.5 A resolution. |journal=The EMBO Journal |language=en |volume=3 |issue=10 |pages=2423–2430 |doi=10.1002/j.1460-2075.1984.tb02149.x|pmid=6499835 |pmc=557703 }}</ref> It is an 𝛼,𝛽-protein with two 6-stranded 𝛽-pleated sheets which form the core of the structure.<ref name=":1">{{Cite web |last=wwPDB.org |title=wwPDB: Worldwide Protein Data Bank |url=http://www.wwpdb.org |access-date=2022-10-26 |website=www.wwpdb.org |language=en}}</ref> These two core sheets run parallel, and all others run antiparallel. The 𝛽-pleated sheets lie in the center of the structure while the 𝛼-helices are denoted by the coils on the periphery. DNase I contains four ion-binding pockets, and requires Ca<sup>2+</sup> and Mg<sup>2+</sup> for hydrolyzing double-stranded DNA.<ref name=":5">{{Cite journal |last1=Guéroult |first1=Marc |last2=Picot |first2=Daniel |last3=Abi-Ghanem |first3=Joséphine |last4=Hartmann |first4=Brigitte |last5=Baaden |first5=Marc |date=2010-11-18 |editor-last=Levitt |editor-first=Michael |title=How Cations Can Assist DNase I in DNA Binding and Hydrolysis |journal=PLOS Computational Biology |language=en |volume=6 |issue=11 |pages=e1001000 |doi=10.1371/journal.pcbi.1001000 |pmid=21124947 |pmc=2987838 |bibcode=2010PLSCB...6E1000G |issn=1553-7358 |doi-access=free }}</ref> Two of the sites strongly bind Ca<sup>2+</sup> while the other two coordinate Mg<sup>2+</sup>. Little has been published on the number and location of the Mg<sup>2+</sup> binding sites, although it has been proposed that Mg<sup>2+</sup> is located near the catalytic pocket and contributes to hydrolysis.<ref>{{Cite journal |last1=Jones |first1=S. J. |last2=Worrall |first2=A. F. |last3=Connolly |first3=B. A. |date=1996-12-20 |title=Site-directed mutagenesis of the catalytic residues of bovine pancreatic deoxyribonuclease I |url=https://pubmed.ncbi.nlm.nih.gov/9000637/ |journal=Journal of Molecular Biology |volume=264 |issue=5 |pages=1154–1163 |doi=10.1006/jmbi.1996.0703 |issn=0022-2836 |pmid=9000637}}</ref> The two Ca<sup>2+</sup> are shown in red in the image. They are bound to DNase I under crystallization conditions and are important for the structural integrity of the molecule by stabilizing the surface loop Asp198 to Thr204 (cyan), and by limiting the region of high thermal mobility in the flexible loop to residues Gly97 to Gly102 (yellow).

[[File:DNase II.png|thumb|Glycoprotein DNase II 3D structure PDB: 5UNB |210x210px]]
'''DNase II Structure:''' DNase II contains a homodimeric quaternary structure that is capable of binding double-stranded DNA within a U-shaped clamp architecture. The interior of the U-shaped clamp is largely electropositive, capable of binding negatively-charged DNA. Similar to DNase I, DNase II structure consists of a mixed 𝛼/𝛽 secondary structure with 9 𝛼-helices and 20 𝛽-pleated sheets.<ref name="foo">{{Cite journal |last1=Varela-Ramirez |first1=Armando |last2=Abendroth |first2=Jan |last3=Mejia |first3=Adrian A. |last4=Phan |first4=Isabelle Q. |last5=Lorimer |first5=Donald D. |last6=Edwards |first6=Thomas E. |last7=Aguilera |first7=Renato J. |date=2017-06-02 |title=Structure of acid deoxyribonuclease |journal=Nucleic Acids Research |volume=45 |issue=10 |pages=6217–6227 |doi=10.1093/nar/gkx222 |issn=0305-1048 |pmc=5449587 |pmid=28369538}}</ref> Although unlike DNase I, DNase II does not require divalent metal ions for catalysis.<ref name="foo"/> The structure consists of protomer A (cyan) and protomer B (green). Each structure consists of two catalytic motifs, which are labeled on protomer B for simplicity: His100 and Lys102 compose the first motif (blue) and His279 and Lys281 compose the second catalytic motif (red). 
[[File:DNase MOA.jpg|thumb|266x266px|Mechanism of action for DNase enzymes]]

