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Translesion DNA synthesis by human DNA polymerase eta on templates containing a pyrimidopurinone deoxyguanosine adduct, 3-(2'-deoxy-beta-d-erythro-pentofuranosyl)pyrimido-[1,2-a]purin-10(3H)-one.

Stafford JB, Eoff RL, Kozekova A, Rizzo CJ, Guengerich FP, Marnett LJ - Biochemistry (2009)

Bottom Line: M(1)dG partially blocked DNA synthesis by polymerase eta.Using steady-state kinetics, we found that insertion of dCTP was the least favored insertion product opposite the M(1)dG lesion (800-fold less efficient than opposite dG).Extension from M(1)dG.dC was equally as efficient as from control primer-templates (dG.dC). dATP insertion opposite M(1)dG was the most favored insertion product (8-fold less efficient than opposite dG), but extension from M(1)dG.dA was 20-fold less efficient than dG.dC.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, A. B. Hancock, Jr., Memorial Laboratory for Cancer Research, Center in Molecular Toxicology, Vanderbilt-Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, USA.

ABSTRACT
M(1)dG (3-(2'-deoxy-beta-d-erythro-pentofuranosyl)pyrimido[1,2-a]purin-10(3H)-one) lesions are mutagenic in bacterial and mammalian cells, leading to base substitutions (mostly M(1)dG to dT and M(1)dG to dA) and frameshift mutations. M(1)dG is produced endogenously through the reaction of peroxidation products, base propenal or malondialdehyde, with deoxyguanosine residues in DNA. The mutagenicity of M(1)dG in Escherichia coli is dependent on the SOS response, specifically the umuC and umuD gene products, suggesting that mutagenic lesion bypass occurs by the action of translesion DNA polymerases, like DNA polymerase V. Bypass of DNA lesions by translesion DNA polymerases is conserved in bacteria, yeast, and mammalian cells. The ability of recombinant human DNA polymerase eta to synthesize DNA across from M(1)dG was studied. M(1)dG partially blocked DNA synthesis by polymerase eta. Using steady-state kinetics, we found that insertion of dCTP was the least favored insertion product opposite the M(1)dG lesion (800-fold less efficient than opposite dG). Extension from M(1)dG.dC was equally as efficient as from control primer-templates (dG.dC). dATP insertion opposite M(1)dG was the most favored insertion product (8-fold less efficient than opposite dG), but extension from M(1)dG.dA was 20-fold less efficient than dG.dC. The sequences of full-length human DNA polymerase eta bypass products of M(1)dG were determined by LC-ESI/MS/MS. Bypass products contained incorporation of dA (52%) or dC (16%) opposite M(1)dG or -1 frameshifts at the lesion site (31%). Human DNA polymerase eta bypass may lead to M(1)dG to dT and frameshift but likely not M(1)dG to dA mutations during DNA replication.

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Summary of translesion bypass of M1dG by human DNA polymerase η. The major in vitro products of human DNA polymerase η bypass of M1dG lesions in DNA include insertion of dC or dA opposite the lesion and “skipping” the lesion site. If subjected to a second round of DNA replication, these bypass products would lead to correct bypass of the lesion (M1dG to dG), a G to T transversion (M1dG to dT), or a −1 frameshift mutation.
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fig7: Summary of translesion bypass of M1dG by human DNA polymerase η. The major in vitro products of human DNA polymerase η bypass of M1dG lesions in DNA include insertion of dC or dA opposite the lesion and “skipping” the lesion site. If subjected to a second round of DNA replication, these bypass products would lead to correct bypass of the lesion (M1dG to dG), a G to T transversion (M1dG to dT), or a −1 frameshift mutation.

Mentions: Our results indicate that the major products of in vitro bypass of M1dG by human pol η result from incorporation of dA opposite M1dG. This substitution would result in an M1dG to dT transversion upon a second round of DNA replication (Figure 7) and agrees with published findings of in vivo replication of M1dG containing DNA in nonreiterated sequences by mammalian and bacterial cells and replication of MDA-treated DNA in E. coli(11,13). Insertion of dA opposite M1dG by human DNA pol η is 130 times more favorable kinetically than insertion of dC opposite the lesion. Insertion of dA is also 24-fold more favorable than insertion of dT and 16-fold more favorable than insertion dG opposite M1dG.


