Chandrashekaran, Siddamadappa and Saravanan, Matheshwaran and Radha, Deshpande R and Nagaraja, Valakunja (2004) $Ca^2^+-$mediated Site-specific DNA Cleavage and Suppression of Promiscuous Activity of KpnI Restriction Endonuclease. In: Journal of Biological Chemistry., 279 (48). pp. 49736-49740.
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The characteristic feature of type II restriction endonucleases (REases) is their exquisite sequence specificity and obligate $Mg^2^+$ requirement for catalysis. Efficient cleavage of DNA only in the presence of $Ca^2^+$ ions, comparable with that of $Mg^2^+$, is previously not described. Most intriguingly, KpnI REase exhibits $Ca^2^+$-dependent specific DNA cleavage. Moreover, the enzyme is highly promiscuous in its cleavage pattern on plasmid DNAs in the presence of $Mn^2^+$ or $Mg^2^+$, with the complete suppression of promiscuous activity in the presence of $Ca^2^+$. KpnI methyltransferase does not exhibit promiscuous activity unlike its cognate REase. The REase binds to oligonucleotides containing canonical and mapped noncanonical sites with comparable affinities. However, the extent of cleavage is varied depending on the metal ion and the sequence. The ability of the enzyme to be promiscuous or specific may reflect an evolutionary design. Based on the results, we suggest that the enzyme KpnI represents an REase evolving to attain higher sequence specificity from an ancient nonspecific nuclease.
|Item Type:||Journal Article|
|Additional Information:||The copyright of this article belongs to American Society for Biochemistry and Molecular Biology.|
|Department/Centre:||Division of Biological Sciences > Microbiology & Cell Biology|
|Date Deposited:||24 Mar 2006|
|Last Modified:||19 Sep 2010 04:24|
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