== Mechanism ==
Some DNases cut, or "cleave", only residues at the ends of DNA molecules. This type of [[Biology:Exonuclease|exonuclease]] is known as [[Biology:Exodeoxyribonuclease|exodeoxyribonuclease]]s. Others cleave anywhere along the chain, known as [[Biology:Endodeoxyribonuclease|endodeoxyribonuclease]]s (a subset of [[Biology:Endonuclease|endonuclease]]s.)<ref>{{cite book |url=http://arbl.cvmbs.colostate.edu/hbooks/genetics/biotech/enzymes/nucleases.html |title=Nucleases: DNase and RNase |vauthors=Bowen RA, Austgen L, Rouge M |date=20 February 2000 |work=Biotechnology and Genetic Engineering |section=Restriction Endonucleases and DNA Modifying Enzymes |access-date=8 January 2017 |archive-url=https://web.archive.org/web/20040805185739/http://arbl.cvmbs.colostate.edu/hbooks/genetics/biotech/enzymes/nucleases.html |archive-date=5 August 2004 |url-status=dead}}</ref>
Some DNases are fairly indiscriminate about the DNA sequence at which they cut, while others, including [[Biology:Restriction enzyme|restriction enzyme]]s, are very sequence-specific. Other DNases cleave only double-stranded DNA, others are specific for single-stranded molecules, and others are active toward both.

The action of DNase occurs in three phases. The initial phase introduces multiple nicks in the phosphodiester backbone. The second phase produces acid-soluble nucleotides. The third phase, which is the terminal phase, consists of reduction of oligonucleotides, causing a [[Biology:Hyperchromic shift|hyperchromic shift]] in the UV data.<ref>{{Citation |last=Bernardi |first=Giorgio |title=11 Spleen Acid Deoxyribonuclease |date=1971-01-01 |url=https://www.sciencedirect.com/science/article/pii/S1874604708603716 |work=The Enzymes |volume=4 |pages=271–287 |editor-last=Boyer |editor-first=Paul D. |series=Hydrolysis |publisher=Academic Press |doi=10.1016/S1874-6047(08)60371-6 |isbn=9780121227043 |language=en |access-date=2022-10-27}}</ref>

=== DNase I Mechanism ===
DNase I predominantly targets double-stranded DNA, and to a lesser extent, some single-stranded DNA for cleavage. DNase I catalyzes nonspecific DNA cleavage by nicking [[Biology:Phosphodiester bond|phosphodiester]] linkages in one of the strands. Its cleavage site lies between the 3′-oxygen atom and the adjacent phosphorus atom, yielding 3′-hydroxyl and 5′-phosphoryl [[Chemistry:Oligonucleotide|oligonucleotide]]s with inversion of configuration at the phosphorus. The DNase enzyme relies on the presence of a [[Chemistry:Valence|divalent]] cation, which is usually Ca<sup>2+</sup>, for proper function. The active site of DNase I includes two [[Chemistry:Histidine|histidine]] residues (His134 and His252) and two acidic residues ([[Chemistry:Glutamic acid|Glu]]78 and [[Chemistry:Aspartic acid|Asp]] 212), all of which are critical for the general acid-base catalysis of phosphodiester bonds.<ref>{{Citation |last1=Lazarus |first1=Robert A. |date=2019 |pages=471–488 |place=Cham |publisher=Springer International Publishing |language=en |doi=10.1007/978-3-030-00710-2_22 |isbn=978-3-030-00709-6 |last2=Wagener† |first2=Jeffrey S.|title=Pharmaceutical Biotechnology |chapter=Recombinant Human Deoxyribonuclease I |pmc=7124075 }}</ref>

=== DNase II Mechanism ===
Deoxyribonuclease II (DNase II) is also known as acid deoxyribonuclease because it has optimal activity in the low pH environment of lysosomes where it is typically found in higher eukaryotes. Some forms of recombinant DNase II display a high level of activity in low pH in the absence of divalent metal ions, similar to eukaryotic DNase II.<ref name="foo" /> Unlike DNase I, DNase II cleaves the phosphodiester bond between the 5'-oxygen atom and the adjacent phosphorus atom, yielding 3΄-phosphorylated and 5΄-hydroxyl nucleotides.

