T4 polynucleotide kinase

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T4 polynucleotide kinase
T4 polynucleotide kinase
according to PDB  1LTQ
other names

T4 polynucleotide kinase, T4-PNK, T4 deoxynucleotide 3'-phosphatase, Enterobacteria phage T4 polynucleotide kinase

Existing structure data : PDB  1LY1 , PDB  1RC8

Mass / length primary structure 301 amino acids , 34,620 Da
Identifier
External IDs
Enzyme classification
EC, category 2.7.1.78
Orthologue (enterobacteria phage T4)
Entrez 1258796
UniProt P06855
Refseq (protein) NP_049834.1
PubMed search 1258796

T4 polynucleotide kinase (T4-PNK) is an enzyme from the group of polynucleotide kinases and comes from the bacteriophage T4 .

properties

T4 polynucleotide kinase has three enzyme activities: a 5'-hydroxyl kinase , a 3'- phosphatase and a 2 ', 3'-cyclic phosphodiesterase . Among other things, it catalyzes the transfer of a phosphate group from adenosine triphosphate (ATP) to 5'-hydroxyl groups of nucleotides and deoxyribonucleotides . In the presence of adenosine diphosphate (ADP) the reaction goes both ways. In the bacteriophage, the T4 polynucleotide kinase is used for immune evasion during the infection of bacteria through a counter reaction to the T4-induced anticodon nuclease , which otherwise interrupts the translation of viral proteins in a T4 infection by breaking down tRNA . The nicks generated by the T4-induced anticodon nuclease are repaired with the T4 DNA ligase following phosphorylation by the T4 polynucleotide kinase . The T4 polynucleotide kinase transfers a phosphate group to tRNA and other nucleic acids with nicks . The pH optimum of the T4 PNK is 6. Cofactors are divalent magnesium ions .

ATP + 5'-dephospho-DNA ADP + 5'-phospho-DNA

Deoxyribonucleoside 3'-phosphate + H 2 O deoxyribonucleoside + phosphate.

Applications

The T4-PNK is used in molecular biology for the phosphorylation of DNA in the course of cloning , which facilitates the subsequent ligation of DNA with the T4-DNA ligase. It is also used to mark DNA at the 5 'end ( end labeling ).

literature

  • ES Miller, E. Kutter, G. Mosig, F. Arisaka, T. Kunisawa, W. Rüger: Bacteriophage T4 genome. In: Microbiology and molecular biology reviews: MMBR. Volume 67, Number 1, March 2003, pp. 86-156, table of contents, PMID 12626685 , PMC 150520 (free full text).

Web links

Individual evidence

  1. ↑ M. Amitsur, R. Levitz, G. Kaufmann: Bacteriophage T4 anticodon nuclease, polynucleotide kinase and RNA ligase reprocess the host lysine tRNA. In: The EMBO journal. Volume 6, Number 8, August 1987, pp. 2499-2503, PMID 2444436 , PMC 553660 (free full text).
  2. ↑ Z. Topcu: An optimized recipe for cloning of the polymerase chain reaction-amplified DNA inserts into plasmid vectors. In: Acta biochimica Polonica. Volume 47, Number 3, 2000, pp. 841-846, PMID 11310983 .
  3. ↑ A. Novogrodsky, J. Hurwitz: The enzymatic phosphorylation of ribonucleic acid and deoxyribonucleic acid. I. Phosphorylation at 5'-hydroxyl termini. In: The Journal of biological chemistry. Volume 241, Number 12, June 1966, pp. 2923-2932, PMID 4287929 .