E. coli restriction-modification system

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Notes of relevance to me taken from [[Escherichia coli & Salmonella]]
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Brief notes of relevance to me taken from [[Escherichia coli & Salmonella]]
*Restriction systems are only found in unicellular organisms.
*Restriction systems are only found in unicellular organisms.
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*'''Classic R-M systems'''
*'''Classic R-M systems'''
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A classi R-M system includes an endonuclease that cleaves a specific DNA sequence and a DNA methyltransferase that methylates either adenosyl or cytosyl residues within the same DNA sequence.
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A classic R-M system includes an endonuclease that cleaves a specific DNA sequence and a DNA methyltransferase that methylates either adenosyl or cytosyl residues within the same DNA sequence.
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**Type 1 R-M systems
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*'''Type I R-M systems'''
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***Relatively rare.
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**Relatively rare.
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***One, three-subunit protein acts as both endonuclease and methylase.
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**One, three-subunit protein acts as both endonuclease and methylase.
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***Requires AdoMet as a cofactor.
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**Requires AdoMet as a cofactor.
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***
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**Endonuclease activity also requires Mg<sup>2+</sup> and ATP.
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**E. coli has a type 1 restriction system encoded by ''hsdR'', ''hsdM'', and ''hsdS''.
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*'''Type II R-M systems'''
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**Relatively abundant - these are the restriction enzymes found in the NEB catalog etc.
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**Separate nuclease and methylation enzymes.
 +
**Requires AdoMet as a cofactor.
 +
**Endonuclease activity also requires Mg<sup>2+</sup>
 +
**Little homology between restriction and methylation enzymes despite the shared DNA recognition sequence.
 +
**Most recognition sequences are close to symmetric.
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**Endonucleases normally act as homodimers, acting on sense and anti-sense strands.
 +
**A subgroup of Type II restriction enzymes (Type IIS) cleave a fixed distance away from the recognition sequence.
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*'''Type III R-M systems'''
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**Relatively rare.
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**Hetero-oligomeric protein catalyzes both the restriction and modification reactions.
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**Requires AdoMet as a cofactor for modification.
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**Endonuclease activity also requires Mg<sup>2+</sup> and ATP.
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**Encoded by ''mod'' and ''res'' genes in ''E. coli''.
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**Only one strand is methylated unlike the other systems.
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**The restriction enzyme requires two recognition sites in inverse orientation.
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*'''Regulation'''
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**There is no evidence of transcriptional regulation of Type I R-M systems in ''E. coli''
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**However, there must be some regulation as the modification subunit is always expressed before the restriction subunit.
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**There is little clear information about the regulation of Type II R-M systems, in part because many of them come from poorly understood bacteria.
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**Type II R-M regulation is also not very well understood.  However, the modification enzyme is again expressed prior to the restriction enzyme.

Revision as of 15:31, 25 July 2005

Brief notes of relevance to me taken from Escherichia coli & Salmonella

  • Restriction systems are only found in unicellular organisms.
    • Either the cell modifies certain sequences so they are not restricted (classic R-M systems)
    • or certain foreign modifications are restricted.
  • Mcr Systems
    • McrBC - cleaves hm5C, m5C and m4C (hydroxymethylated or methylated cytosine at positions 4 or 5). The modified bases must be paired about 40-80bp apart. (genes mcrB and mcrC)
    • McrA - cleaves DNA mehylated by HpaII and SssI methylases. (encoded by the prophagelike e14 element)
  • Classic R-M systems

A classic R-M system includes an endonuclease that cleaves a specific DNA sequence and a DNA methyltransferase that methylates either adenosyl or cytosyl residues within the same DNA sequence.

  • Type I R-M systems
    • Relatively rare.
    • One, three-subunit protein acts as both endonuclease and methylase.
    • Requires AdoMet as a cofactor.
    • Endonuclease activity also requires Mg2+ and ATP.
    • E. coli has a type 1 restriction system encoded by hsdR, hsdM, and hsdS.
  • Type II R-M systems
    • Relatively abundant - these are the restriction enzymes found in the NEB catalog etc.
    • Separate nuclease and methylation enzymes.
    • Requires AdoMet as a cofactor.
    • Endonuclease activity also requires Mg2+
    • Little homology between restriction and methylation enzymes despite the shared DNA recognition sequence.
    • Most recognition sequences are close to symmetric.
    • Endonucleases normally act as homodimers, acting on sense and anti-sense strands.
    • A subgroup of Type II restriction enzymes (Type IIS) cleave a fixed distance away from the recognition sequence.
  • Type III R-M systems
    • Relatively rare.
    • Hetero-oligomeric protein catalyzes both the restriction and modification reactions.
    • Requires AdoMet as a cofactor for modification.
    • Endonuclease activity also requires Mg2+ and ATP.
    • Encoded by mod and res genes in E. coli.
    • Only one strand is methylated unlike the other systems.
    • The restriction enzyme requires two recognition sites in inverse orientation.
  • Regulation
    • There is no evidence of transcriptional regulation of Type I R-M systems in E. coli
    • However, there must be some regulation as the modification subunit is always expressed before the restriction subunit.
    • There is little clear information about the regulation of Type II R-M systems, in part because many of them come from poorly understood bacteria.
    • Type II R-M regulation is also not very well understood. However, the modification enzyme is again expressed prior to the restriction enzyme.
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