Perspective: pre-chemistry conformational changes in DNA polymerase mechanisms
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In recent papers, there has been a lively exchange concerning theories for enzyme catalysis, especially the role of protein dynamics/pre-chemistry conformational changes in the catalytic cycle of enzymes. Of particular interest is the notion that substrate-induced conformational changes that assemble the polymerase active site prior to chemistry are required for DNA synthesis and impact fidelity (i.e., substrate specificity). High-resolution crystal structures of DNA polymerase β representing intermediates of substrate complexes prior to the chemical step are available. These structures indicate that conformational adjustments in both the protein and substrates must occur to achieve the requisite geometry of the reactive participants for catalysis. We discuss computational and kinetic methods to examine possible conformational change pathways that lead from the observed crystal structure intermediates to the final structures poised for chemistry. The results, as well as kinetic data from site-directed mutagenesis studies, are consistent with models requiring pre-chemistry conformational adjustments in order to achieve high fidelity DNA synthesis. Thus, substrate-induced conformational changes that assemble the polymerase active site prior to chemistry contribute to DNA synthesis even when they do not represent actual rate-determining steps for chemistry.
KeywordsEnzyme catalysis Intrinsic protein dynamics Pre-chemistry conformational adjustments Nucleotidyl transfer DNA polymerase β Catalytic cycle chemical step
Research described in this Article was supported in part by Philip Morris USA Inc. and Philip Morris International and by NSF award MCB-0316771, and NIH award R01 ES012692 to T.S., and Research Project Numbers Z01-ES050158 and Z01-ES050161 to S.H.W. in the Intramural Research Program of the National Institutes of Health, National Institute of Environmental Health Sciences and was in association with NIH award 1U19CA105010.
- 2.Kamerlin SCL, Warshel A (2010) At the dawn of the 21st century: is dynamics the missing link for understanding enzyme catalysis? Proteins 78:1339Google Scholar
- 34.Foley MC, Arora K, Schlick T (2012) Intrinsic motions of DNA polymerases underlie their remarkable specificity and selectivity and suggest a hybrid substrate binding mechanism. In: Schlick T (ed) Innovations in bimolecular modeling and simulations, vol 2., Royal Society of ChemistryLondon, UK, pp 81–110CrossRefGoogle Scholar
- 35.Arora K, Brooks III CL, Schlick T (2012) Computational support for hybrid induced fit and conformational selection mechanisms. In preparationGoogle Scholar
- 54.Zhang H, Cao W, Zakharova E, Konigsberg W, De La Cruz EM (2007) Fluorescence of 2-aminopurine reveals rapid conformational changes in the RB69 DNA polymerase-primer/template complexes upon binding and incorporation of matched deoxynucleoside triphosphates. Nucl Acids Res 35:6052CrossRefGoogle Scholar