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Biomedical subjects

D Elad

Publications and source records attributed to D Elad.

At least 109 records · Page 6Linked to original sources
PubMed1979

Endonucleolytic activity directed towards 8-(2-hydroxy-2-propyl) purines in double-stranded DNA.

Photoalkylation of circular covalently closed DNA from phage PM2 with isopropyl alcohol by using a free radical photoinitiator and UV light of lambda greater than 305 nm led to the specific 8-substitution of purine moieties in the DNA, yielding 8-(2-hydroxy-2-propyl)adenine and 8-(2-hydroxy-2-propyl)guanine as the only detectable damage in the DNA. Using this specifically photoalkylated DNA as a substrate, we discovered in extracts of Micrococcus luteus an endonucleolytic activity that is directed towards 8-(2-hydroxy-2-propyl) purines in DNA. The activity is not a combination of a DNA-glycosylase and an apurinic site endonuclease. It is not inhibited by single-stranded DNA, by UV- or gamma-irradiated single-stranded DNA, or by normal or depurinated double-stranded DNA. however, gamma- or UV-(254 nm) irradiated double-stranded DNAs to inhibit the activity, hinting at the possibility of a common type of lesion in these damaged DNAs. Divalent cations are not required for the incising activity, and it is fully active in 1 mM EDTA, whereas caffeine and ATP cause inhibition. Extracts of mutant M. luteus lacking pyrimidine-dimer-directed endonucleases were found to contain the endonucleolytic activity in levels comparable to those present in the wild type. After the incision, we could demonstrate the specific excision of the 8-alkylated purines from the damaged DNA. The special conformational consequences of the 8-alkylation of purines, at the nucleotide level, namely their nonregular syn conformation, suggest that it is the distortion in the DNA that is recognized by the endonuclease.

Alkylation↗
PubMed1979

Enzymatic insertion of purine bases into depurinated DNA in vitro.

An enzymatic activity that inserts purines into depurinated DNA was found in a soluble enzyme extract of Escherichia coli. This activity brings about the insertion of adenine and guanine into the appropriate apurinic sites in double-stranded DNA by using the corresponding deoxyribonucleoside triphosphates as the purine donors. Magnesium ions are required for this activity, it is inhibited by caffeine, and it does not act on depurinated single-stranded DNA. The insertion activity described here may represent a step in a repair mechanism, "base-insertion repair," whereby apurinic sites (which may occur in double-stranded DNA either due to the removal of damaged purines with specific glycosylases or by spontaneous depurination) are directly filled with the correct missing purine base.

Caffeine↗
PubMed1978

Light-induced free-radical reactions of nucleic acid constituents. Effect of sequence and base--base interactions on the reactivity of purines and pyrimidines in ribonucleotides.

The reaction with 2-propanol of purines and pyrimidines, induced photochemically with light of lambda greater than 300 nm and di-tert-butyl peroxide as an initiator, was applied to a variety of adenosine-, guanosine-, and uridine-containing ribonucleotides in order to determine the rules which govern the reactivity of the heterocyclic bases of nucleotides. The reactivity of the purine moieties was found to depend on the conformation of the appropriate nucleotide (anti or syn) and on the site of binding of the phosphate group to the ribose moiety. Adenosine moieties (assuming an anti conformation) blocked at their 3'-hydroxyl reacted faster than those blocked at their 5'-hydroxyl. The reactivity of the guanosine moieties (tending to assume a syn conformation) was independent of the site of binding of the phosphate. The uridine moieties of the various nucleotides exhibited a wide range of reactivity. A correlation between the reactivity of the uridines and their involvement in stacking interactions with next- and second-neighboring purines could be made. Thus, the uridine moieties of U-U-U, G-U, U-G, A-U-A, and A-U-G were reactive, while those of A-U and A-U-U were unreactive. The relative reactivity of uridine moieties of nucleotides can, therefore, be used as a measure of the extent of pyrimidine-purine stacking and vice versa.

Alkylation↗
PubMed1978

Photochemical dehydrogenation of 5, 6-dihydropyrimidines.

5, 6-Dihydropyrimidines can be oxidized photochemically to the original pyrimidines with light of lambda greater than 300 nm in the presence of transition metal salts which act as sensitizers. A hydrogen atom abstraction by hydroxyl radical is suggested as the reaction mechanism.

Copper↗
PubMed1977

Light-induced free radical alkylation of polynucleotides and their enzymatic digestion.

Ultraviolet light-induced free radical alkylation with 2-propanol or D-ribose, initiated with di-tert-butyl peroxide, of poly (G), poly (U20G), and poly(A) led to the substitution of the appropriate group for the H-8 atom of the purines and addition across the 5,6-double bond of the pyrimidines. The alkylated polynucleotides were subjected to nucleolytic digestion with several nucleases. T1-RNase digestion of poly(G) irradiated with 2-propanol gave a mixture of the modified and non-modified mononucleotides. Similarly, pancreatic RNase digestion of the irradiated poly(U20G) resulted in a mixture of the appropriate mononucleotides. A T2-RNase treatment of poly(A) irradiated with 2-propanol gave the modified Ado-21:3'-P, while T2-RNase digestion of poly(A) irradiated with D-ribose led to the cyclic modified mononucleotides, in addition to the modified mononucleotides.

Alkylation↗
PubMed1976

Reactivity and selectivity in light-induced free radical reactions of 2-propanol with purine and pyrimidine mononucleotides and dinucleoside monophosphates.

Photoalkylation reactions with 2-propanol, initiated with di-tert-butyl peroxide, of a variety of purine and pyrimidine mononucleotides and dinucleoside monophosphates lead to the substitution of an alpha-hydroxyisopropyl group for the H-8 atom of adenosine and the addition of the alcohol across the 5,6-double bond of the pyrimidines. Adenosine moieties blocked at their 3'-hydroxyl group are alkylated faster than those blocked at their 5'-hydroxyl. The reactivity of the uridine moieties of 3'-UMP, 5'-UMP, and uridylyl-(3',5')-uridine is not affected by the location of the phosphate group. However, the uridine moiety of uridylyl-(3',5')-adenosine is modified faster than that of adenylyl-(3',5')-uridine. It is suggested that steric hindrance imposed by the phosphate group determines the reactivity of adenosine moieties, while base stacking involving adenosine determines the reactivity of uridine moieties. These two effects play a major role in controlling the nature and degree of the selectivity of these photoalkylation reactions for either adenosine or uridine. Cytidine has been found to be inert in these reactions.

1-Propanol↗