Recombinant NgTet1 (1-321) protein

Catalog No: 81148 Format: 100 µg $435 Buy
Catalog No: 81848 Format: 1 mg $2,700 Buy

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Expressed In: E. coli Protein Species: Naegleria

Contents

A representative Technical Data Sheet (TDS) is provided here. Please refer to the lot-specific TDS you will receive with your order for the lot-specific buffer contents and protein concentration.

Background

Ten Eleven Translocation (TET) family proteins are cytosine oxygenases that catalyze the conversion of 5-methylcytosine (5-mC) into 5-hydroxymethylcytosine (5-hmC). 5-hmC can be further oxidized into 5-formylcytosine (5-fC) and 5-carboxylcytosine (5-caC) by TET proteins. Methylation at the C5 position of cytosine is an epigenetic modification of the mammalian genome that plays an important role in transcriptional regulation. TET proteins are responsible for iinitiating the enzymatic deamination process leading to cytosine demethylation.

The Tet family of dioxygenases is widely distributed across the tree of life, including in the heterolobosean amoeboflagellate Naegleria gruberi . NgTet1 (Naegleria Tet-like protein) is a homologue of mammalian Tet proteins. Like mammalian Tet proteins, NgTet1 acts on 5-mC and generates 5-hmC, 5-fC and 5-caC.

Application Notes

Recombinant NgTet1 protein is suitable for use in cytosine oxygenase assays, 5-mC sequencing, enzyme kinetics, inhibitor screening, and selectivity profiling..

TET Activity Assay Conditions: 250 nM of 30bp DNA oligos (sequence: 5'-CAGTAACTGTGGTC(mC)GGTAACTGACTTGCA-3', contains a 5mCpG site) was incubated with 100 nM recombinant TET1 (1418-2136) protein in buffer containing 50 mM HEPES, pH 8.0, 100 μM Fe(NH4)2(SO4)2, 2 mM ascorbate, 1 mM alpha-ketoglutarate, 1 mM ATP, 1 mM TCEP at 37C for 1 hr. Activity was detected by Dot-blot.

Protein Details

Recombinant NgTet1 protein was expressed in E. coli cells as the full length protein (accession number XP_002667965.1) with an N-terminal 6xHis Tag. The molecular weight of NgTet1 is 38.7 kDa.

 

Recombinant NgTet1 protein
10% SDS-PAGE Coomassie staining
MW: 38.7 kDa
Purity: >85%

Recombinant NgTet1 activity assay
1 µg of 30 nt ssDNA oligos with a 5mCpG site were incubated with different concentrations of NgTet1 in a reaction system at 34°C for 1 hour. Sample was concentrated to 10 µl, and 1 µl was spotted onto nylon membrane. 5hmC Ab (Cat. No. 39769) and 5-caC Ab (Cat. No. 61225) were used to detect products. NgTet1 activity assay was detected by Dot-Blot..

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.