2br0
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DNA ADDUCT BYPASS POLYMERIZATION BY SULFOLOBUS SOLFATARICUS DPO4. ANALYSIS AND CRYSTAL STRUCTURES OF MULTIPLE BASE-PAIR SUBSTITUTION AND FRAMESHIFT PRODUCTS WITH THE ADDUCT 1,N2-ETHENOGUANINE
OverviewOverview
1,N(2)-Etheno(epsilon)guanine is a mutagenic DNA lesion derived from lipid, oxidation products and also from some chemical carcinogens. Gel, electrophoretic analysis of the products of primer extension by Sulfolobus, solfataricus P2 DNA polymerase IV (Dpo4) indicated preferential, incorporation of A opposite 3'-(1,N(2)-epsilon-G)TACT-5', among the four, dNTPs tested individually. With the template 3'-(1,N(2)-epsilon-G)CACT-5', both G and A were incorporated. When primer extension was done in the, presence of a mixture of all four dNTPs, high pressure liquid, chromatography-mass spectrometry analysis of the products indicated that, (opposite 3'-(1,N(2)-epsilon-G)CACT-5') the major product was 5'-GTGA-3', and the minor product was 5'-AGTGA-3'. With the template, 3'-(1,N(2)-epsilon-G)TACT-5', the following four products were identified, by high pressure liquid chromatography-mass spectrometry: 5'-AATGA-3', 5'-ATTGA-3', 5'-ATGA-3', and 5'-TGA-3'. An x-ray crystal structure of Dpo4, was solved (2.1 A) with a primer-template and A placed in the primer to be, opposite the 1,N(2)-epsilon-G in the template 3'-(1,N(2)-epsilon-G)TACT, 5'. The added A in the primer was paired across the template T with, classic Watson-Crick geometry. Similar structures were observed in a, ternary Dpo4-DNA-dATP complex and a ternary Dpo4-DNA-ddATP complex, with, d(d)ATP opposite the template T. A similar structure was observed with a, ddGTP adjacent to the primer and opposite the C next to 1,N(2)-epsilon-G, in 3'-(1,N(2)-epsilon-G)CACT-5'. We concluded that Dpo4 uses several, mechanisms, including A incorporation opposite 1,N(2)-epsilon-G and also a, variation of dNTP-stabilized misalignment, to generate both base pair and, frameshift mutations.
About this StructureAbout this Structure
2BR0 is a Single protein structure of sequence from Sulfolobus solfataricus with CA and DG3 as ligands. Active as DNA-directed DNA polymerase, with EC number 2.7.7.7 Structure known Active Site: AC1. Full crystallographic information is available from OCA.
ReferenceReference
DNA adduct bypass polymerization by Sulfolobus solfataricus DNA polymerase Dpo4: analysis and crystal structures of multiple base pair substitution and frameshift products with the adduct 1,N2-ethenoguanine., Zang H, Goodenough AK, Choi JY, Irimia A, Loukachevitch LV, Kozekov ID, Angel KC, Rizzo CJ, Egli M, Guengerich FP, J Biol Chem. 2005 Aug 19;280(33):29750-64. Epub 2005 Jun 17. PMID:15965231
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