Restriction endonucleases

Jim Owens jow at helix.nih.gov
Wed Sep 14 17:24:52 EST 1994


I wish I had seen the start of this thread so I would know whether to
muddy the waters or not.  

Oh, well.  What the hell.

In article <1994Sep7.093349.1 at leif> , vdrover at kean.ucs.mun.ca writes:
>	Restriction endonucleases (RE) are, as stated earlier, protein
>molecules (enzymes) that cleave DNA at site specific locations. Most are
>isolated from bacteria and are thus named. For instance, EcoRI was
isolated 
>from E.coli. If i remember correctly, they help guard a cell from
invasion by 
>foreign DNA, as in the case of a viral attack. Although most restriction 
>enzymes recognize 6 nucleotides, some recognize 4 and 8 nucleotides, the 
>former obviously having a higher probability of finding the proper
sequence
>. A couple of details about RE's:

Here are some more details:

What Vic Drover describes in his post are Type II restriction
endonucleases.  There are also Type I and Type III which do not cut at
specific sites.  All three types protect the bacterial cell from foreign
DNA as part of a two part bacterial "immune" system.  The other part is
the modification step.  Each endonuclease has a corresponding
modification enzyme.  The modification step makes the host DNA
recognizable as self.  This involves methylation (in the Types I, II and
III restriction systems) of bases in the sequence recognized by the
endonuclease.  

Methylation is not the only modification that is made to host DNA.  Back
when I was just a babe in school (1960), there were reports of
glycosylation of bacteriophage DNA in Scientific American.  This seemed
be yet another restriction-modification system, but I've never seen any
more about this.

Type I enzymes recognize a particular sequence string but cut the DNA
some distance away from this site.  It is not a fixed distance.  There
are some other differences, like Type I systems have single, 3-subunit
proteins with both methylase and nuclease activities; the third subunit
is for sequence recognition.  They require ATP hydrolysis,
S-adenosylmethionine and Mg2+ for activity.  Type III enzymes have both
activities on the same protein, like Type I, but the cleavage is near to
the recognition sequence and is predictable, like Type II.  Although they
require ATP for activity, there is little hydrolysis of ATP.

The first descriptions of restriction-modification systems were published
in the very early 1950s.  The initial observation was that transfections
of one strain of E. coli were much more efficient than in another.  The
genetics of this phenomenon was worked out before 1955 and seems to match
that of Type I systems.  If anyone is interested, I can search my
archives for the relevant details of who, what, where and when.

Good luck,

Jim Owens



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