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Sloan-Kettering - Projects: DNA Damage Recognition and Repair by DNA Ligases DNA is susceptible to damage caused by errors committed during replication and by environmental factors, such as radiation, oxidants, or alkylating agents. DNA ligase I is a 919-amino acid polypeptide, expressed in all tissues, which catalyzes the joining of Okazaki fragments during DNA replication and also plays a role in DNA repair. DNA ligases IIIa (922 amino acids) and IIIb (862 amino acids) are the products of a single gene; they differ in amino acid sequence only at their carboxyl termini as a consequence of alternative mRNA splicing. www.mskcc.org /mskcc/html/10426.cfm   (1788 words)

DNA ligase - Wikipedia, the free encyclopedia The mechanism of DNA ligase in connecting broken DNA strands is to form covalent phosphodiester bonds between 3' hydroxyl ends of one nucleotide with the 5' phosphate end of another. DNA ligase I: ligates Okazaki fragments during lagging strand DNA replication and some recombinant fragments. DNA ligase III: complexes with DNA repair protein XRCC1 to aid in sealing base excision mutations and recombinant fragments. en.wikipedia.org /wiki/DNA_ligase   (476 words)

Ligase - Wikipedia, the free encyclopedia In biochemistry, a ligase (from the Latin verb ligāre — "to bind" or "to glue together") is an enzyme that can catalyse the joining of two molecules by forming a new chemical bond, with accompanying hydrolysis of adenosine triphosphate or other similar molecules. The common names of ligase enzymes often include the word "ligase", such as DNA ligase, an enzyme commonly used in molecular biology laboratories to join together DNA fragments. Ligases are classified as EC 6 in the EC number classification of enzymes. en.wikipedia.org /wiki/Ligase   (263 words)

DNA ligase: Facts and details from Encyclopedia Topic   (Site not responding. Last check: ) DNA ligase: Facts and details from Encyclopedia Topic In biochemistry, a ligase is an enzyme that can catalyse the joining of two molecules ("ligation" or "glue together") by forming a new chemical bond, with... A polymerase (ec 2.7.7.6/7/19/48/49) is an enzyme whose central function is associated with polymers of nucleic acids such as rna and dna.... www.absoluteastronomy.com /encyclopedia/d/dn/dna_ligase2.htm   (445 words)

DNA Ligase DNA ligases catalyze formation of a phosphodiester bond between the 5' phosphate of one strand of DNA and the 3' hydroxyl of the another. A DNA ligase from E. coli is also available, but is not commonly used. Additional information on the use of T4 DNA ligase is presented in the section on DNA ligation. arbl.cvmbs.colostate.edu /hbooks/genetics/biotech/enzymes/ligase.html   (139 words)

ChemCases: Cisplatin - 14. DNA The three-dimensional structure of DNA was deduced in 1953 by James Watson and Francis Crick. One of these is DNA polymerase I, which catalyzes the step-by-step addition of activated deoxyribonucleotide units to a DNA chain; the activated deoxyribonucleotide units for adenine, guanine, thymine, and cytosine are represented as dATP, dGTP, dTTP, and dCTP, respectively. DNA is damaged in a variety of ways, and all cells have mechanisms by which to repair the damage. www.chemcases.com /cisplat/cisplat14.htm   (1201 words)

DNA Ligase, T4 T4 DNA ligase is primarily used in joining DNA molecules with compatible cohesive termini, or blunt ended double stranded DNA to one another or to synthetic linkers. The joining of DNA fragments with protruding 5' termini that are not compatible (for instance, restriction digestion of DNA with Xba I and Hind III) can be accomplished by partial filling of the recessed 3' termini in controlled reactions using the Klenow fragment of E. The assay of T4 DNA ligase is based on the capacity of the enzyme to catalyze an exchange reaction between pyrophosphate and ATP. dwb.unl.edu /Teacher/NSF/C08/C08Links/www.worthington-biochem.com/manual/D/DNAT4L.html   (622 words)