== Applications ==

=== Laboratory applications ===
DNase is commonly used when [[Biology:Protein purification|purifying proteins]] that are extracted from [[Biology:Prokaryote|prokaryotic]] organisms. Protein extraction often involves the degradation of the [[Biology:Cell membrane|cell membrane]]. It is common for the degraded and fragile cell membrane to be [[Biology:Lysis|lysed]], releasing unwanted DNA and the desired proteins. The resulting DNA-protein extract is highly viscous and difficult to purify, in which case DNase is added to break it down.<ref>{{Cite book |title=Fundamental Laboratory Approaches for Biochemistry and Biotechnology |vauthors=Ninfa AJ, Ballou DP, Benore M |publisher=John Wiley & Sons, Inc. |year=2010 |isbn=978-0-470-08766-4 |edition=2nd |location=United States of America |pages=234}}</ref> The DNA is hydrolyzed but the proteins are unaffected and the extract can undergo further purification.

=== Treatment ===
Extracellular DNA (ecDNA) is DNA that is found in blood circulation. It appears as a result of [[Biology:Apoptosis|apoptosis]], [[Biology:Necrosis|necrosis]], or [[Biology:Neutrophil extracellular traps|neutrophil extracellular traps]] (NET)-osis of blood and tissue cells, but can also arise from the active secretion from living cells. EcDNA and their designated [[Biology:DNA-binding protein|DNA binding proteins]] are able to activate DNA-sensing receptors, [[Biology:Pattern recognition receptor|pattern recognition receptor]]s (PRRs). PRRs are able to stimulate pathways that cause an inflammatory immune response. As a result, several studies of inflammatory diseases have found that there are high concentrations of ecDNA in blood plasma. For this reason, DNase has proven to be a possible treatment for the reduction of ecDNA in the blood plasma. DNases can be excreted both intracellularly and extracellularly and can cleave the DNA [[Biology:Phosphodiester bond|phosphodiester bond]]. This function can be used to maintain a low ecDNA concentration, therefore treating inflammation. Illnesses that result from DNA residue in blood have been targeted using the "breaking-down properties" of DNase. Studies have shown DNase to be able to act as a treatment by decreasing the viscosity of mucus.<ref name=":6">{{Cite journal |last1=Simpson |first1=G. |last2=Roomes |first2=D. |last3=Heron |first3=M. |date=2006 |title=Effects of streptokinase and deoxyribonuclease on the viscosity of human surgical and empyema pus |url=https://pubmed.ncbi.nlm.nih.gov/10858409/ |journal=Chest |volume=117 |issue=6 |pages=1728–1733 |doi=10.1378/chest.117.6.1728 |issn=0012-3692 |pmid=10858409}}</ref><ref>J.B. Armstrong, J.C. White Liquefaction of viscous purulent exudates by deoxyribonuclease Lancet, 259 (1950), pp. 739-742</ref> Administration of DNase varies dependent on the disease. It can and has been administered [[Chemistry:Dornase alfa|orally]], intrapleurally, [[Chemistry:Intravenous therapy|intravenously]], [[Medicine:Intraperitoneal injection|intraperitoneally]], and via [[Biology:Inhalation|inhalation]].<ref name=":7">{{Cite journal |last1=Lauková |first1=Lucia |last2=Konečná |first2=Barbora |last3=Janovičová |first3=Ľubica |last4=Vlková |first4=Barbora |last5=Celec |first5=Peter |date=2020-07-11 |title=Deoxyribonucleases and Their Applications in Biomedicine |journal=Biomolecules |language=en |volume=10 |issue=7 |pages=1036 |doi=10.3390/biom10071036 |pmid=32664541 |pmc=7407206 |issn=2218-273X|doi-access=free }}</ref> Several studies continue to examine the application of DNase as treatment as well as ways to monitor health. For example, recently, DNase derived from [[Medicine:Pathogenic bacteria|pathogenic bacteria]] has been used as an indicator for wound infection monitoring.<ref>{{cite journal |display-authors=6 |vauthors=Xiong Z, Achavananthadith S, Lian S, Madden LE, Ong ZX, Chua W, Kalidasan V, Li Z, Liu Z, Singh P, Yang H, Heussler SP, Kalaiselvi SM, Breese MB, Yao H, Gao Y, Sanmugam K, Tee BC, Chen PY, Loke W, Lim CT, Chiang GS, Tan BY, Li H, Becker DL, Ho JS |date=2021 |title=A wireless and battery-free wound infection sensor based on DNA hydrogel |journal=Science Advances |language=EN |volume=7 |issue=47 |pages=eabj1617 |doi=10.1126/sciadv.abj1617 |pmc=8604401 |pmid=34797719|bibcode=2021SciA....7.1617X }}</ref>

