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DpnI
DpnI (pronounced "''D-P-N one''") is a Type IIM restriction endonuclease isolated from ''Streptococcus pneumonae'' (formerly ''Diplococcus pneumonae''). It recognizes and cuts methylated DNA at the sequence G m6A↓TC. Structure The structure of DpnI comprises an N-terminal catalytic domain and a C-terminal winged helix DNA binding domain, both of which show specificity for the methylated GATC sequence. The catalytic domain is disordered in solution and becomes ordered upon binding DNA. Uses in molecular biology DpnI is commonly used to digest template DNA after site-directed mutagenesis. Most strains of ''E. coli'' used in molecular biology express Dam methylase, a protein that methylates DNA at the sequence GATC. Adding DpnI to the product of a PCR reaction digests only the template DNA, as the template DNA was isolated from ''E. coli'' and will have methylation at this sequence while the newly synthesized DNA will not. DpnI is widely available commercially, both alone ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
DNA Adenine Methyltransferase Identification
DNA adenine methyltransferase identification, often abbreviated DamID, is a molecular biology protocol used to map the binding sites of DNA- and chromatin-binding proteins in eukaryotes. DamID identifies binding sites by expressing the proposed DNA-binding protein as a fusion protein with DNA methyltransferase. Binding of the protein of interest to DNA localizes the methyltransferase in the region of the binding site. Adenine methylation does not occur naturally in eukaryotes and therefore adenine methylation in any region can be concluded to have been caused by the fusion protein, implying the region is located near a binding site. DamID is an alternate method to ChIP-on-chip or ChIP-seq. Description Principle N6-methyladenine (m6A) is the product of the addition of a methyl group (CH3) at position 6 of the adenine. This modified nucleotide is absent from the vast majority of eukaryotes, with the exception of ''C. elegans'', but is widespread in bacterial genomes, as part of ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
DpnII Restriction Endonuclease Family
In molecular biology, the DpnII restriction endonuclease family is a family of restriction endonucleases which includes DpnII from '' Diplococcus pneumoniae''. These enzymes recognise the double-stranded DNA unmethylated sequence In mathematics, a sequence is an enumerated collection of objects in which repetitions are allowed and order matters. Like a set, it contains members (also called ''elements'', or ''terms''). The number of elements (possibly infinite) is cal ... GATC and cleave before G-1, where it encompasses the full length of the protein. References {{restriction enzyme Protein families ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Restriction Enzyme
A restriction enzyme, restriction endonuclease, REase, ENase or'' restrictase '' is an enzyme that cleaves DNA into fragments at or near specific recognition sites within molecules known as restriction sites. Restriction enzymes are one class of the broader endonuclease group of enzymes. Restriction enzymes are commonly classified into five types, which differ in their structure and whether they cut their DNA enzyme substrate (biology), substrate at their recognition site, or if the recognition and cleavage sites are separate from one another. To cut DNA, all restriction enzymes make two incisions, once through each backbone chain, sugar-phosphate backbone (i.e. each strand) of the DNA double helix. These enzymes are found in bacteria and archaea and provide a defense mechanism against invading viruses. Inside a prokaryote, the restriction enzymes selectively cut up ''foreign'' DNA in a process called ''restriction digestion''; meanwhile, host DNA is protected by a modification ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Site-directed Mutagenesis
Site-directed mutagenesis is a molecular biology method that is used to make specific and intentional mutating changes to the DNA sequence of a gene and any gene products. Also called site-specific mutagenesis or oligonucleotide-directed mutagenesis, it is used for investigating the structure and biological activity of DNA, RNA, and protein molecules, and for protein engineering. Site-directed mutagenesis is one of the most important laboratory techniques for creating DNA libraries by introducing mutations into DNA sequences. There are numerous methods for achieving site-directed mutagenesis, but with decreasing costs of oligonucleotide synthesis, artificial gene synthesis is now occasionally used as an alternative to site-directed mutagenesis. Since 2013, the development of the CRISPR/Cas9 technology, based on a prokaryotic viral defense system, has also allowed for the editing of the genome, and mutagenesis may be performed ''in vivo'' with relative ease. History Early atte ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Enzymes
An enzyme () is a protein that acts as a biological catalyst by accelerating chemical reactions. The molecules upon which enzymes may act are called substrates, and the enzyme converts the substrates into different molecules known as products. Almost all metabolic processes in the cell need enzyme catalysis in order to occur at rates fast enough to sustain life. Metabolic pathways depend upon enzymes to catalyze individual steps. The study of enzymes is called ''enzymology'' and the field of pseudoenzyme analysis recognizes that during evolution, some enzymes have lost the ability to carry out biological catalysis, which is often reflected in their amino acid sequences and unusual 'pseudocatalytic' properties. Enzymes are known to catalyze more than 5,000 biochemical reaction types. Other biocatalysts include catalytic RNA molecules, also called ribozymes. They are sometimes described as a ''type'' of enzyme rather than being ''like'' an enzyme, but even in the d ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Restriction Modification System
