S. cerevisiae RNA polymerase II CTD repeat YSPTSPS (phospho S2) peptide (ab12793)

Overview

Description

  • NatureSynthetic
  • Amino Acid Sequence
    • AccessionP04050
    • SpeciesSaccharomyces cerevisiae
    • Modificationsphospho S2

Specifications

Our Abpromise guarantee covers the use of ab12793 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

  • Applications

    Western blot

    Blocking - Blocking peptide for Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S2) antibody - ChIP Grade (ab5095)

  • FormLiquid
  • Additional notes

    - First try to dissolve a small amount of peptide in either water or buffer. The more charged residues on a peptide, the more soluble it is in aqueous solutions.
    - If the peptide doesn’t dissolve try an organic solvent e.g. DMSO, then dilute using water or buffer.
    - Consider that any solvent used must be compatible with your assay. If a peptide does not dissolve and you need to recover it, lyophilise to remove the solvent.
    - Gentle warming and sonication can effectively aid peptide solubilisation. If the solution is cloudy or has gelled the peptide may be in suspension rather than solubilised.
    - Peptides containing cysteine are easily oxidised, so should be prepared in solution just prior to use.

  • Concentration information loading...

Preparation and Storage

  • Stability and Storage

    Shipped at 4°C. Upon delivery aliquot and store at -20°C or -80°C. Avoid repeated freeze / thaw cycles.

    Information available upon request.

General Info

  • Alternative names
    • DNA directed RNA polymerase II A
    • DNA-directed RNA polymerase II largest subunit RNA polymerase II 220 kd subunit
    • DNA-directed RNA polymerase II subunit A
    • DNA-directed RNA polymerase II subunit RPB1
    • DNA-directed RNA polymerase III largest subunit
    • hRPB220
    • hsRPB1
    • POLR2
    • Polr2a
    • POLRA
    • Polymerase (RNA) II (DNA directed) polypeptide A
    • Polymerase (RNA) II (DNA directed) polypeptide A 220kDa
    • RNA polymerase II subunit B1
    • RNA-directed RNA polymerase II subunit RPB1
    • RPB1
    • RPB1_HUMAN
    • RPBh1
    • RpIILS
    • RPO2
    • RPOL2
    see all
  • FunctionDNA-dependent RNA polymerase catalyzes the transcription of DNA into RNA using the four ribonucleoside triphosphates as substrates. Largest and catalytic component of RNA polymerase II which synthesizes mRNA precursors and many functional non-coding RNAs. Forms the polymerase active center together with the second largest subunit. Pol II is the central component of the basal RNA polymerase II transcription machinery. It is composed of mobile elements that move relative to each other. RPB1 is part of the core element with the central large cleft, the clamp element that moves to open and close the cleft and the jaws that are thought to grab the incoming DNA template. At the start of transcription, a single-stranded DNA template strand of the promoter is positioned within the central active site cleft of Pol II. A bridging helix emanates from RPB1 and crosses the cleft near the catalytic site and is thought to promote translocation of Pol II by acting as a ratchet that moves the RNA-DNA hybrid through the active site by switching from straight to bent conformations at each step of nucleotide addition. During transcription elongation, Pol II moves on the template as the transcript elongates. Elongation is influenced by the phosphorylation status of the C-terminal domain (CTD) of Pol II largest subunit (RPB1), which serves as a platform for assembly of factors that regulate transcription initiation, elongation, termination and mRNA processing. Acts as an RNA-dependent RNA polymerase when associated with small delta antigen of Hepatitis delta virus, acting both as a replicate and transcriptase for the viral RNA circular genome.
  • Sequence similaritiesBelongs to the RNA polymerase beta' chain family.
  • DomainThe C-terminal domain (CTD) serves as a platform for assembly of factors that regulate transcription initiation, elongation, termination and mRNA processing.
  • Post-translational
    modifications
    The tandem heptapeptide repeats in the C-terminal domain (CTD) can be highly phosphorylated. The phosphorylation activates Pol II. Phosphorylation occurs mainly at residues 'Ser-2' and 'Ser-5' of the heptapeptide repeat and is mediated, at least, by CDK7 and CDK9. CDK7 phosphorylation of POLR2A associated with DNA promotes transcription initiation by triggering dissociation from DNA. Phosphorylation also takes place at 'Ser-7' of the heptapeptide repeat, which is required for efficient transcription of snRNA genes and processing of the transcripts. The phosphorylation state is believed to result from the balanced action of site-specific CTD kinases and phosphatases, and a 'CTD code' that specifies the position of Pol II within the transcription cycle has been proposed. Dephosphorylated by the protein phosphatase CTDSP1.
    Among tandem heptapeptide repeats of the C-terminal domain (CTD) some do not match the Y-S-P-T-S-P-S consensus, the seventh serine residue 'Ser-7' being replaced by a lysine. 'Lys-7' in these non-consensus heptapeptide repeats can be alternatively acetylated, methylated and dimethylated. EP300 is one of the enzyme able to acetylate 'Lys-7'. Acetylation at 'Lys-7' of non-consensus heptapeptide repeats is associated with 'Ser-2' phosphorylation and active transcription. It may regulate initiation or early elongation steps of transcription specially for inducible genes.
    Methylated at Arg-1810 prior to transcription initiation when the CTD is hypophosphorylated, phosphorylation at Ser-1805 and Ser-1808 preventing this methylation. Symmetrically or asymmetrically dimethylated at Arg-1810 by PRMT5 and CARM1 respectively. Symmetric or asymmetric dimethylation modulates interactions with CTD-binding proteins like SMN1/SMN2 and TDRD3. SMN1/SMN2 interacts preferentially with the symmetrically dimethylated form while TDRD3 interacts with the asymmetric form. Through the recruitment of SMN1/SMN2, symmetric dimethylation is required for resolving RNA-DNA hybrids created by RNA polymerase II, that form R-loop in transcription terminal regions, an important step in proper transcription termination. CTD dimethylation may also facilitate the expression of select RNAs. Among tandem heptapeptide repeats of the C-terminal domain (CTD) some do not match the Y-S-P-T-S-P-S consensus, the seventh serine residue 'Ser-7' being replaced by a lysine. 'Lys-7' in these non-consensus heptapeptide repeats can be alternatively acetylated, methylated and dimethylated. Methylation occurs in the earliest transcription stages and precedes or is concomitant to 'Ser-5' and 'Ser-7' phosphorylation.
    Ubiquitinated by WWP2 leading to proteasomal degradation (By similarity). Following UV treatment, the elongating form of RNA polymerase II (RNA pol IIo) is ubiquitinated UV damage sites without leading to degradation: ubiquitination is facilitated by KIAA1530/UVSSA and promotes RNA pol IIo backtracking to allow access to the nucleotide excision repair machinery.
  • Cellular localizationNucleus.
  • Information by UniProt

