Recombinant ALKBH3 protein

Catalog No: 81130 Format: 100 µg ¥50,000 Buy
Catalog No: 81830 Format: 1 mg ¥400,000 Buy
Expressed In: E. coli Protein Species: Human

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Contents

Recombinant ALKBH3 protein is supplied in 25 mM Tris pH 8.0, 300 mM NaCl, 10% glycerol, 0.5 mM TCEP.

Background

ALKBH3 (AlkB Homolog 3, Alpha-Ketoglutarate Dependent Dioxygenase) is a member of ALKBH protein family. It mediates demethylation of DNA and RNA containing 1-methyladenosine (m1A). It can repair alkylated DNA containing 1-methyladenosine (m1A) and 3-methylcytosine (m3C) by oxidative demethylation. ALKBH3 has a strong preference for single-stranded DNA. It is able to process alkylated m3C within double-stranded regions via its interaction with ASCC3, which promoting DNA unwinding to generate single-stranded substrate needed for ALKBH3. ALKBH3 also acts on RNA via demethylating N(1)-methyladenosine (m1A) RNA, an epigenetic internal modification of messenger RNAs (mRNAs) highly enriched within 5-untranslated regions (UTRs) and in the vicinity of start codons. The oxidative demethylation requires molecular oxygen, alpha-ketoglutarate and iron.

Application Notes

Recombinant ALKBH3 protein is suitable for use in enzyme kinetics, inhibitor screening, and selectivity profiling.

Protein Details

Full length ALKBH3 protein (accession number NP_631917.1) was expressed in E.coli cells with an N-terminal 6xHis tag. The molecular weight of ALKBH3 is 36.9 kDa.

 

Recombinant ALKBH3 protein
12.5% SDS-PAGE Coomassie staining
MW: 36.9 kDa
Purity: >90%

ALKBH3 dioxygenase activity
1 µg of ssDNA oligos (5’-AAAGCAG(1mA) ATTCGAAAAAGCGAAA-3’) was incubated with varying concentrations of ALKBH3 in a reaction system including 50 mM HEPES-NaOH pH 8.0, 50 µM Fe(NH4)2(SO4)2, 1 mM 2-oxoglutarate, 2 mM ascorbate and 1 mM TCEP for 30 min at 37°C. Then ssDNA oligos were annealed with equimolar of non-methylated complement strand followed by 1 µg EcoRI digestion for 45 min at 37°C.

% reaction products were run on a 20% Native PAGE gel and stained by ethidium bromide.

Storage

Recombinant proteins in solution are temperature sensitive and must be stored at -80°C to prevent degradation. Avoid repeated freeze/thaw cycles and keep on ice when not in storage.

Guarantee

This product is for research use only and is not for use in diagnostic procedures. This product is guaranteed for 6 months from date of arrival.