Structure and function relationships in mammalian DNA polymerases

Hoitsma, N.M.; Whitaker, A.M.; Schaich, Matthew A; Smith, M.R.; Fairlamb, Max S.; Freudenthal, Bret

doi:10.1007/s00018-019-03368-y

Cited by 27 publications

(25 citation statements)

References 292 publications

(347 reference statements)

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“…DNA polymerases bind the DNA and incoming nucleotide substrates and arrange them such that the 3′-oxygen of the primer termini and the α-phosphate of the incoming nucleotide are properly oriented for the nucleotidyl transfer reaction (1)(2)(3)(8)(9)(10). This occurs in two sequential steps: DNA binding and subsequent dNTP binding.…”

Section: Organization Of Substrates and Products Within The Rev1 Active Sitementioning

confidence: 99%

“…In the case of many DNA polymerases, the nucleotide-binding step couples the association of the nucleotide and metal ions to conformational changes in the polymerase (1)(2)(3)(8)(9)(10). Typically, this involves an open-to-closed transition of the finger subdomain around the incoming nucleotide and the template base.…”

Section: Organization Of Substrates and Products Within The Rev1 Active Sitementioning

confidence: 99%

“…In the product complex of other DNA polymerases, the incorporated nucleotide is base-paired with the templating base (1)(2)(3)(8)(9)(10). The base-paired product is then released from the active site through reopening of the finger subdomain of the polymerase.…”

Section: Organization Of Substrates and Products Within The Rev1 Active Sitementioning

confidence: 99%

“…DNA polymerase | DNA repair | translesion synthesis C ells are tasked with efficiently and faithfully replicating the genome in each round of cell division (1)(2)(3). During replication, the replicative DNA polymerases encounter a variety of DNA lesions that can stall the replication fork, promoting genomic instability (4,5).…”

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confidence: 99%

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Visualizing Rev1 catalyze protein-template DNA synthesis

Weaver

Cortez

Khoang

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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During DNA replication, replicative DNA polymerases may encounter DNA lesions, which can stall replication forks. One way to prevent replication fork stalling is through the recruitment of specialized translesion synthesis (TLS) polymerases that have evolved to incorporate nucleotides opposite DNA lesions. Rev1 is a specialized TLS polymerase that bypasses abasic sites, as well as minor-groove and exocyclic guanine adducts. Lesion bypass is accomplished using a unique protein-template mechanism in which the templating base is evicted from the DNA helix and the incoming dCTP hydrogen bonds with an arginine side chain of Rev1. To understand the protein-template mechanism at an atomic level, we employed a combination of time-lapse X-ray crystallography, molecular dynamics simulations, and DNA enzymology on the Saccharomyces cerevisiae Rev1 protein. We find that Rev1 evicts the templating base from the DNA helix prior to binding the incoming nucleotide. Binding the incoming nucleotide changes the conformation of the DNA substrate to orient it for nucleotidyl transfer, although this is not coupled to large structural changes in Rev1 like those observed with other DNA polymerases. Moreover, we found that following nucleotide incorporation, Rev1 converts the pyrophosphate product to two monophosphates, which drives the reaction in the forward direction and prevents pyrophosphorolysis. Following nucleotide incorporation, the hydrogen bonds between the incorporated nucleotide and the arginine side chain are broken, but the templating base remains extrahelical. These postcatalytic changes prevent potentially mutagenic processive synthesis by Rev1 and facilitate dissociation of the DNA product from the enzyme.