==== Respiratory diseases ====
'''[[Medicine:Cystic fibrosis|Cystic fibrosis]]''' is a [[Medicine:Genetic disorder|genetic disorder]] that affects the production of mucus, sweat, and digestive fluids, causing them to become more viscous rather than [[Physics:Lubricant|lubricant]]. DNase enzymes can be inhaled using a [[Medicine:Nebulizer|nebulizer]] by [[Medicine:Cystic fibrosis|cystic fibrosis]] sufferers. DNase enzymes help because [[Biology:White blood cell|white blood cell]]s accumulate in the mucus, and, when they break down, they release DNA, which adds to the 'stickiness' of the mucus. DNase enzymes break down the DNA, and the mucus is much easier to clear from the lungs. Specifically, DNase I, also known as FDA approved drug [[Chemistry:Dornase alfa|Pulmozyme]] (also known as dornase alfa) is used as a treatment to increase pulmonary function.

Other respiratory illness such as [[Medicine:Asthma|asthma]],<ref>{{Cite journal |last1=Boogaard |first1=R. |last2=Smit |first2=F. |last3=Schornagel |first3=R. |last4=Vaessen-Verberne |first4=A. a. P. H. |last5=Kouwenberg |first5=J. M. |last6=Hekkelaan |first6=M. |last7=Hendriks |first7=T. |last8=Feith |first8=S. W. W. |last9=Hop |first9=W. C. J. |last10=de Jongste |first10=J. C. |last11=Merkus |first11=P. J. F. M. |date=2008 |title=Recombinant human deoxyribonuclease for the treatment of acute asthma in children |journal=Thorax |volume=63 |issue=2 |pages=141–146 |doi=10.1136/thx.2007.081703 |issn=1468-3296 |pmid=17675321|s2cid=309718 |doi-access=free }}</ref> pleural empyema,<ref name=":6" /> and [[Medicine:Chronic obstructive pulmonary disease|chronic obstructive pulmonary disease]] have also been found to be positively affected by DNases properties.

Furthermore, recent studies show that intrapleural [[Biology:Tissue plasminogen activator|tissue plasminogen activator]] (tPA), a protein that is responsible for the breakdown of blood clots, combined with deoxyribonuclease increase pleural drainage, decreases hospital length of stay, and decreases the need for surgery in parapneumonic effusions and empyema.

==== Other diseases ====
'''[[Medicine:Sepsis|Sepsis]]''' is a life-threatening inflammatory disease caused by the body's extreme response to an infection. The body begins to attack itself as an inflammatory response encompasses the human body. As a result, high levels of ecDNA have been associated with the bloodstream and therefore, researchers have looked to DNase as an appropriate treatment. Studies have shown that DNase was successful in disrupting NETs and decreasing inflammatory responses. More research on the type and time of administration is needed to further establish DNase as an official treatment.<ref>{{Cite journal |last1=Chen |first1=QiXing |last2=Ye |first2=Ling |last3=Jin |first3=YuHong |last4=Zhang |first4=Ning |last5=Lou |first5=TianZheng |last6=Qiu |first6=ZeLiang |last7=Jin |first7=Yue |last8=Cheng |first8=BaoLi |last9=Fang |first9=XiangMing |date=2012 |title=Circulating nucleosomes as a predictor of sepsis and organ dysfunction in critically ill patients |journal=International Journal of Infectious Diseases |volume=16 |issue=7 |pages=e558–564 |doi=10.1016/j.ijid.2012.03.007 |issn=1878-3511 |pmid=22609014|doi-access=free }}</ref><ref name=":8">{{Cite journal |last1=Janovičová |first1=Ľubica |last2=Čonka |first2=Jozef |last3=Lauková |first3=Lucia |last4=Celec |first4=Peter |date=2022-10-01 |title=Variability of endogenous deoxyribonuclease activity and its pathophysiological consequences |url=https://www.sciencedirect.com/science/article/pii/S089085082200055X |journal=Molecular and Cellular Probes |language=en |volume=65 |pages=101844 |doi=10.1016/j.mcp.2022.101844 |pmid=35803442 |s2cid=250328196 |issn=0890-8508}}</ref><ref>Mai, Safiah H. C†; Khan, Momina*†; Dwivedi, Dhruva J.†‡; Ross, Catherine A.§; Zhou, Ji†; Gould, Travis J†; Gross, Peter L†‡; Weitz, Jeffrey I†‡; Fox-Robichaud, Alison E†‡∥; Liaw, Patricia C†‡∥ for the Canadian Critical Care Translational Biology Group. Delayed but not Early Treatment with DNase Reduces Organ Damage and Improves Outcome in a Murine Model of Sepsis. Shock: August 2015 - Volume 44 - Issue 2 - p 166–172
{{doi|10.1097/SHK.0000000000000396}}</ref>

