RNA 5' Pyrophosphohydrolase (RppH)

Description

The bacterial RNA 5´ Pyrophosphohydrolase (RppH) removes pyrophosphate from the 5´ end of triphosphorylated RNA to leave a 5´ monophosphate RNA (1). The RppH protein was also known as NudH/YgdP which can split Ap5A to ADP and ATP (2).

Product Source

An E. coli strain containing a clone of the E. coli RppH gene.

Reagents Supplied

The following reagents are supplied with this product:

Store at (°C)Concentration
NEBuffer 2-2010X

Properties and Usage

Unit Definition

One unit is the amount of enzyme that converts 1 µg 300 mer RNA transcript into a XRN-1 digestible RNA in 30 minutes at 37°C.

Reaction Conditions

1X NEBuffer 2
Incubate at 37°C

1X NEBuffer 2:
50 mM NaCl
10 mM Tris-HCl
10 mM MgCl2
1 mM DTT
pH 7.9 @ 25°C

Storage Temperature

-20°C

Storage Conditions

20 mM Tris-HCl
200 mM NaCl
1 mM DTT
0.1 mM EDTA
50% Glycerol
0.01% Triton® X-100
pH 7.5 @ 25°C

Notes

  1. Since RppH is active in the presence of Mg2+, it can be incubated with RNA Ligase or XRN-1.

References

  1. Deana, A. et al. (2008). Nature. 451, 355-358.
  2. Bessman, M.J. et al. (2001). JBC. 276, 37834.
  3. Neri F, Rapelli S, Krepelova A, Incarnato D, Parlato C, Basile G, et al. (2017). Intragenic DNA Methylation prevents spurious transcription initiation. Nature. 543, 72–7.
  4. Hetzel, Jonathan et al. (2016). Nascent RNA Sequencing Reveals Distinct Features in Plant Transcription. Proceeding of the National Academy of Sciences. 113.43, 12316–12321.

FAQs

  1. Can RppH replace Tobacco Acid Pyrophosphatase (TAP) as a decapping enzyme?

Tech Tips

We strongly recommend wearing gloves, using nuclease-free tubes and reagents, and thoroughly cleaning pipettes and bench surfaces to avoid RNase contamination.

Protocols

  1. Decapping eukaryotic mRNA with RppH (NEB #M0356)

Datacards

The Product Summary Sheet, or Data Card, includes details for how to use the product, as well as details of its formulation and quality controls. The following file naming structure is used to name the majority of these document files: [Catalog Number]Datasheet-Lot[Lot Number]. For those product lots not listed below, please contact NEB at info@neb.com or fill out the Technical Support Form for appropriate document.

Interactive Tools

Quality Control

Quality Control Assays

The following Quality Control Tests are performed on each new lot and meet the specifications designated for the product. Individual lot data can be found on the Product Summary Sheet/Datacard or Manual which can be found in the Supporting Documents section of this page. Further information regarding NEB product quality can be found here.
  • Endonuclease Activity (Nicking):
    The product is tested in a reaction containing a supercoiled DNA substrate. After incubation for 4 hours the percent converted to the nicked form is determined by agarose gel electrophoresis.
  • Exonuclease Activity (Radioactivity Release):
    The product is tested in a reaction containing a radiolabeled mixture of single and double-stranded DNA. After incubation for 4 hours the exonuclease activity is determined by the % release of radioactive nucleotides.
  • Phosphatase Activity (PNPP):
    The product is tested in a reaction containing a p-nitrophenyl phosphate (PNPP), a chromogenic substrate for most phosphatases. After incubation the percent degradation is determined by spectrophotometric analysis of released p-nitrophenol at 405 nm.
  • RNase Activity (2 Hour Digestion):
    The product is tested in a reaction containing a RNA substrate.  After incubation for 2 hour there is no detectable degradation of the RNA substrate as determined by gel electrophoresis.

Safety Data Sheet

The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.

Datacards

The Product Summary Sheet, or Data Card, includes details for how to use the product, as well as details of its formulation and quality controls. The following file naming structure is used to name the majority of these document files: [Catalog Number]Datasheet-Lot[Lot Number]. For those product lots not listed below, please contact NEB at info@neb.com or fill out the Technical Support Form for appropriate document.