S. cerevisiae RNA polymerase II CTD repeat YSPTSPS (phospho S2) peptide images

  • ab12793 RNA polymerase II CTD repeat YSPTSPS peptide - phospho S2 0.1 ug/ml was mixed with GST tagged recombinant proteins in vitro in binding buffer for 1 hour at room temp. Glutathione coated magnetic beads were then mixed into the buffer and removed after another hour incubation. The GST tagged recombinant proteins and any bound peptide was eluted by incubating with free glutathione. The peptides analyzed by dotblot using primary antibody ab5095(1/3000) and an undiluted conjugated HRP-Goat anti-rabbit secondary antibody.
    This experiment was conducted with ab12795, ab18488 and an another peptide as a negative control.

    See Abreview

References for S. cerevisiae RNA polymerase II CTD repeat YSPTSPS (phospho S2) peptide (ab12793)

This product has been referenced in:
  • Freter R  et al. Adult stem cells exhibit global suppression of RNA polymerase II serine-2 phosphorylation. Stem Cells 28:1571-80 (2010). Blocking . Read more (PubMed: 20641035) »
  • Racanelli AC  et al. A mouse gene that coordinates epigenetic controls and transcriptional interference to achieve tissue-specific expression. Mol Cell Biol 28:836-48 (2008). Blocking . Read more (PubMed: 17998333) »

See all 2 Publications for this product

Product Wall

Application Other

Review text: This peptide was mixed with GST tagged recombinant proteins in vitro in binding buffer for 1 hour at room temp. Glutathione coated magnetic beads were then mixed into the buffer and removed after another hour incubation. The GST tagged recombinant proteins and any bound peptide was eluted by incubating with free glutathione. Eluted proteins were analyzed by western blot and peptides analyzed by dotblot.

