Modifications of Circular DNA by Photoalkylation

Abstract

The effects of photoalkylation on superhelical PM2 DNA were examined. The chief product was 8-(2-hydroxy-2-propyl)guanine, formed exclusively in sequences of alternating purines and pyrimidines. Other purine damages included 8-(2-hydroxy-2-propyl)adenine and smaller quantities of two uncharacterized adenine products. DNA strand breaks were formed with increasing irradiation. A small quantity of thymine-containing photodimers was formed. Photoalkylation of poly(dG-dC):poly(dG-dC) reduced the concentration of salt required to effect inversion of the circular dichroic spectrum. This suggests that photoalkylation induces the transition of poly(dG-dC):poly(dG-dC) from the right-handed B form of DNA to the left-handed Z form.

Endonucleolytic Incision of Uvb‐irradiated Dna

“…DNA was irradiated with a Bausch & Lomb unit equipped with a 200-W Osram Hg bulb. Broadspectrum UV irradiation was conducted through a Schott WG 320 filter at an incident dose of 6.6 x einstein, cm*/min as previously described (Duker et al, 1982(Duker et al, , 1985Weiss and Duker, 1986). Monochromatic UV irradiation of PM2 phage DNA was performed at the wavelengths and doses indicated through a Bausch & Lomb highintensity monochromator as previously described (Duke1 et al, 1981).…”

Photoalkylated DNA and ultraviolet-irradiated DNA are incised at cytosines by endonuclease III

“…The DNA was purified as described (6) and stored in 0.1 M KH2PO4, pH 7.0. DNA was photoalkylated in 1.3 M isopropanol and 2X di-tert-butylperoxide for 5 minutes as previously described (2,3). Mixed-wavelength irradiation was with a Bausch and Lomb unit equipped with a 200 W Osram Hg bulb and a Schott W G 320 filter at an incident dose of 6.6 x 10-5 einstein/cm2Imin (2,3).…”

“…DNA was photoalkylated in 1.3 M isopropanol and 2X di-tert-butylperoxide for 5 minutes as previously described (2,3). Mixed-wavelength irradiation was with a Bausch and Lomb unit equipped with a 200 W Osram Hg bulb and a Schott W G 320 filter at an incident dose of 6.6 x 10-5 einstein/cm2Imin (2,3). The fiDo wing damaged moieties were present in the photoalkylated substrate: eight 842-hydroxy-2-propyl)guanines, three 842-hydroxy-2propyl)adenines and four thymine dimers per molecule (3).…”

“…Mixed-wavelength irradiation was with a Bausch and Lomb unit equipped with a 200 W Osram Hg bulb and a Schott W G 320 filter at an incident dose of 6.6 x 10-5 einstein/cm2Imin (2,3). The fiDo wing damaged moieties were present in the photoalkylated substrate: eight 842-hydroxy-2-propyl)guanines, three 842-hydroxy-2propyl)adenines and four thymine dimers per molecule (3). Broad spectrum ultraviolet irradiation was performed under identical conditions, except that both isopropanol and di-tert-butylperoxide were absent from the reaction mixture.…”

“…Photoalkylation is the additon of alcohols to C-8 positions of purines in ultra violet-irradiated DNA (1). In addition to 8-(2-hydroxy-2-propyl)purines, pyrimidine dimers and uncharacterized adenine adducts are also formed in photoalkylated DNA (2,3). An endonuclease activity that incises photoalkylated DNA was found in crude extracts of M. luteus (4).…”