Recombinant Human Bifunctional polynucleotide phosphatase/kinase (PNKP)

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Code CSB-EP857031HU
MSDS
Size $224
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  • (Tris-Glycine gel) Discontinuous SDS-PAGE (reduced) with 5% enrichment gel and 15% separation gel.
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Product Details

Purity
Greater than 90% as determined by SDS-PAGE.
Target Names
PNKP
Uniprot No.
Research Area
Epigenetics and Nuclear Signaling
Alternative Names
2''(3'')-polynucleotidase; 2'(3')-polynucleotidase; Bifunctional polynucleotide phosphatase/kinase; DEM 1; DEM1; DNA 5' kinase/3' phosphatase; DNA 5''-kinase/3''-phosphatase; EIEE10; Homo sapiens polynucleotide kinase 3' phosphatase (PNKP); MCSZ; PNK 1; PNK1; Pnkp; PNKP DNA kinase; PNKP_HUMAN; Polynucleotide 3'-phosphatase; Polynucleotide 5' hydroxyl kinase; Polynucleotide 5''-hydroxyl-kinase; Polynucleotide kinase 3 prime phosphatase; Polynucleotide kinase 3' phosphatase; Polynucleotide Kinase; Polynucleotide kinase-3''-phosphatase
Species
Homo sapiens (Human)
Source
E.coli
Expression Region
1-521aa
Target Protein Sequence
MGEVEAPGRLWLESPPGGAPPIFLPSDGQALVLGRGPLTQVTDRKCSRTQVELVADPETRTVAVKQLGVNPSTTGTQELKPGLEGSLGVGDTLYLVNGLHPLTLRWEETRTPESQPDTPPGTPLVSQDEKRDAELPKKRMRKSNPGWENLEKLLVFTAAGVKPQGKVAGFDLDGTLITTRSGKVFPTGPSDWRILYPEIPRKLRELEAEGYKLVIFTNQMSIGRGKLPAEEFKAKVEAVVEKLGVPFQVLVATHAGLYRKPVTGMWDHLQEQANDGTPISIGDSIFVGDAAGRPANWAPGRKKKDFSCADRLFALNLGLPFATPEEFFLKWPAAGFELPAFDPRTVSRSGPLCLPESRALLSASPEVVVAVGFPGAGKSTFLKKHLVSAGYVHVNRDTLGSWQRCVTTCETALKQGKRVAIDNTNPDAASRARYVQCARAAGVPCRCFLFTATLEQARHNNRFREMTDSSHIPVSDMVMYGYRKQFEAPTLAEGFSAILEIPFRLWVEPRLGRLYCQFSEG
Note: The complete sequence including tag sequence, target protein sequence and linker sequence could be provided upon request.
Mol. Weight
73.1kDa
Protein Length
Full Length
Tag Info
N-terminal 6xHis-SUMO-tagged
Form
Liquid or Lyophilized powder
Note: We will preferentially ship the format that we have in stock, however, if you have any special requirement for the format, please remark your requirement when placing the order, we will prepare according to your demand.
Buffer
If the delivery form is liquid, the default storage buffer is Tris/PBS-based buffer, 5%-50% glycerol.
Note: If you have any special requirement for the glycerol content, please remark when you place the order.
If the delivery form is lyophilized powder, the buffer before lyophilization is Tris/PBS-based buffer, 6% Trehalose, pH 8.0.
Reconstitution
We recommend that this vial be briefly centrifuged prior to opening to bring the contents to the bottom. Please reconstitute protein in deionized sterile water to a concentration of 0.1-1.0 mg/mL.We recommend to add 5-50% of glycerol (final concentration) and aliquot for long-term storage at -20°C/-80°C. Our default final concentration of glycerol is 50%. Customers could use it as reference.
Troubleshooting and FAQs
Storage Condition
Store at -20°C/-80°C upon receipt, aliquoting is necessary for mutiple use. Avoid repeated freeze-thaw cycles.
Shelf Life
The shelf life is related to many factors, storage state, buffer ingredients, storage temperature and the stability of the protein itself.
Generally, the shelf life of liquid form is 6 months at -20°C/-80°C. The shelf life of lyophilized form is 12 months at -20°C/-80°C.
Lead Time
3-7 business days
Notes
Repeated freezing and thawing is not recommended. Store working aliquots at 4°C for up to one week.
Datasheet & COA
Please contact us to get it.
Description