DNA Structure, Replication and Eukaryotic Chromatin Structure> DNA replication is semi-conservative, one strand serves as the template for the second strand. DNA single-stranded binding proteins - These proteins bind to the DNA as a tetramer and stabilize the single-stranded structure that is generated by the action of the helicases. DNA Ligase - Nicks occur in the developing molecule because the RNA primer is removed and synthesis proceeds in a discontinuous manner on the lagging strand. www.ndsu.nodak.edu /instruct/mcclean/plsc431/eukarychrom/eukaryo2.htm   (727 words)

Taq DNA Ligase, Ligases, Intl, NEB Taq DNA Ligase catalyzes the formation of a phosphodiester bond between juxtaposed 5´ phosphate and 3´ hydroxyl termini of two adjacent oligonucleotides which are hybridized to a complementary target DNA. Taq DNA Ligase is active at elevated temperatures (45°C-65°C) (1,2). Taq DNA Ligase is not a substitute for T4 DNA Ligase. www.neb.com /nebecomm/products_intl/productM0208.asp   (787 words)

Ampligase® Thermostable DNA Ligase, EPICENTRE® Biotechnologies Thermostable DNA Ligase catalyzes NAD-dependent ligation of adjacent 3´-hydroxylated and 5´-phosphorylated termini in duplex DNA structures that are stable at high temperatures. Ampligase DNA Ligase is used in a two-step thermal cycling reaction that generates oligonucleotide multimers when nucleotide repeats are present in a DNA template. Ampligase DNA Ligase is free of detectable exo- and endonuclease and RNase activities. www.epibio.com /item.asp?ID=265&CatID=4&SubCatID=38   (732 words)

DNA Replication DNA replication begins with the "unzipping" of the parent molecule as the hydrogen bonds between the base pairs are broken. A molecule of a DNA polymerase binds to one strand of the DNA and begins moving along it in the 3' to 5' direction, using it as a template for assembling a leading strand of nucleotides and reforming a double helix. DNA replication begins at a single, fixed location in this molecule, the replication origin, proceeds at about 1000 nucleotides per second, and thus is done in no more than 40 minutes. users.rcn.com /jkimball.ma.ultranet/BiologyPages/D/DNAReplication.html   (845 words)

Genetic repair mechanism clears the way for sealing DNA breaks The enzyme, DNA ligase, repairs the millions of DNA breaks generated during the normal course of a cell's life, for example, linking together the abundant DNA fragments formed during replication of the genetic material in dividing cells. DNA is surprisingly reactive and under continuous assault from environmental toxins and reactive cellular metabolites. The closed conformation of DNA ligase bound to DNA was imaged in a separate study previously reported by Ellenberger's group. www.physorg.com /news80487896.html   (750 words)

BCH5425 Molecular Biology and Biotechnology   (Site not responding. Last check: ) DNA may have to be prepared from E. coli strains which are dam-dcm- in order to be cut by these enzymes. However, this chain termination by incorporation of dideoxy nucleotides is the basis of the Sanger method of DNA sequencing, as well as therapies to try to inhibit viral replication. Ligases catalyze the formation of a phosphodiester bond between juxtaposed 5' phosphate and 3' hydroxyl termini of nucleotides (potentially RNA or DNA depending on the ligase). wine1.sb.fsu.edu /bch5425/lect04/lect04.htm   (1379 words)

Lucigen Corporation Recombinant T4 DNA Ligase, Cloning Qualified   (Site not responding. Last check: ) Lucigen's T4 DNA Ligase is a recombinant enzyme formulated and tested for high activity in blunt- and cohesive-end ligations for cloning and linker addition. DNA Ligase is effective for cloning cDNAs, PCR products and genomic DNAs using plasmid, cosmid or BAC vectors in all cloning and library construction projects. Recombinant T4 DNA Ligase, Cloning Qualifi ed is supplied at 2,000 U/ml in a storage buffer of 50% glycerol, 50 mM Tris-HCl (pH 7.5), 100 mM NaCl, 0.1mM EDTA, 1 mM DTT, 0.1% Triton X-100. www.lucigen.com /products/Enzymes/T4DNAligase.html   (280 words)

