Apyrimidinic sites

Recently, Sage and Haseltine (49) have quantitatively determined the spectrum of DNA lesions induced by reactions of BPDE with DNA. They found that alkali-labile lesions account for about 40% of the DNA adducts. There was a striking correlation between the mutation frequencies induced by BPDE in lacl and the frequencies of alkali sensitive lesions at G, A, and C residues. Apurinic/apyrimidinic sites are common alkali-sensitive lesions. Earlier work by Drink-water e al. (48) had also shown that treatment of DNA with BPDE generated apurinic/apyrimidinic sites. 

Lindahl, T. (1979). DNA glycoslylases, endonucleases for apurinic/apyrimidinic sites and base excision repair. Proc. Nuc. Acid Res. Mol. Biol. 22 109-118. 

Deoxyribonuclease V [EC 3.1.22.3], also known as endodeoxyribonuclease V, catalyzes the endonucleolytic cleavage of DNA at apurinic or apyrimidinic sites to products with a 3 -phosphate. 

DNA-(apurinic or apyrimidinic site) lyase [EC 4.2.99.18, formerly EC 3.1.25.2] acts on the C-Q-P bond 3 to the apurinic or apyrimidinic site in DNA. This bond is broken by a /3-elimination reaction, leaving a 3 -terminal unsaturated sugar and a product with a terminal 5 -phosphate. Note that this nicking of the phosphodiester bond is a lyase-type reaction, not hydrolysis. 

The production of apurinic and apyrimidinic sites can both be caused by alkylating agents. When, for example, the N7 of guanine is alkylated, this causes the bond from base to sugar to become labile and so the base can be lost completely. Thus, apurinic sites are formed. Insertion of an incorrect base at this site can then cause a mutation. Loss of purines and pyrimidines can also occur spontaneously. 

A glycosylase enzyme recognizes and removes the damaged base to give an apurinic or apyrimidinic site. 

Apurinic/apyrimidinic site production in SV40 DNA in vitro + NT 2.0 Drinkwater ei a/. (1980)... 

Drinkwater, N.R.. Miller. E.C. Miller, J.A. (1980) Estimation of apurinic/apyrimidinic sites and phosphotriesters in deoxyribonucleic acid treated with electrophilic carcinogens and mutagens. Biochemistry, 19, 5087-5092... 

Base-Excision Repair Every cell has a class of enzymes called DNA glycosylases that recognize particularly common DNA lesions (such as the products of cytosine and adenine deamination see Fig. 8-33a) and remove the affected base by cleaving the Af-glycosyl bond. This cleavage creates an apurinic or apyrimidinic site in the DNA, commonly referred to as an AP site or abasic... 

Removal of abnormal bases Abnormal bases, such as uracil, which can occur in DNA either by deamination of cytosine or improper incorporation of dUTP instead of dTTP during DNA synthesis, are recognized by specific glycosylases that hydrolytically cleave them from the deoxyribose-phosphate backbone of the strand. This leaves an apyrimidinic site (or apurinic, if a purine was removed), referred to as an AP-site. 

As the word suggests, an abasic site (apurinic/apyrimidinic site, AP) lacks the base, but the DNA backbone is more or less intact. The simplest AP is a 2-dR site, formed... 

Povirk LF, Goldberg IH (1985a) Detection of neocarzinostatin chromophore-deoxyribose adducts as exonuclease-resistant sites in defined-sequence DNA. Biochemistry 24 4035-4040 Povirk LF, Goldberg IH (1985b) Endonuclease-resistant apyrimidinic sites formed by neocarzinostatin at cytosine residues in DNA evidence for a possible role in mutagenesis. Proc Natl Acad Sci USA 82 3182-3186... 

Apurinic/apyrimidinic site containing a 2-deoxyribonolactone unit Double-stranded Double-strand break Double-stranded DNA... 

Fresco P, Shacker F, Kortenkamp A. 1995. The reductive conversion of chromium (VI) by ascorbate gives rise to apurinic/apyrimidinic sites in isolated DNA. Chem Res Toxicol 8 884-890. 

The removal of an altered base by the glycosylase results in the formation of an abasic site (AP site) (sometimes also referred to as an apurinic or apyrimidinic site). The AP site is acted upon by an AP endonuclease that makes an incision in the damaged strand by hydrolyzing the phosphodiester bond 5 to the AP site (Figure 23.5). Alternatively, some glycosylases have an associated lyase activity that incises... 

The catalytic action of these glycosylases results in formation of a nucleotide that has lost its base. The result is an apyrimidinic site (AF site). The AP site consists only of a nesidueofdwxyribose-phosphate. What happens next in the DNArepair pathway The deoxyribosc-phosphate is subsequently snipped out of the strand of DNA by an enzyme called etidottuclease. The full name for the enzyme is AP endonuclease (APE). The action of AP endonuclease results in a small gap in the double-stranded DMA helix, which is then filled in by DNA polymerase to yield repaired DNA. 

Sagher, D. and Strauss, B. (1983) Insertion of nucleotides opposite apurinic/apyrimidinic sites in deoxyribonucleic acid during in vitro synthesis. Uniqueness of adenine nucleotides. Biochemistry, 22,4518-4526. 

P.B., Chiang, S.Y., Row, Y.W., and Swenberg, J.A. (1998) Highly sensitive apurinic/apyrimidinic site assay can detect spontaneous and chemically induced depurination under physiological conditions. Cancer Res., 58, 222-225. 

R. D. (2005) DDB1-DDB2 (xeroderma pigmentosum group E) protein complex recognizes a cyclobutane pyrimidine dimer, mismatches, apurinic/ apyrimidinic sites, and compound lesions in DNA. J. Biol. Chem., 280, 39982-39989. 

P.B., Chiang, S.Y., Row, Y.W., and Swenberg, J.A. (1998) Highly sensitive apurinic/apyrimidinic site assay can detect spontaneous and chemically induced depurination under physiological conditions. Cancer Res., 58, 222-225. Loeb, L.A. and Preston, B.D. (1986) Mutagenesis by apurinic/apyrimidinic sites. Annu. Rev. Genet., 20, 201-230. Xue, L. and Greenberg, M.M. (2007)... 

Kokoska, R.J., McCulloch, S.D., and Kunkel, T.A. (2003) The efficiency and specificity of apurinic/apyrimidinic site bypass by human DNA polymerase r and Sulfolobus solfataricus Dpo4. J. Biol. Chem., 278, 50537-50545. 

Figure 16.4 DNA lesions used in TLS studies with gapped plasmids. BP-G, B[o]P-guanine adduct AP site, apurinic/apyrimidinic site 4-OHEN-C, 4-hydroxyequilenin-C adduct M12, dodecamethylene Cisplatin-GG, c/s-diamminedichloroplatinum(ll)-GG intrastrand crosslink ...

---