Structure of purine-purine mispairs during misincorporation and extension by Escherichia coli DNA polymerase I

Biochemistry
Angie M Kretulskie, Thomas E Spratt

Abstract

The mechanism by which purine-purine mispairs are formed and extended was examined with the high-fidelity Klenow fragment of Escherichia coli DNA polymerase I with the proofreading exonuclease activity inactivated. The structures of the purine-purine mispairs were examined by comparing the kinetics of mispair formation with adenine versus 7-deazaadenine and guanine versus 7-deazaguanine at four positions in the DNA, the incoming dNTP, the template base, and both positions of the terminal base pair. A decrease in rate associated with a 7-deazapurine substitution would suggest that the nucleotide is in a syn conformation in a Hoogsteen base pair with the opposite base. During mispair formation, the k(pol)/K(d) values for the insertion of dATP opposite A (dATP/A) as well as dATP/G and dGTP/G were decreased greater than 10-fold with the deazapurine in the dNTP. These results suggest that during mispair formation the newly forming base pair is in a Hoogsteen geometry with the incoming dNTP in the syn conformation and the template base in the anti conformation. During mispair extension, the only decrease in k(pol)/K(d) was associated with the G/G base pair in which 7-deazaguanine was in the template strand. These results as well as p...Continue Reading

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Citations

Jul 12, 2008·Biochemistry·Lindsey DeCarloThomas E Spratt
Aug 10, 2007·Molekuliarnaia biologiia·A A NeschastnovaN G Dolinnaia
Apr 4, 2014·Physical Chemistry Chemical Physics : PCCP·Ol'ha O Brovarets', Dmytro M Hovorun
Mar 13, 2014·Journal of the American Chemical Society·Wen-Jin WuMing-Daw Tsai
May 31, 2012·Angewandte Chemie·Tatsuya FunaiHidehito Urata
Oct 29, 2008·Journal of Molecular Biology·Benedetta A Sampoli BenítezTamar Schlick
Jan 10, 2017·Chemical Research in Toxicology·A S Prakasha GowdaThomas E Spratt
Dec 5, 2017·Frontiers in Molecular Biosciences·Anthony J Berdis
Oct 13, 2017·Chembiochem : a European Journal of Chemical Biology·Rania Abu El AsrarPiet Herdewijn

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