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2 Type II Restriction Endonucleases

Richard J. Roberts, Stephen E. Halford


The type II restriction endonucleases, commonly referred to as restriction enzymes, are among the most prolific and best known of all the nucleases. They are defined as double-strand nucleases that recognize specific DNA sequences and cleave at a defined point within or close to that sequence. They require Mg++ as a cofactor. They are the workhorses of the genetic engineer, and acronyms such as EcoRI and BamHI have become essential vocabulary for the molecular biologist. Sadly, those same molecular biologists often know much less about the basic biology, biochemistry, and molecular biology of this fascinating class of endonucleases, which is the subject of this chapter. Modrich and Roberts (1982) have summarized progress in this field up to 1982. Recently, more specialized reviews of restriction-modification systems have focused on the genetics (Wilson and Murray 1991), biochemistry (Bennett and Halford 1989), biology (Heitman 1993), and structural aspects (Anderson 1993).

The first type II restriction endonuclease was called endonuclease R (Kelly and Smith 1970; Smith and Wilcox 1970) and is now known to have consisted of a mixture of HindII and HindIII. For this discovery, Hamilton Smith of Johns Hopkins University received the Nobel prize in medicine in 1977. A cosharer in that prize, Werner Arber, had previously postulated the existence of restriction endonucleases (Arber and Dussoix 1962; Dussoix and Arber 1962) as a means of explaining earlier genetic results (Luria and Human 1952; Bertani and Weigle 1953). Soon after Smith’s discovery, a number of other restriction endonucleases were...

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