Key Difference – T4 vs T7 DNA Ligase
DNA ligaseis an importantenzymeused in molecular biology techniques. It functions as a molecular glue to joinnucleotidesby forming phosphodiester bonds between nucleotides. The phosphodiester bonds are formed between the 3’hydroxylend of the sugar moiety and the 5’ endphosphategroup. DNA ligase is vital in the replication process to join the Okazaki fragments of the lagging strand and during DNA repair mechanisms andin vitrocloning experiments to join the desired gene of interest to thevectorgenome. There are two main DNA ligases used currently by molecular biologists: T4 and T7 DNA ligases.T4 DNA ligase is one of the first enzymes to be isolated from the T4bacteriophage. T7 DNA ligase, which is a smaller protein, is an enzyme isolated from T7 bacteriophage. This is the key difference between T4 and T7 DNA ligases.
CONTENTS
1.Overview and Key Difference
2.What is the Action of DNA Ligase
3.What is T4 DNA Ligase
4.What is T7 DNA Ligase
4.Similarities Between T4 and T7 DNA Ligase
5.Side by Side Comparison – T4 vs T7 DNA Ligase in Tabular Form
6.Summary
What is the Action of DNA Ligase?
DNA结扎发生在三步反应。的first step is a nucleophilic attack on the alpha-phosphorus ofATPgenerating a covalent enzyme-adenylate intermediate in which AMP is linked to the terminal side-chain nitrogen oflysine. In the second step, a nucleophilic attack occurs on the phosphorus of the enzyme-adenylate by the 5’-phosphate-terminated DNA substrate, releasing lysine and forming DNA-adenylate. In the final step, DNA-adenylate is attacked by 3’- OH of another DNA strand, releasing AMP and joining the polynucleotides.
What is T4 DNA Ligase?
T4 Ligase was the first ligase to be isolated and identified by Meselson, Weigle, and Kellenberger and was the first ever commercial ligase produced. T4 DNA ligase is an ATP dependent enzyme which catalyzes the phosphodiester bond formation. It is a singlepolypeptidechain with 487 amino acid residues and has a molecular weight of about 77 kDa. The optimum pH for its activity lies in the range of 7.5 – 8.
T4 DNA Ligase, as its name suggests, is present in bacteriophage T4 and was isolated when phage produced the enzyme upon infection withE. coli.的enzyme ligates double stranded DNA duplexes with blunt ends and the fragments to be bound should be in close proximity to each other.
What is T7 DNA Ligase?
T7 ligase is 41 kDa protein produced by T7 Bacteriophage and is isolated by infectingE. colicells. The main function of T7 ligase is to ligate DNA duplexes through phosphodiester linkages and join nearby fragments of DNA during cloning experiments. The optimum pH for T7 DNA ligase lies in the range of 7.0 – 7.2. T7 ligase is ATP dependent. T7 DNA ligases are also capable of annealing DNA and RNA hybrids, giving rise to abnormal conformations during transcription; however, this phenomenon is still under research byin vitrostudies.
What are the Similarities Between T4 and T7 DNA Ligase?
- T4 and T7 DNA Ligase participate in a ligation reaction by annealing DNA duplexes through phosphodiester bond formation between 3’ OH end and the 5’ phosphate end of the DNA fragment.
- Both enzymes ligateblunt endedDNA duplexes which are in close proximity to each other.
- Both enzymes are ATP dependent.
- 的se enzymes are isolated by infectingE. colicells with the particular bacteriophage.
- Applications of both enzymes include:
- Molecularcloning– to ligate the gene of interest to the vector
- DNA repair – to repair double stranded niches
- In vitroDNA replication
- Ligase chain reaction
What is the Difference Between T4 and T7 DNA Ligase?
T4 vs T7 DNA Ligase |
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T4 DNA Ligase type of DNA ligase isolated from the bacteriophage T4 and is involved in ligating adjacent DNA fragments via phosphodiester bond formation. | T7 DNA Ligase type of DNA ligase isolated from the bacteriophage T7 and is involved in ligating adjacent DNA fragments via phosphodiester bond formation. |
Size of the Protein | |
Size of T4 DNA ligase is 77 kDa – larger compared to T7 DNA ligase. | Size of T7 DNA ligase is 41 kDa. |
Optimum pH | |
pH range which works T4 DNA ligase is 7.5 – 8.0. | pH range which works T7 DNA ligase is 7.0 – 7.2. |
Summary – T4 vs T7 DNA Ligase
DNA ligases are an important class of enzymes involved in molecular cloning to producerecombinant DNAmolecules containing the gene of interest. Ligation reaction is an energy utilizing reaction and there are novel research studies underway to improve the accuracy via modulating the available ligases. T4 and T7 DNA ligases are two types of important DNA ligase enzymes used in molecular biological techniques. T7 DNA ligase is also a result of research which was based on the primary ligase isolated from T4 infectedE. coli.T7 is portrayed more efficiency and accuracy in carrying out its function due to its smaller size. This is the difference between T4 and T7 ligase.
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References:
1. “DNA Ligase, T4.” DNA Ligase, T4 – Worthington Enzyme Manual,Available here.Accessed 22 Aug. 2017.
2. Doherty, Aidan J., et al. “Bacteriophage T7 DNA Ligase OVEREXPRESSION, PURIFICATION, CRYSTALLIZATION, AND CHARACTERIZATION.” Journal of Biological Chemistry, 10 May 1996,Available here. Accessed 22 Aug. 2017.
Image Courtesy:
1. “Figure 3. Splint Aid Single Stranded mRNA-DNA T4 DNA Ligase Ligation.”(CC BY 3.0)viaCommons Wikimedia
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