Mapping and Use of a Sequence that Targets DNA Ligase I to Sites of DNA Replication In Vivo -- Cardoso et al. 139 (3): ... DNA ligase I with the affinity-purified rabbit antibodies. I fused at the COOH terminus of the GFP. DNA ligase I is associated with the 21 S complex of enzymes for DNA synthesis in HeLa cells. www.jcb.org /cgi/content/full/139/3/579   (6894 words)

RCSB Protein Data Bank   (Site not responding. Last check: ) If one strand in the DNA breaks, it is not a disaster, but it can lead to problems when the DNA double helix is unwound during the processes of transcription and replication. DNA ligase reconnects DNA strands when they are broken. In both cases, a lysine in the DNA ligase forms a bond to the phosphate in the cofactor, holding onto the AMP portion and discarding the rest. pdbbeta.rcsb.org /pdb/static.do?p=education_discussion/molecule_of_the_month/pdb55_1.html   (580 words)

Binding of Nucleotides by T4 DNA Ligase and T4 RNA Ligase: Optical Absorbance and Fluorescence Studies -- Cherepanov ... DNA ligase and RNA ligase from the bacteriophage T4 catalyze the formation of the phosphodiester bond between two molecules T4 DNA ligase was incubated both in the presence and the absence of 0.05 mg/ml BSA, at +4°C and +20°C. The enzyme was diluted in the reaction buffer to a final concentration of 1 U/ml (~6 nM). The activity was measured after addition of 1 µl of the T4 DNA ligase incubation mixture to 14 µl of standard assay containing 1 µM of the nicked dsDNA. www.biophysj.org /cgi/content/full/81/6/3545   (7081 words)

Drosophila gene families: DNA replication enzymes and protein cofactors DNA synthesis is initiated by the bifunctional pol-alpha:primase complex, a heterotetrameric phosphoprotein. The pol-alpha:primase serves exclusively to initiate DNA synthesis on the lagging strand, and dissociation from the DNA provides a primer terminus for assembly of the PCNA/pol-delta complex, which serves to extend the RNA/DNA primers originally synthesized by pol-alpha:primase. The heterodimeric DNA polymerase-delta is involved in the elongation stage of DNA replication, acting both on the leading and lagging strands. www.flybase.org /allied-data/lk/interactive-fly/aignfam/dnarep.htm   (999 words)

Evaluation of NAD+-Dependent DNA Ligase of Mycobacteria as a Potential Target for Antibiotics -- Korycka-Machala et al. ... ATP-dependent DNA ligases are encoded in the genomes of mycobacteria, DNA ligases confirmed by in vitro studies (10, 27, 28, 33). DNA ligase and the pyridine nucleotide cycle in Salmonella typhimurium. aac.asm.org /cgi/content/full/51/8/2888   (4216 words)

DNA Ligase III antibody [6G9] (ab587) datasheet   (Site not responding. Last check: ) If DNA Ligase III antibody [6G9] (ab587) does not perform as described on this datasheet, notify us within 90 days of delivery using this form, so that we can help you or offer you a replacement or a refund. DNA Ligase III gene product migrates as a 100kDa (a-form) protein in somatic tissues, is ubiquitously expressed and complexes with XRCC-1 and a 97kDa (ß-form) protein that is found in testis, and is involved in rejoining DNA single-strand breaks and in the base excision repair pathway. Role of the DNA ligase III zinc finger in polynucleotide binding and ligation. www.abcam.com /?datasheet=587   (585 words)

Cloning and Functional Characterization of an NAD+-Dependent DNA Ligase from Staphylococcus aureus -- Kaczmarek et al. ... aureus DNA ligase with thermolysin and purification of the DNA ligase fragments are Sequence of the DNA ligase-encoding gene from Thermus scotoductus and conserved motifs in DNA ligases. Eukaryotic DNA ligase: purification and properties of the enzyme from bovine thymus, and immunochemical studies of the enzyme from animal tissues. jb.asm.org /cgi/content/full/183/10/3016   (4943 words)

