Nuclear magnetic resonance investigation of primer--template models: formation of a pyrimidine bulge upon misincorporation.
Our previous studies have shown that misaligned structures can occur upon misincorporation of a dNTP opposite thymine templates. The formation of misaligned structures during DNA replication, if not repaired properly, can be bypassed and extended by low-fidelity polymerases and ultimately lead to mutations. In this study, the base pair structures at the replicating sites of a set of primer-template models which mimic the situation upon misincorporation of a dNTP opposite cytosine templates have been determined. High-resolution NMR structural results show that misaligned structures with a C-bulge can be formed upon incorporation of dCTP, dTTP, and dATP opposite 5'-GC, 5'-AC, and 5'-TC templates, respectively. The stabilities of misaligned structures depend on the types of terminal base pairs at the replicating sites. Together with the structural findings in thymine templates, we conclude that terminal G.C and C.G base pairs always contribute a larger stabilizing effect to the misaligned structures containing a pyrimidine bulge than terminal A.T and T.A base pairs. Misalignment and thus deletion mutation are more likely to occur if misincorporation of a nucleotide opposite a pyrimidine template can cause template slippage to form a terminal G.C or C.G base pair. Although misalignment also occurs when the newly formed terminal base pair is an A.T base pair or a T.A base pair, both misaligned and mismatched conformers coexist, which can lead to deletion and substitution mutations, respectively.
Chi
Lai Man
LM
Department of Chemistry, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong.
Lam
Sik Lok
SL
D000225:Adenine / Q000737:chemistry; D020137:Base Pair Mismatch; D020029:Base Pairing; D003596:Cytosine / Q000737:chemistry*; D017931:DNA Primers; D004261:DNA Replication; D003838:Deoxyadenine Nucleotides / Q000378:metabolism; D003845:Deoxycytosine Nucleotides / Q000378:metabolism; D006147:Guanine / Q000737:chemistry; D008957:Models, Genetic; D019906:Nuclear Magnetic Resonance, Biomolecular; D013698:Templates, Genetic; D013936:Thymidine / Q000737:chemistry; D013942:Thymine Nucleotides / Q000378:metabolism
D017931:DNA Primers; D003838:Deoxyadenine Nucleotides; D003845:Deoxycytosine Nucleotides; D013942:Thymine Nucleotides; C024107:2'-deoxycytidine 5'-triphosphate; D006147:Guanine; D003596:Cytosine; D000225:Adenine; C026600:2'-deoxyadenosine triphosphate; C024157:thymidine 5'-triphosphate; D013936:Thymidine