Cutting and joining DNA molecules - Principles of Gene Manipulation - page 29

products of the hsd R, hsd M and hsd S genes. The
methylation and cutting reactions both require ATP
and S -adenosylmethionine as cofactors. The recog-
nition sequences are quite long with no recognizable
features such as symmetry. The enzyme also cuts
unmodified DNA at some distance from the recogni-
tion sequence. However, because the methylation
reaction is performed by the same enzyme which
mediates cleavage, the target DNA may be modified
before it is cut. These features mean that type I sys-
tems are of little value for gene manipulation (see
also Box 3.1). However, their presence in E. coli
strains can affect recovery of recombinants (see
p. 33). Type III enzymes have symmetrical recog-
nition sequences but otherwise resemble type I
systems and are of little value.
Most of the useful R-M systems are of type II. They
have a number of advantages over type I and III
systems. First, restriction and modification are medi-
ated by separate enzymes so it is possible to cleave
DNA in the absence of modification. Secondly, the
restriction activities do not require cofactors such as
ATP or S -adenosylmethionine, making them easier
to use. Most important of all, type II enzymes recog-
nize a defined, usually symmetrical, sequence and
cut within it . Many of them also make a staggered
break in the DNA and the usefulness of this will
become apparent. Although type IIs systems have
similar cofactors and macromolecular structure to
those of type II systems, the fact that restriction
occurs at a distance from the recognition site limits
their usefulness.
The classification of R-M systems into types I to III
is convenient but may require modification as new
discoveries are made. For example, the Eco 571 sys-
tem comprises a single polypeptide which has both

restriction and modification activities (Petrusyte
et al. 1988). Other restriction systems are known
which fall outside the above classification. Examples
include the mcr and mrr systems (see p. 34) and
homing endonucleases. The latter are double-
stranded DNases derived from introns or inteins
(Belfort & Roberts 1997). They have large, asym-
metric recognition sequences and, unlike standard
restriction endonucleases, tolerate some sequence
degeneracy within their recognition sequence.

Nomenclature

The discovery of a large number of restriction and
modification systems called for a uniform system of
nomenclature. A suitable system was proposed by
Smith and Nathans (1973) and a simplified version
of this is in use today. The key features are:
• The species name of the host organism is identi-
fied by the first letter of the genus name and the first
two letters of the specific epithet to generate a three-
letter abbreviation. This abbreviation is always
written in italics.
• Where a particular strain has been the source
then this is identified.
• When a particular host strain has several different
R-M systems, these are identified by roman numerals.
Some examples are given in Table 3.2.
Homing endonucleases are named in a similar
fashion except that intron-encoded endonucleases
are given the prefix 'I-' (e.g. I- Ceu I) and intein
endonucleases have the prefix 'PI-' (e.g. Pl- Psp I).
Where it is necessary to distinguish between the
restriction and methylating activities, they are given
the prefixes 'R' and 'M', respectively, e.g. R. Sma I and
M. Sma I.

Table 3.2 Examples of restriction endonuclease nomenclature.

Enzyme Enzyme source Recognition sequence

Sma I Serratia marcescens ,1st enzyme CCCGGG

Hae III Haemophilus aegyptius, 3rd enzyme GGCC

Hin dII Haemophilus influenzae , strain d, 2nd enzyme GTPyPuAC

Hin dIII Haemophilus influenzae , strain d, 3rd enzyme AAGCTT

Bam HI Bacillus amyloliquefaciens , strain H, 1st enzyme GGATCC