T4 DNA Ligase  T4 DNA Ligase catalyzes the formation of a phosphodiester bond between juxtaposed 5'-phosphate and 3'-hydroxyl termini in duplex DNA or RNA. One cohesive-end ligation unit is defined as the amount of enzyme required to give 50% ligation of HindIII fragments of Lambda DNA in 30min at 16°C in 20µl of the assay mixture (50mM Tris-HCl, pH 7.5), 10mM MgCl2, 10mM DTT, 1mM ATP, 25µg/ml BSA and a 5'-DNA termini concentration of 0.12µM (300µg/ml). The ratio of Weiss unit to cohesive-end ligation unit is determined by conversion of [5'-33P]-labeled termini of HindIII fragments of Lambda DNA to a phosphates-resistant form. www.cinnagen.com /T4_DNA_Ligase.htm   (273 words)

Ligases: T4 DNA Ligases The T4 DNA Ligase catalyzes the formation of a phosphodiester bond between juxtaposed 5'-phosphate and 3'-hydroxyl termini in duplex DNA or RNA with blunt or cohesive-end termini. The T4 DNA Ligase binding to DNA may result in a band shift in agarose gels. It is necessary to remove the enzyme from the ligation mixture by chloroform extraction prior to electro-transformation of bacterial cells with DNA. www.fermentas.com /catalog/modifyingenzymes/t4dnaligase.htm   (654 words)

RCSB Protein Data Bank If one strand in the DNA breaks, it is not a disaster, but it can lead to problems when the DNA double helix is unwound during the processes of transcription and replication. Polymerases only work in one direction when they copy a DNA strand, so one of the two strands is copied in a series of small pieces that need to be linked up into a continuous strand. In both cases, a lysine in the DNA ligase forms a bond to the phosphate in the cofactor, holding onto the AMP portion and discarding the rest. www.rcsb.org /pdb/static.do?p=education_discussion/molecule_of_the_month/pdb55_1.html   (580 words)

Dynamic mechanism of nick recognition by DNA ligase -- Cherepanov and de Vries 269 (24): 5993 -- FEBS Journal   (Site not responding. Last check: ) DNA ligases are the enzymes responsible for the repair of single-stranded The AMP moiety of Chlorella virus DNA ligase adenylate is shown in purple. B-form DNA is colored yellow, A-DNA is blue and the enzyme is shown in green. content.febsjournal.org /cgi/content/full/269/24/5993   (4776 words)

Recombinant DNA Technology Problem Set   (Site not responding. Last check: ) DNA ligase - enzyme that cuts DNA, creating sticky ends. A sample of human DNA is also digested with EcoRI to produce pieces with the same sticky ends. DNA ligase is used to covalently link the fragments. www.biology.arizona.edu /molecular_bio/problem_sets/Recombinant_DNA_Technology/02t.html   (136 words)

The Human DNA Ligase III Gene Encodes Nuclear and Mitochondrial Proteins -- Lakshmipathy and Campbell 19 (5): 3869 -- ... Expression constructs harboring HA-tagged DNA ligase III proteins were used to transiently transfect Cos-1 cells, and subcellular localization was visualized by immunocytochemistry using fluorescein-labeled anti-HA monoclonal antibody (confocal images of cells transfected with pLIG-FL-1 or pLIG-FL-2 are displayed beneath the expression constructs). DNA end-joining reactions were carried out on 5 µg of mitochondrial extracts prepared from human cells transfected with pREP4 (lane c) or pREP4dnlAS (lane a) as described in Materials and Methods. A DNA ligase from mitochondria of rat liver Biochem. mcb.asm.org /cgi/content/full/19/5/3869   (5060 words)

Efficient RNA 5'-adenylation by T4 DNA ligase to facilitate practical applications -- Wang and Silverman 12 (6): 1142 ... The yields of 5'-AppRNA for DNA overhang lengths of 1 nt, 5 nt, and 10 nt were 8%, 26%, and 37% under these conditions, indicating that the longer 10-nt overhang is preferred. A 20-nt DNA overhang led to approximately the same 5'-adenylation yield as a 10-nt overhang (data not shown). The corresponding DNA oligo was a 60-mer complementary to nt 102–151 www.rnajournal.org /cgi/content/full/12/6/1142   (2972 words)

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