'''[[Medicine:Lupus|Systemic lupus erythematosus]] (SLE)''' is an [[Medicine:Autoimmune disease|autoimmune disease]] that results in auto-antibody generation causing inflammation that results in damage to organs, joints, and kidneys. SLE has been linked with low levels of DNase I as [[Biology:Apoptosis|apoptotic cells]] become self-[[Biology:Antigen|antigen]]s in this disease. DNase I has been investigated as a possible treatment to decrease the amount of apoptotic debris in the human system. It has been suggested that their difficulty might be due to the inability for the enzyme to break down the cell membrane of [[Biology:Chromatin|chromatin]]. Studies have shown conflicting results on this treatment, however, further research is being conducted to examine the therapeutic benefits of DNase I.<ref name=":7" /><ref name=":8" /><ref>Fernando Martínez Valle, Eva Balada, Josep Ordi-Ros, Miquel Vilardell-Tarres, ''DNase 1 and systemic lupus erythematosus'', Autoimmunity Reviews, Volume 7, Issue 5,2008, Pages 359–363, {{ISSN|1568-9972}}, {{doi|10.1016/j.autrev.2008.02.002}}.</ref>

'''Anti-tumor treatment.''' DNase is known to hold anti-tumor effects due to its ability to break down DNA. High levels of DNA are found to be in cancer patients' blood, suggesting that DNase I might be a possible treatment. There is still a lack of understanding as to why there are such high levels of ecDNA and whether or not DNase will act as an effective treatment. Several mice studies have shown positive results in anti-tumor progression utilizing intravenous DNase I. However, more investigations need to be carried out before being introduced to the public.<ref>{{Cite journal |last1=Trejo-Becerril |first1=Catalina |last2=Pérez-Cardenas |first2=Enrique |last3=Gutiérrez-Díaz |first3=Blanca |last4=De La Cruz-Sigüenza |first4=Desiree |last5=Taja-Chayeb |first5=Lucía |last6=González-Ballesteros |first6=Mauricio |last7=García-López |first7=Patricia |last8=Chanona |first8=José |last9=Dueñas-González |first9=Alfonso |date=2016-07-26 |title=Antitumor Effects of Systemic DNase I and Proteases in an ''In Vivo'' Model |journal=Integrative Cancer Therapies |language=en |volume=15 |issue=4 |pages=NP35–NP43 |doi=10.1177/1534735416631102 |pmid=27146129 |pmc=5739158 |issn=1534-7354}}</ref><ref name=":7" />

== Assays ==
DNA absorbs [[Physics:Ultraviolet|ultraviolet (UV) light]] with a wavelength of maximal absorbance near 260&nbsp;nm.  This absorption is due to the pi electrons in the [[Chemistry:Aromaticity|aromatic]] bases of the DNA.  In dsDNA, or even regions of [[Biology:RNA|RNA]] where double-stranded structure occurs, the bases are stacked parallel to each other, and the overlap of the base [[Chemistry:Molecular orbital|molecular orbital]]s leads to a decrease in [[Physics:Absorbance|absorbance]] of UV light. This phenomenon is called the [[Physics:Hypsochromic shift|hypochromic effect]]. When DNase liberates [[Biology:Nucleotide|nucleotide]]s from dsDNA, the bases are no longer stacked as they are in dsDNA, so that orbital overlap is minimized and UV absorbance increases. This increase in absorbance underlies the basis of the Kunitz unit of DNase activity. One Kunitz unit is defined as the amount of enzyme added to 1&nbsp;mg/ml salmon sperm DNA that causes an increase in absorbance of 0.001 per minute at the wavelength of 260&nbsp;nm when acting upon highly [[Chemistry:Polymerization|polymerized]] DNA at 25&nbsp;°C in a 0.1 M NaOAc (pH 5.0) buffer. The unit's name recognizes the Russian-American biochemist Moses Kunitz, who proposed the standard test in 1946.<ref>{{cite web |last1=Rowlett |first1=Russ |title=K |url=https://www.ibiblio.org/units/dictK.html |website=A Dictionary of Units of Measurement |publisher=University of North Carolina at Chapel Hill |access-date=27 October 2022}}</ref>

A standard enzyme preparation should be run in parallel with an unknown because standardization of DNA preparations and their degree of [[Chemistry:Polymerization|polymerization]] in solution is not possible.