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Section: Organization Of Substrates and Products Within The Rev1 Active Sitementioning

confidence: 99%

Section: Organization Of Substrates and Products Within The Rev1 Active Sitementioning

confidence: 99%

Section: Organization Of Substrates and Products Within The Rev1 Active Sitementioning

confidence: 99%

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confidence: 99%

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Visualizing Rev1 catalyze protein-template DNA synthesis

Weaver

Cortez

Khoang

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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“…DNA primer enzyme encoded by ORF35 has 99 % homology with Xanthomonas phage XAA_VB_Phi31, which can catalyze the synthesis of RNA primer. Different DNA polymerases play different roles in DNA replication[23]. The DNA polymerase encoded by ORF30 is mainly used for DNA replication and repair and exhibited 99 % identity to the S.maltophilia phage Ponderosa.…”

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confidence: 99%

Biological Characteristics And Genomic Analysis of A Novel Bacteriophage BUCT609 Infecting Stenotrophomonas Maltophilia

Han

Fan

et al. 2021

Preprint

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Stenotrophomonas maltophilia is widely distributed in nature and has a high isolation rate in nosocomial infections. Due to its potential application and important role in clinical practice, the relevant studies of S.maltophilia have received much attention. S.maltophilia phage BUCT609 (GenBank: MW960043) was isolated from hospital sewage with S.maltophilia strain No. 3015 as a host. Morphologically, it can be inferred as Podoviridae phage from the result of transmission electron microscopy (TEM). The electron microscopy also shows that the phage has an isometric capsid ~50 nm in diameter. The one-step growth curve demonstrated that the incubation period of 10 min and the burst size is 382 pfu/cell when its optimal multiplicity of infection (MOI) is 0.01. Phage BUCT609 had a high survival rate at pH 3 to 10 and tolerant temperature from 4 ℃ to 55 ℃. Next-Generation Sequencing (NGS) results demonstrated that its complete genome is linear double-stranded DNA of 43145 bp in length, and the GC content is 58 %. It has very little resemblance to other phages. The BlastN analysis shows that the genome of phage BUCT609 shares 22 % homology with S.maltophilia phage Ponderosa (GenBank: MK903280.1), and it encodes 56 putative proteins, of which only 25 have annotated function. Phage BUCT609 with a relatively large burst size and excellent survival ability in a wide pH and temperature range, suggests BUCT609 is a potential alternative for multi-drug resistance S.maltophilia therapy.

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Computational investigations of polymerase enzymes: Structure, function, inhibition, and biotechnology

Geronimo

Vidossich

Donati

et al. 2021

WIREs Comput Mol Sci

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DNA and RNA polymerases (Pols) are central to life, health, and biotechnology because they allow the flow of genetic information in biological systems. Importantly, Pol function and (de)regulation are linked to human diseases, notably cancer (DNA Pols) and viral infections (RNA Pols) such as COVID‐19. In addition, Pols are used in various applications such as synthesis of artificial genetic polymers and DNA amplification in molecular biology, medicine, and forensic analysis. Because of all of this, the field of Pols is an intense research area, in which computational studies contribute to elucidating experimentally inaccessible atomistic details of Pol function. In detail, Pols catalyze the replication, transcription, and repair of nucleic acids through the addition, via a nucleotidyl transfer reaction, of a nucleotide to the 3′‐end of the growing nucleic acid strand. Here, we analyze how computational methods, including force‐field‐based molecular dynamics, quantum mechanics/molecular mechanics, and free energy simulations, have advanced our understanding of Pols. We examine the complex interaction of chemical and physical events during Pol catalysis, like metal‐aided enzymatic reactions for nucleotide addition and large conformational rearrangements for substrate selection and binding. We also discuss the role of computational approaches in understanding the origin of Pol fidelity—the ability of Pols to incorporate the correct nucleotide that forms a Watson–Crick base pair with the base of the template nucleic acid strand. Finally, we explore how computations can accelerate the discovery of Pol‐targeting drugs and engineering of artificial Pols for synthetic and biotechnological applications. This article is categorized under: Structure and Mechanism > Reaction Mechanisms and Catalysis Structure and Mechanism > Computational Biochemistry and Biophysics Software > Molecular Modeling

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Structure and function relationships in mammalian DNA polymerases

Cited by 27 publications

References 292 publications

Visualizing Rev1 catalyze protein-template DNA synthesis

Visualizing Rev1 catalyze protein-template DNA synthesis

Biological Characteristics And Genomic Analysis of A Novel Bacteriophage BUCT609 Infecting Stenotrophomonas Maltophilia

Computational investigations of polymerase enzymes: Structure, function, inhibition, and biotechnology

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