Restriction Enzymes 109tion sequence. Type III enzymes, such as HinfllI (7), are ATP and Mg 2÷
dependent, but do not show a stringent requirement for S-adenosyl-
methionine (Table 1). The classification of restriction enzymes as Type
I or Type II enzymes was proposed by Boyer (8). The currently used
and accepted nomenclature for individual restriction enzymes was
introduced by Smith and Nathans (9): It uses a three-letter abbrevia-
tion (in italics) for the organism from which the enzyme has been
isolated. Often a fourth letter is included to designate a particular
strain. A roman numeral is added to differentiate enzymes of different
specificity isolated from the same source. For example, HindlI is the
name for the second restriction enzyme isolated from H. influenzae
strain d. The official nomenclature includes a capital letter R followed
by a raised dot in front of the enzyme symbol, e.g., RoHpalI, to distin-
guish the restriction enzyme from the corresponding modification
enzyme, e.g., M°HpalI. Most often, however, the letter Ro is not used,
such that HpalI, for example, denotes the restriction endonuclease and
not the modification methyltransferase.
The Type II restriction enzymes that recognize defined DNA sequences
4 to 8 bp in length and cleave the DNA within or close to the recog-
nition sequence have turned out to be of paramount importance for the
analysis and engineering of the genetic material. They will be the
subject of this chapter. Excellent reviews have been published that
deal with the genetics (10) and enzymology of restriction enzymes
(11-14). Two volumes of the series Gene Amplification and Analysis
deal with restriction enzymes (15,16). Experimental details for the use
of restriction enzymes can be found in various laboratory manuals
(e.g., 17-21), as well as in vol. 65 of Methods in Enzymology (22).
Comprehensive lists of the properties and sources of known restriction
enzymes appear regularly (23,24).
The present chapter will focus on the structure and mechanism of
action of restriction enzymes. Information will be emphasized that is
important for the use of restriction enzymes for various purposes.
- Enzymology of Type II Restriction Enzymes
(EC 3.1.21.4)
Well over 1200 different restriction enzymes have been discovered
in a wide variety of prokaryotes, (see compilation by Roberts [24]);
among them, many are isoschizomers, i.e., enzymes that recognize the