This experiment was conducted with ab12795, ab18488 and an unrelated non-Abcam peptide as a negative control.
Sample: Human Recombinant protein

Primary antibody (in addition to 'RNA polymerase II CTD repeat YSPTSPS peptide - phospho S2')
Primary antibody: Abcam primary antibody: Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S2) antibody - ChIP Grade (ab5095)
Dilution: 1/3000

Secondary antibody
Name: Non-Abcam antibody was used: anti-rabbit
Host species: Goat
Clonality: Polyclonal
Conjugation: Horse Radish Peroxidase

Additional data
Notes: Ab5095 was used to detect peptides in this experiment.
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Submitted Aug 13 2012

Application Other

Review text: This peptide was used in an in vitro pulldown assay. It was biotinylated and linked to streptavidin coated magnetic beads. This complex was then mixed with our proteins of interest under a variety of conditions. Proteins which bound to the CTD-S2p peptide were eluted and assessed by western blot.

This peptide was used along side ab18488, ab12795, and an unrelated non-abcam peptide and binding to each CTD was compared.
Sample: Human Recombinant protein

Primary antibody (in addition to 'RNA polymerase II CTD repeat YSPTSPS peptide - phospho S2')
Primary antibody: None used

Secondary antibody
Secondary antibody: None used
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Submitted Aug 07 2012

Application Western blot

Review text: This peptide was used as a positive control for ab5095 (anti S2p-CTD)
Sample: Human Recombinant protein (ab12793 peptide)
Loading amount: 0.25 µg
Specification: ab12793 peptide
Gel Running Conditions: Reduced Denaturing (16% tris-tricine (Schagger et al 2006))
Blocking step: BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 23°C

Other product details
Incubation time: 1 hour(s) and 0 minute(s) · Temperature: 23°C

Primary antibody (in addition to 'RNA polymerase II CTD repeat YSPTSPS peptide - phospho S2')
Primary antibody: Abcam primary antibody: Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S2) antibody - ChIP Grade (ab5095)
Dilution: 1/3000

Secondary antibody
Name: Non-Abcam antibody was used: anti-rabbit
Host species: Goat
Clonality: Polyclonal
Conjugation: Horse Radish Peroxidase
Dilution: 1/20000

Detection
Detection method: west pico
Exposure: 30 second(s)
Bands: Specific: 4 kDa

Additional data
Additional Notes: Band was detected at 4kDa but should appear at 1.7kDa. Probably due to uncertain running of a very low MW product. Only 1 band should be present in the sample and only one band was detected.
Abcam response: Thank you for submitting your Abreview. While you have been able to detect the 1.4 kDa band, we would not typically recommend this peptide for use as a WB positive control because it is very difficult to successfully run and transfer peptides of this size. The peptide is commonly used for blocking purposes.
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Submitted Aug 01 2012

Application Other

Review text: This peptide was used for dot blots to quality control anti-CTD antibodies. It was detected by ab5095 but not with anti-S5-CTD antibody.
Sample: Human Recombinant protein

Primary antibody (in addition to 'RNA polymerase II CTD repeat YSPTSPS peptide - phospho S2')
Primary antibody: Abcam primary antibody: Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S2) antibody - ChIP Grade (ab5095)
Dilution: 1/3000

Secondary antibody
Name: Non-Abcam antibody was used: anti-rabbit
Host species: Goat
Clonality: Polyclonal
Conjugation: Horse Radish Peroxidase
Dilution: 1/20000

Additional data
Notes: 0.1ug of peptide was spotted onto PVDF and membrane blocked with 2% BSA prior to antibody incubation.
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Submitted May 25 2012

Thank you for your enquiry. ab18488 was synthesised as >90 purity by HPLC analysis, although by weight they will contain salts and buffer components. They are supplied as a solution in the buffer stated on the datasheet. ab18488 was synthesised as ...

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