Pyrophosphate hydrolysis is an intrinsic and critical step of the DNA synthesis reaction

Nucleic Acids Research
Jithesh Kottur, Deepak T Nair

Abstract

DNA synthesis by DNA polymerases (dPols) is central to duplication and maintenance of the genome in all living organisms. dPols catalyze the formation of a phosphodiester bond between the incoming deoxynucleoside triphosphate and the terminal primer nucleotide with the release of a pyrophosphate (PPi) group. It is believed that formation of the phosphodiester bond is an endergonic reaction and PPi has to be hydrolyzed by accompanying pyrophosphatase enzymes to ensure that the free energy change of the DNA synthesis reaction is negative and it can proceed in the forward direction. The fact that DNA synthesis proceeds in vitro in the absence of pyrophosphatases represents a long-standing conundrum regarding the thermodynamics of the DNA synthesis reaction. Using time-resolved crystallography, we show that hydrolysis of PPi is an intrinsic and critical step of the DNA synthesis reaction catalyzed by dPols. The hydrolysis of PPi occurs after the formation of the phosphodiester bond and ensures that the DNA synthesis reaction is energetically favorable without the need for additional enzymes. Also, we observe that DNA synthesis is a two Mg2+ ion assisted stepwise associative SN2 reaction. Overall, this study provides deep temporal i...Continue Reading

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Citations

Sep 24, 2019·Nucleic Acids Research·Mary K JohnsonDeepak T Nair
May 18, 2020·FEBS Letters·Andrey do Nascimento VieiraMartina Preiner
Aug 23, 2020·Nature Communications·Nigel C BrissettAidan J Doherty
Oct 2, 2020·Proceedings of the National Academy of Sciences of the United States of America·Tyler M WeaverBret D Freudenthal
Jun 8, 2021·Chemical Science·Zi Fei XuHui Ming Ge
Jul 28, 2021·Molecular Aspects of Medicine·Sonam BhatiaDeepak T Nair
May 9, 2019·Journal of the American Chemical Society·Yao-Kai ChangMing-Daw Tsai

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