| Summary: | 碩士 === 國立中興大學 === 生命科學系所 === 99 === Polyamines play an essential role in cell growth. Polyamines that are fully protonated under physiological conditions can interact with nucleic acids and stabilize DNA structure. Actinomycin D (ACTD) is an anticancer antibiotic which binds to double-stranded DNA preferably at the GpC sequence via intercalation of the planar chromophore. This drug has been used clinically for the treatment of highly malignant tumors, such as Wilms’ tumor and gestational trophoblastic neoplasia. Our research is to clarify the influence of polyamines on the ACTD acting on DNA duplex. The binding constants between ACTD and DNA determined by surface plasmon resonance showed spermine is able to decrease the DNA-binding activity of ACTD. According to UV-vis absorption spectra, spermine results in hyperchromic effect on the ACTD-DNA complexes. Circular dichroism (CD) studies revealed that ACTD–DNA complexes showed a red shift from 275 to 288 nm, indicative of a conformational transition to an A‐form structure. In the presence of spermine, DNA conformation does not change upon ACTD binding and spermine can stabilize ACTD-DNA complexes, according to melting temperature studies. Klenow fragment and T7 RNA polymerase assays showed that spermine is able to affect the inhibition of ACTD on the DNA and RNA synthesis. BrdU assay and cytotoxicity experiments indicated depletion of intracellular polyamines by methylglyoxal bis(guanylhydrazone) (MGBG) enhanced the cytotoxicity of ACTD and increased the inhibition of ACTD on the DNA synthesis of the cell. In summary, our results are important in understanding the influence of polyamines on the action of ACTD to their DNA targets.
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