Translesion DNA synthesis by human DNA polymerase eta on templates containing a pyrimidopurinone deoxyguanosine adduct, 3-(2'-deoxy-beta-d-erythro-pentofuranosyl)pyrimido-[1,2-a]purin-10(3H)-one.

Stafford JB, Eoff RL, Kozekova A, Rizzo CJ, Guengerich FP, Marnett LJ - Biochemistry (2009)

Summary of translesion bypass of M1dG by human DNA polymerase η. The major in vitro products of human DNA polymerase η bypass of M1dG lesions in DNA include insertion of dC or dA opposite the lesion and “skipping” the lesion site. If subjected to a second round of DNA replication, these bypass products would lead to correct bypass of the lesion (M1dG to dG), a G to T transversion (M1dG to dT), or a −1 frameshift mutation.
© Copyright Policy - open-access - ccc-price
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2651650&req=5

fig7: Summary of translesion bypass of M1dG by human DNA polymerase η. The major in vitro products of human DNA polymerase η bypass of M1dG lesions in DNA include insertion of dC or dA opposite the lesion and “skipping” the lesion site. If subjected to a second round of DNA replication, these bypass products would lead to correct bypass of the lesion (M1dG to dG), a G to T transversion (M1dG to dT), or a −1 frameshift mutation.
Mentions: Our results indicate that the major products of in vitro bypass of M1dG by human pol η result from incorporation of dA opposite M1dG. This substitution would result in an M1dG to dT transversion upon a second round of DNA replication (Figure 7) and agrees with published findings of in vivo replication of M1dG containing DNA in nonreiterated sequences by mammalian and bacterial cells and replication of MDA-treated DNA in E. coli(11,13). Insertion of dA opposite M1dG by human DNA pol η is 130 times more favorable kinetically than insertion of dC opposite the lesion. Insertion of dA is also 24-fold more favorable than insertion of dT and 16-fold more favorable than insertion dG opposite M1dG.

Bottom Line: M(1)dG partially blocked DNA synthesis by polymerase eta.Using steady-state kinetics, we found that insertion of dCTP was the least favored insertion product opposite the M(1)dG lesion (800-fold less efficient than opposite dG).Extension from M(1)dG.dC was equally as efficient as from control primer-templates (dG.dC). dATP insertion opposite M(1)dG was the most favored insertion product (8-fold less efficient than opposite dG), but extension from M(1)dG.dA was 20-fold less efficient than dG.dC.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, A. B. Hancock, Jr., Memorial Laboratory for Cancer Research, Center in Molecular Toxicology, Vanderbilt-Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, USA.

ABSTRACT
M(1)dG (3-(2'-deoxy-beta-d-erythro-pentofuranosyl)pyrimido[1,2-a]purin-10(3H)-one) lesions are mutagenic in bacterial and mammalian cells, leading to base substitutions (mostly M(1)dG to dT and M(1)dG to dA) and frameshift mutations. M(1)dG is produced endogenously through the reaction of peroxidation products, base propenal or malondialdehyde, with deoxyguanosine residues in DNA. The mutagenicity of M(1)dG in Escherichia coli is dependent on the SOS response, specifically the umuC and umuD gene products, suggesting that mutagenic lesion bypass occurs by the action of translesion DNA polymerases, like DNA polymerase V. Bypass of DNA lesions by translesion DNA polymerases is conserved in bacteria, yeast, and mammalian cells. The ability of recombinant human DNA polymerase eta to synthesize DNA across from M(1)dG was studied. M(1)dG partially blocked DNA synthesis by polymerase eta. Using steady-state kinetics, we found that insertion of dCTP was the least favored insertion product opposite the M(1)dG lesion (800-fold less efficient than opposite dG). Extension from M(1)dG.dC was equally as efficient as from control primer-templates (dG.dC). dATP insertion opposite M(1)dG was the most favored insertion product (8-fold less efficient than opposite dG), but extension from M(1)dG.dA was 20-fold less efficient than dG.dC. The sequences of full-length human DNA polymerase eta bypass products of M(1)dG were determined by LC-ESI/MS/MS. Bypass products contained incorporation of dA (52%) or dC (16%) opposite M(1)dG or -1 frameshifts at the lesion site (31%). Human DNA polymerase eta bypass may lead to M(1)dG to dT and frameshift but likely not M(1)dG to dA mutations during DNA replication.

Show MeSH
Related in: MedlinePlus