Intact human bifunctional polynucleotide phosphatase/kinase (PNKP) cDNA (1-521aa) with an N-terminal 6xHis-SUMO-tag was expressed in the E.coli. The forming protein is the recombinant full-length human PNKP protein. The purity of this protein is greater than 90% determined by SDS-PAGE. Under reducing conditions, the gel showed a molecular weight band of about 90 kDa. This recombinant PNKP protein may be applied for specific antibody production or in the studies of epigenetics and nuclear signaling.

PNKP is a DNA repair factor possessing both DNA 5’-kinase and DNA 3’-phosphatase activities to modify the ends of a DNA break before ligation. In addition to its role in DNA repair, PNKP plays an important role in normal neurogenesis and also exerts crucial genome maintenance functions after its completion. PNKP mutations are associated with a spectrum of neurodevelopmental disorders, such as microcephaly, seizures, and developmental delay (MCSZ) to ataxia with oculomotor apraxia type 4 (AOA4).

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Target Background

Function
Plays a key role in the repair of DNA damage, functioning as part of both the non-homologous end-joining (NHEJ) and base excision repair (BER) pathways. Through its two catalytic activities, PNK ensures that DNA termini are compatible with extension and ligation by either removing 3'-phosphates from, or by phosphorylating 5'-hydroxyl groups on, the ribose sugar of the DNA backbone.
Gene References into Functions
  1. Despite presence of an alternative 3'-phosphatase, loss of PNKP significantly sensitizes cells to 3'-phosphate-terminated DSBs, due to a 3'-dephosphorylation defect. PMID: 29807321
  2. PNKP mutation in two siblings is associated with progressive ataxia, abnormal saccades, sensorimotor neuropathy and dystonia consistent with ataxia with oculomotor apraxia disorders. PMID: 28552035
  3. we have identified a mutation in PNKP, leading to a phenotype of microcephaly with primordial dwarfism. PMID: 27232581
  4. XRCC1 and PNKP interact via a high-affinity phosphorylation-dependent interaction site in XRCC1 and a forkhead-associated domain in PNKP. Data suggest a second PNKP interaction site in XRCC1 that binds PNKP with lower affinity and independently of XRCC1 phosphorylation. (XRCC1 = X-ray repair cross complementing protein 1; PNKP = polynucleotide kinase 3'-phosphatase) PMID: 28821613
  5. In a recombinant PNKP-XRCC4-LigIV complex, stable binding of PNKP requires XRCC4 phosphorylation. Only one PNKP protomer binds per XRCC4 dimer. Both the PNKP FHA and catalytic domains contact the XRCC4 coiled-coil and LigIV BRCT repeats. A surface on the PNKP phosphatase domain may contact XRCC4-LigIV. A mutation on this surface (E326K) impairs PNKP recruitment to damaged DNA and causes microcephaly with seizures. PMID: 28453785
  6. Mutations in TDP1 and APTX have been linked to Spinocerebellar ataxia with axonal neuropathy (SCAN1) and Ataxia-ocular motor apraxia 1 (AOA1), respectively, while mutations in PNKP are considered to be responsible for Microcephaly with seizures (MCSZ) and Ataxia-ocular motor apraxia 4 (AOA4). PMID: 27470939
  7. the role for PNKP in maintaining brain function and how perturbation in its activity can account for the varied pathology of neurodegeneration or microcephaly present in microcephaly with seizures and ataxia with oculomotor apraxia 4 respectively. PMID: 27125728
  8. In 11 Portuguese patients, PNKP mutations cause ataxia with oculomotor apraxia type 4. PMID: 26970421
  9. Here we report that purified wild-type (WT) ATXN3 stimulates, and by contrast the mutant form specifically inhibits, PNKP's 3' phosphatase activity in vitro. ATXN3-deficient cells also show decreased PNKP activity PMID: 25633985
  10. We now report that the mutant ATXN3 protein interacts with and inactivates PNKP (polynucleotide kinase 3'-phosphatase), an essential DNA strand break repair enzyme PMID: 25590633