'''Single Radial Enzyme Diffusion (SRED)'''
This simple method for DNase I activity measurement was introduced by Nadano et al. and is based on the digestion of DNA in the [[Biology:Agarose gel electrophoresis|agarose gel]] by DNase, which is present in samples punched into the gel.<ref name=":7" /> DNase activity is represented by the size of a dispensed circular well in an agarose gel layer, in which DNA stained by [[Chemistry:Ethidium bromide|ethidium bromide]] is uniformly distributed. After the incubation, a circular dark zone is formed as the enzyme diffuses from the well radially into the gel and cleaves DNA. SRED underwent many modifications, which led to an increase in sensitivity and safety, such as the replacement of ethidium bromide with [[Chemistry:SYBR Green I|SYBR Green I]] or other DNA gel stains.<ref>{{cite journal |last1=Yasuda |first1=T. |last2=Takeshita |first2=H |last3=Nakazato |first3=E. |last4=Nakajima |first4=T. |last5=Hosomi |first5=O. |last6=Nakashima |first6=Y. |last7=Kishi |first7=K. |title=Activity Measurement for Deoxyribonucleases I and II with Picogram Sensitivity Based on DNA/SYBR Green I Fluorescence. |journal=Anal. Biochem. |date=1998 |volume=255 |issue=2 |pages=274–276 |doi=10.1006/abio.1997.2496|pmid=9451514 }}</ref>

'''Colorimetric DNase I Activity Assay'''

Kinetic colorimetric DNase I activity assay is developed for the assessment of the stability of the human recombinant DNase I (Pulmozyme). The method was adjusted from a colorimetric endpoint enzyme activity assay based on the degradation of a DNA/methyl green complex.<ref>{{cite journal |last1=Horney |first1=D.L. |last2=Webster |first2=D.A. |title=Deoxyribonuclease: A sensitive assay using radial diffusion in agarose containing methyl green-DNA complex |journal=Biochim. Biophys. Acta |date=1971 |volume=247 |issue=1 |pages=54–61 |doi=10.1016/0005-2787(71)90806-9|pmid=4946282 }}</ref>

== See also ==

* Extracellular DNA (ecDNA) 
* [[Biology:Deoxyribonuclease I|Deoxyribonuclease I (DNase I)]]
* [[Biology:Deoxyribonuclease II|Deoxyribonuclease II (DNase II)]]
* [[Biology:Endonuclease|Endonuclease]]
* [[Biology:Enzyme|Enzyme]]
* [[Chemistry:Hydrolysis|Hydrolysis]] 
* [[Biology:Phosphodiester bond|Phosphodiester linkages]]

== References ==
<references />

== External links ==

* [https://meshb.nlm.nih.gov/record/ui?ui=D003851 Deoxyribonucleases] at the US National Library of Medicine [[Medicine:Medical Subject Headings|Medical Subject Headings]] (MeSH)
* [https://www.thermofisher.com/us/en/home/brands/thermo-scientific/molecular-biology/thermo-scientific-restriction-modifying-enzymes/modifying-enzymes-thermo-scientific/deoxyribonucleases-dnases.html Deoxyribonucleases (DNases)] from Thermo Fisher Scientific Inc. Website
* [https://www.sigmaaldrich.com/US/en/technical-documents/protocol/genomics/dna-and-rna-purification/deoxyribonuclease-i Deoxyribonuclease I] from [[Company:Sigma-Aldrich|Sigma-Aldrich]] Solutions website 
* [https://www.ebi.ac.uk/interpro/entry/InterPro/IPR018057/ Deoxyribonuclease I, active site] from [[Biology:InterPro|InterPro]] Website

{{Esterases}}
{{Enzymes}}

[[Category:EC 3.1.21]]

{{Sourceattribution|Deoxyribonuclease}}