The Effects of Anti-tumor Antibiotic CC-1065 on Restriction Enzymes Involved in Processing the DNA
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CC-1065 is a potent new anti-tumor antibiotic discovered in Upjohn Laboratories, produced by Streptomyces zelensis. This drug interacts strongly with double stranded DNA at adenine-thymine sites in the minor groove and is thought to exert its cytotoxic effects through superstabilization of the DNA helix. The manner in which it binds to DNA suggests possible mechanisms of its cytotoxicity such as stiffening of the helix by CC-1065 which prevents free association of the DNA with the enzyme of proper enzyme activity, physical masking of an enzyme restriction site, and inhibition of normal unwinding and enzymatic processes occurring during replication. In this project the effects of CC-1065 on restriction enzymes specific for known sequences on 𝜙𝜲174RF phage DNA were investigated. The 𝜙𝜲174RF DNA was treated with various concentrations of CC-1065 so as to yield CC-1065 molecule to nucleotide ratios of 1:50, 1:200, and 1:800. After this the samples were treated with enzyme. A control was also prepared containing ,𝜙𝜲174RF DNA and enzyme ( no drug ), another was prepared containing the CC-1065 and ,𝜙𝜲174RF DNA alone ( no enzyme,). These samples were incubated for 1,2, or 3 hours at 37º C and run through agarose of polyacrylamide gel electrophoresis. It was found that AT specific enzymes were more greatly inhibited by CC-1065 than GC specific enzymes. It was also found that CC-1065 does inhibit some cleavage sites on the 𝜙𝜲174RF DNA preferentially to others. Further experimentation is necessary to determine the length of the adenine-thymine run required by the drug, and to investigate the long-range inductive effects of the drug.