  11. We identified homozygous or compound-heterozygous PNKP mutations in eight of the nine Portuguese families we studied, suggesting that, in Portugal, mutations in PNKP are the most frequent cause of ataxia with oculomotor apraxia. PMID: 25728773
  12. we show that modest inhibition of PNKP in a PTEN knockout background enhances cellular radiosensitivity, suggesting that such a "synthetic sickness" approach involving the combination of PNKP inhibition with radiotherapy PMID: 23883586
  13. Mutations in PNKP have previously been associated with a syndrome of microcephaly, seizures and developmental delay (MIM 613402), and is now associated with a neurodegenerative disorder. PMID: 23224214
  14. the interaction between PNKP and XRCC1 has roles in the retention of XRCC1 at DNA damage sites and in DNA alkylation damage repair PMID: 22992732
  15. The data suggest that all four known mutations associated with microcephaly, seizures and developmental delay reduce the cellular stability and level of PNKP protein, with three mutations likely ablating cellular DNA 5'-kinase activity and all of the mutations greatly reducing cellular DNA 3'-phosphatase activity. PMID: 22508754
  16. the critical role of NEIL2 and PNKP in maintenance of the mammalian mitochondrial genome. PMID: 22130663
  17. PNKP distorts target DNA structures to access damaged substrate DNA ends, thus providing a molecular mechanism for the involvement of PNKP in the repair of both single- and double-strand breaks. PMID: 22171004
  18. Results reveal that ionizing radiation-induced phosphorylation of PNKP by ATM and DNA-PK regulates PNKP function at DNA double strand breaks. PMID: 21824916
  19. Studies indicate that PNKP serves a crucial role in the repair of DNA strand breaks through interactions with other DNA repair proteins, notably XRCC1 and XRCC4. PMID: 21353781
  20. CK2-mediated phosphorylation of XRCC4 can have different effects on PNKP activity. PMID: 20852255
  21. The neurological abnormalities in individuals with microcephaly, early onset, intractable seizures and develomental delays may reflect a role for PNKP in several DNA repair pathways. PMID: 20118933
  22. Involvement of human polynucleotide kinase in double-strand break repair by non-homologous end joining PMID: 12032095
  23. First direct physical evidence for ternary complex formation involving a polynucleotide kinase, AMP-PNP, and an oligonucleotide, supports a reaction mechanism in which ATP and DNA bind simultaneously to the enzyme. PMID: 14556639
  24. Data show that polynucleotide kinase is associated with the PARP-1-dependent end-joining pathway, and show functional parallels between the PARP-1 and DNA-PK-dependent end-joining processes. PMID: 16364363
  25. polynucleotide kinase participates in repair of DNA double-strand breaks by nonhomologous end joining but not homologous recombination PMID: 17638872
  26. XRCC1 enhances the capacity of hPNK to discriminate between strand breaks with 5'-OH termini and those with 5'-phosphate termini; and XRCC1 stimulates hPNK activity by displacing hPNK from the phosphorylated DNA product PMID: 17650498
  27. The PNKP T5644G variant does not seem to be involved in adenoma recurrence in the Polyp Prevention Trial. PMID: 18414202
  28. the FHA domain of PNK binds specifically, and with high affinity to a multiply phosphorylated motif in XRCC1 containing a pSer-pThr dipeptide, and forms a 2:1 PNK:XRCC1 complex. PMID: 19155274

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Involvement in disease
Microcephaly, seizures, and developmental delay (MCSZ); Ataxia-oculomotor apraxia 4 (AOA4)
Subcellular Location
Nucleus. Chromosome.
Protein Families
DNA 3' phosphatase family
Tissue Specificity
Expressed in many tissues with highest expression in spleen and testis, and lowest expression in small intestine. Expressed in higher amount in pancreas, heart and kidney and at lower levels in brain, lung and liver.
Database Links

HGNC: 9154

OMIM: 605610

KEGG: hsa:11284

STRING: 9606.ENSP00000323511

UniGene: Hs.78016

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