Difference between revisions of "Part:BBa K1470001"
 
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<partinfo>BBa_K1470001 short</partinfo>
 
<partinfo>BBa_K1470001 short</partinfo>
  
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Puromycin is an antibodic derived from Streptomyces alboniger. The substance affects the cell both in protein translation and mitochondrial protein import. Due to this, it's toxic to many organisms such as bacteria, Trypanosoma and humans.
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Puromycin is an antibodic derived from the bacteria <i>Streptomyces alboniger</i>. The substance affects the cell both in protein translation and mitochondrial protein import [1]. Due to this, it's toxic to many organisms such as bacteria, <i>Trypanosoma</i> and humans.
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<br>
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A small part of its structure resembles to aminoacetylated tRNA and is recognized by ribosomes. the nascent chain containing puromycin leads to a translational stop.
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A small part of its structure resembles to aminoacetylated tRNA and is recognized by ribosomes. The nascent chain containing puromycin leads to a translational stop.
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<br>
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Streptomyces alboniger protects itself from puromycin with a acetyl transferase. It uses acetyl-CoA to bind the the acetyl group with the amino group. In this state puromycin cannot be recognized by the ribosom anymore.
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<i>Streptomyces alboniger</i> protects itself from puromycin with an acetyl transferase. It uses acetyl-CoA to bind the acetyl moiety to the amino group. In this state puromycin cannot be recognized by the ribosom anymore [2].
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<br>
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This enzyme is often used for selecting stable eucaryotic cell lines. Because of the blocked protein biosynthesis nearly over 99 % of unimfected cells will be dead within tows days.
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This enzyme is often used for selecting stable eucaryotic cell lines. Because of the blocked protein biosynthesis more than 99 % of uninfected cells will be dead within two days.
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We transduced HEK-293T cells via viral vectors including puromycin N-acetyl-transferase as a selection marker. After incubating the cells in medium with 0,5 µg /mL up to 2 µg /mL Puromycin for two days, transduced cells were vital. Hence untransduced cells, which served as a control died within two days.
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<br>
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==References==
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<small>
  
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[1] NEUBERT, D. AND LEHNINGER, A. L.: The effect of oligomycin, gramicidin, and other antibiotics on reversal of
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mitochondrial swelling by adenosine triphosphate. Biochim. biophys. Acta 62: 556-565, 1962.<br>
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[2] DARKEN MA.: Puromycin inhibition of protein synthesis. Pharmacol Rev. 16:223-43,Sep 1964.<br>
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</small>
 
<!-- Add more about the biology of this part here
 
<!-- Add more about the biology of this part here
 
===Usage and Biology===
 
===Usage and Biology===

Latest revision as of 18:44, 2 November 2014

puromycin N-acetyl-transferase

Puromycin is an antibodic derived from the bacteria Streptomyces alboniger. The substance affects the cell both in protein translation and mitochondrial protein import [1]. Due to this, it's toxic to many organisms such as bacteria, Trypanosoma and humans.
A small part of its structure resembles to aminoacetylated tRNA and is recognized by ribosomes. The nascent chain containing puromycin leads to a translational stop.
Streptomyces alboniger protects itself from puromycin with an acetyl transferase. It uses acetyl-CoA to bind the acetyl moiety to the amino group. In this state puromycin cannot be recognized by the ribosom anymore [2].
This enzyme is often used for selecting stable eucaryotic cell lines. Because of the blocked protein biosynthesis more than 99 % of uninfected cells will be dead within two days. We transduced HEK-293T cells via viral vectors including puromycin N-acetyl-transferase as a selection marker. After incubating the cells in medium with 0,5 µg /mL up to 2 µg /mL Puromycin for two days, transduced cells were vital. Hence untransduced cells, which served as a control died within two days.

References

[1] NEUBERT, D. AND LEHNINGER, A. L.: The effect of oligomycin, gramicidin, and other antibiotics on reversal of mitochondrial swelling by adenosine triphosphate. Biochim. biophys. Acta 62: 556-565, 1962.
[2] DARKEN MA.: Puromycin inhibition of protein synthesis. Pharmacol Rev. 16:223-43,Sep 1964.
 Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 481