The restriction modification system (RM system) is found in bacteria and archaea, and provides a defense against foreign DNA, such as that borne by bacteriophages. Bacteria have restriction enzymes, also called restriction endonucleases, which cleave double-stranded DNA at specific points into fragments, which are then degraded further by other endonucleases. This prevents infection by effectively destroying the foreign DNA introduced by an infectious agent (such as a bacteriophage). Approximately one-quarter of known bacteria possess RM systems and of those about one-half have more than one type of system. As the sequences recognized by the restriction enzymes are very short, the bacterium itself will almost certainly contain some within its genome. In order to prevent destruction of its own DNA by the restriction enzymes, methyl groups are added. These modifications must not interfere with the DNA base-pairing, and therefore, usually only a few specific bases are modified ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
List Of Restriction Enzyme Cutting Sites
A restriction enzyme or restriction endonuclease is a special type of biological macromolecule that functions as part of the "immune system" in bacteria. One special kind of restriction enzymes is the class of " homing endonucleases", these being present in all three domains of life, although their function seems to be very different from one domain to another. The classical restriction enzymes cut up, and hence render harmless, any unknown (non- cellular) DNA that enters a bacterial cell as a result of a viral infection. They recognize a specific DNA sequence, usually short (3 to 8 bp), and cut it, producing either blunt or overhung ends, either at or nearby the recognition site. Restriction enzymes are quite variable in the short DNA sequences they recognize. An organism often has several different enzymes, each specific to a distinct short DNA sequence. __TOC__ Restriction enzymes catalog The list includes some of the most studied examples of restriction endoncleases. ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Kinase
In biochemistry, a kinase () is an enzyme that catalyzes the transfer of phosphate groups from high-energy, phosphate-donating molecules to specific substrates. This process is known as phosphorylation, where the high-energy ATP molecule donates a phosphate group to the substrate molecule. As a result, kinase produces a phosphorylated substrate and ADP. Conversely, it is referred to as dephosphorylation when the phosphorylated substrate donates a phosphate group and ADP gains a phosphate group (producing a dephosphorylated substrate and the high energy molecule of ATP). These two processes, phosphorylation and dephosphorylation, occur four times during glycolysis. Kinases are part of the larger family of phosphotransferases. Kinases should not be confused with phosphorylases, which catalyze the addition of inorganic phosphate groups to an acceptor, nor with phosphatases, which remove phosphate groups (dephosphorylation). The phosphorylation state of a molecule, whether it ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Ligase
In biochemistry, a ligase is an enzyme that can catalyze the joining ( ligation) of two molecules by forming a new chemical bond. This is typically via hydrolysis of a small pendant chemical group on one of the molecules, typically resulting in the formation of new C-O, C-S, or C-N bonds. For example, DNA ligase can join two complementary fragments of nucleic acid by forming phosphodiester bonds, and repair single stranded breaks that arise in double stranded DNA during replication. In general, a ligase catalyzes the following dehydration reaction, thus joining molecules A and B: A-OH + B-H → A–B + H2O Nomenclature The naming of ligases is inconsistent and so these enzymes are commonly known by several different names. Generally, the common names of ligases include the word "ligase", such as in DNA ligase, an enzyme commonly used in molecular biology laboratories to join together DNA fragments. However, many common names use the term "synthetase" or "synthase" instead, ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Streptococcus Pneumoniae
''Streptococcus pneumoniae'', or pneumococcus, is a Gram-positive, spherical bacteria, hemolysis (microbiology), alpha-hemolytic member of the genus ''Streptococcus''. ''S. pneumoniae'' cells are usually found in pairs (diplococci) and do not form Bacterial morphological plasticity, spores and are non motile. As a significant human pathogenic bacterium ''S. pneumoniae'' was recognized as a major cause of pneumonia in the late 19th century, and is the subject of many humoral immunity studies. ''Streptococcus pneumoniae'' resides asymptomatically in healthy carriers typically colonizing the respiratory tract, sinuses, and nasopharynx, nasal cavity. However, in susceptible individuals with immunocompromised, weaker immune systems, such as the elderly and young children, the bacterium may become pathogenic and spread to other locations to cause disease. It spreads by direct person-to-person contact via respiratory droplets and by auto inoculation in persons carrying the bacteria in ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
DNA Adenine Methylase
DNA adenine methylase, (Dam) (also site-specific DNA-methyltransferase (adenine-specific), , ''modification methylase'', ''restriction-modification system'') is an enzyme that adds a methyl group to the adenine of the sequence 5'-GATC-3' in newly synthesized DNA. Immediately after DNA synthesis, the daughter strand remains unmethylated for a short time. It is an orphan methyltransferase that is not part of a restriction-modification system and regulates gene expression. This enzyme catalyses the following chemical reaction : S-adenosyl-L-methionine + DNA adenine \rightleftharpoons S-adenosyl-L-homocysteine + DNA 6-methylaminopurine This is a large group of enzymes unique to prokaryotes and bacteriophages. The ''E. coli'' DNA adenine methyltransferase enzyme (Dam), is widely used for the chromatin profiling technique DamID, in which the Dam is fused to a DNA-binding protein of interest and expressed as a transgene in a genetically tractable model organism to identify protein bi ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
