Recombinant Mononucleosomes (H2A.Z) - biotinylated

Catalog No: 31583 Format: 20 µg $435 Buy
Expressed In: E. coli Protein Species: Human

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Contents

Recombinant Mononucleosomes (H2A.Z) - biotinylated (20 µg protein + 20 µg DNA) are supplied in 10 mM Tris-HCl, pH 8.0, 1 mM EDTA, 2 mM DTT and 20% glycerol. Please refer to product insert upon arrival for lot-specific concentration.

Background

In vivo, the nucleosome is the basic structural unit of chromatin. It is comprised of about 146 bp of DNA wrapped around a core of eight histones of four different types: H2A, H2B, H3 and H4. Histones are subject to post-translational modifications, such as methylation, acetylation, phosphorylation, mono-ubiquitination, etc. Histone modifications influence multiple chromatin templated processes such as gene transcription, DNA repair and recombination. Besides the “major“ histones, there are some histone variants in specific regions of chromatin or in specific cell types. Histone variants are involved in multiple biological processes including chromosome segregation, DNA repair, transcriptional regulation and mRNA processing.

H2A.Z (also known as H2AFZ, Histone Family Member Z) is a histone H2A family member. It is highly conserved from yeast to human, with 90% of its primary sequence preserved among different species, showing only 60% homology with canonical histone H2A. H2A.Z is found in approximately 10% of mammalian nucleosomes. H2A.Z has been one of the most studied H2A variants in recent years. It plays an important role in different biological processes such as DNA replication, DNA repair, transcription regulation, cell cycle, spermatogenesis, chromosome segregation, centromere structure and maintenance of heterochromatic/euchromatic status. However, different studies reported diverse conclusions in the nucleosome stability and transcriptional regulation. The contradictory roles of H2A.Z in vivo might be explained by different combinations of H2A.Z with other epigenetic regulators, PTMs on H2A.Z and interaction with chromatin binding proteins.

Nucleosomes are more physiologically relevant substrates than histones and histone-derived peptides for in vitro studies. More importantly, some histone methyltransferases are significantly more active, as well as specific, when using nucleosomal substrates in HMT assays, such as DOT1L and NSD family enzymes. Nucleosomes are also widely used in histone methyltransferase screening assays to identify small molecular inhibitors for drug discovery.

Application Notes

Recombinant Mononucleosomes (H2A.Z) - biotinylated are suitable for use as substrates in the study of enzyme kinetics, inhibitor screening, and selectivity profiling.

Protein Details

Recombinant Mononucleosomes (H2A.Z) - biotinylated, consist of 167 bp of 601 DNA with a 5’ biotin tag and two molecules each of histones H2A.Z that includes amino acids 1-128 (end) (accession number NP_002097.1), H2B that includes amino acids 1-126 (end) (accession number NP_003509.1), H3 that includes amino acids 1-136 (end) (accession number NP_003520.1), and H4 that includes amino acids 1-103 (end) (accession number NP_003539.1). All of these histones were expressed in E. coli cells. The molecular weight of the histone octamer is ~108.3 kDa.

 
Recombinant Mononucleosomes (H2A.Z) - biotinylated, SDS-PAGE.

Recombinant Mononucleosomes (H2A.Z) - biotinylated, protein gel.
12.5% SDS-PAGE gel stained with Coomassie Blue
Purity: ≥ 95%

Streptavidin pull-down for Recombinant Mononucleosomes (H2A.Z) - biotinylated.

Streptavidin pull-down for Recombinant Mononucleosomes (H2A.Z) - biotinylated.
Mononucleosomes (H2A.Z) were pulled down by streptavidin beads. Input mononucleosomes (Lane 1) and the mononucleosomes pulled down by streptavidin (Lane 2) were run on a 12.5% SDS-PAGE gel and stained with Coomassie Blue. The SDS-PAGE gel result shows that almost all of biotinylated mononucleosomes (H2A.Z) are pulled down by streptavidin beads.
* indicates streptavidin. Nucleosomes = Mononucleosomes.

Recombinant Mononucleosomes (H2A.Z) - biotinylated, DNA gel.

Recombinant Mononucleosomes (H2A.Z) - biotinylated, DNA gel.
Biotinylated mononucleosomes (H2A.Z) were run on a 2% agarose gel and stained with ethidium bromide. Lane 1: DNA marker. Lane 2: free 601 DNA. Lane 3: Intact mononucleosomes. Intact mononucleosomes migrate much higher than free 601 DNA.
The agarose gel result shows that almost all of 601 DNA wraps histone octamers to form mononucleosomes. Nucleosomes = Mononucleosomes.

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.